Infusions of muscimol into the lateral septum do not reduce rats' defensive behaviors toward a cat odor stimulus.

PubMed

Chee, San-San A; Patel, Ronak; Menard, Janet L

2015-01-01

The lateral septum (LS) is implicated in behavioral defense. We tested whether bilateral infusions of the GABAA receptor agonist muscimol into the LS suppress rats' defensive responses to cat odor. Rats received intra-LS infusions of either saline or muscimol (40 ng/rat) and were exposed to either a piece of a cat collar that had been previously worn by a cat or to a control (cat odor free) collar. Rats exposed to the cat odor collar displayed more head-out postures, while intra-LS application of muscimol reduced the number of head-out postures. However, this reduction was also present in rats exposed to a control (cat odor free) collar. This latter finding suggests that despite its involvement in other defensive behaviors (e.g., open arm avoidance in the elevated plus maze), the LS does not selectively regulate rats' receptor defensive responding to the olfactory cues present in our cat odor stimulus. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

GABA(A) receptors in the pontine reticular formation of C57BL/6J mouse modulate neurochemical, electrographic, and behavioral phenotypes of wakefulness.

PubMed

Flint, RaShonda R; Chang, Theresa; Lydic, Ralph; Baghdoyan, Helen A

2010-09-15

Drugs that potentiate transmission at GABA(A) receptors are widely used to enhance sleep and to cause general anesthesia. The mechanisms underlying these effects are unknown. This study tested the hypothesis that GABA(A) receptors in the pontine reticular nucleus, oral part (PnO) of mouse modulate five phenotypes of arousal: sleep and wakefulness, cortical electroencephalogram (EEG) activity, acetylcholine (ACh) release in the PnO, breathing, and recovery time from general anesthesia. Microinjections into the PnO of saline (vehicle control), the GABA(A) receptor agonist muscimol, muscimol with the GABA(A) receptor antagonist bicuculline, and bicuculline alone were performed in male C57BL/6J mice (n = 33) implanted with EEG recording electrodes. Muscimol caused a significant increase in wakefulness and decrease in rapid eye movement (REM) and non-REM (NREM) sleep. These effects were reversed by coadministration of bicuculline. Bicuculline administered alone caused a significant decrease in wakefulness and increase in NREM sleep and REM sleep. Muscimol significantly increased EEG power in the delta range (0.5-4 Hz) during wakefulness and in the theta range (4-9 Hz) during REM sleep. Dialysis delivery of bicuculline to the PnO of male mice (n = 18) anesthetized with isoflurane significantly increased ACh release in the PnO, decreased breathing rate, and increased anesthesia recovery time. All drug effects were concentration dependent. The effects on phenotypes of arousal support the conclusion that GABA(A) receptors in the PnO promote wakefulness and suggest that increasing GABAergic transmission in the PnO may be one mechanism underlying the phenomenon of paradoxical behavioral activation by some benzodiazepines.

Involvement of the CA1 GABAA receptors in MK-801-induced anxiolytic-like effects: an isobologram analysis.

PubMed

Naseri, Mohammad-Hasan; Hesami-Tackallou, Saeed; Torabi-Nami, Mohammad; Zarrindast, Mohammad-Reza; Nasehi, Mohammad

2014-06-01

There seems to be a close relationship between hippocampal N-methyl-D-aspartic acid (NMDA) and GABAA receptors with respect to the modulation of behavior that occurs in the CA1 region of the hippocampus. This study investigated the possible involvement of the CA1 GABAA receptors in anxiolytic-like effects induced by (+)-MK-801 (a noncompetitive antagonist of the NMDA subtype of the glutamate receptor). Male Wistar rats were subjected to the elevated plus-maze apparatus and open arm time (%OAT), and open arm entries (%OAE) for anxiety-related behaviors, and closed arm entries that correspond to the locomotor activity were assessed. An intra-CA1 injection of (+)-MK-801 (2 μg/rat) and muscimol (0.5 μg/rat; a GABAA receptor agonist) increased %OAT and %OAE by themselves while not altering the closed arm entries, indicating an anxiolytic-like effect of these drugs. Injection of bicuculline (0.1, 0.25, and 0.5 μg/rat; a GABAA receptor antagonist) did not alter any of the anxiety-related parameters. An intra-CA1 injection of a subthreshold dose of muscimol (0.1 μg/rat) or bicuculline (0.5 μg/rat), 5 min before injection of subthreshold and effective doses of (+)-MK-801 (0.5, 1 and 2 μg/rat), increased and decreased the anxiolytic-like effect of (+)-MK-801, respectively. The isobologram analysis of these findings suggested a synergistic anxiety-like effect of intra-CA1 (+)-MK-801 and muscimol. In conclusion, the CA1 GABAA receptors appear to be involved in anxiolytic-like behaviors induced by (+)-MK-801.

Role of the amygdala GABA-A receptors in ACPA-induced deficits during conditioned fear learning.

PubMed

Nasehi, Mohammad; Roghani, Farnaz; Ebrahimi-Ghiri, Mohaddeseh; Zarrindast, Mohammad-Reza

2017-05-01

The basolateral amygdala (BLA) is a key structure for the emotional processing and storage of memories associated with emotional events, especially fear. On the other hand, endocannabinoids and CB1 receptors play a key role in learning and memory partly through long-term synaptic depression of GABAergic synapses in the BLA. The aim of this study was to explore the effects of GABA-A receptor agonist and antagonist in the fear-related memory acquisition deficits induced by ACPA (a selective CB1 cannabinoid receptor agonist). This study used context and tone fear conditioning paradigms to assess fear-related memory in male NMRI mice. Our results showed that the pre-training intraperitoneal administration of ACPA (0.5mg/kg) or (0.1 and 0.5mg/kg) decreased the percentage of freezing time in the contextual and tone fear conditioning, respectively. This indicated an impaired context- or tone-dependent fear memory acquisition. Moreover, the pre-training intra-BLA microinjection of GABA-A receptor agonist, muscimol, at 0.05 and 0.5μg/mouse impaired context-dependent fear memory, while the same doses of GABA-A antagonist, bicuculline, impaired tone-dependent fear memory. However, a subthreshold dose of muscimol or bicuculline increased the effect of ACPA at 0.1 and 0.5 or 0.05mg/kg on context- or tone-dependent fear memory, respectively. In addition, bicuculline at the lower dose increased the ACPA response on locomotor activity compared to its respective group. Such findings highlighted an interaction between BLA GABAergic and cannabinoidergic systems during the acquisition phase of conditioned fear memories. Copyright © 2017 Elsevier Inc. All rights reserved.

Muscimol microinjected in the arcuate nucleus affects metabolism, body temperature & ventilation.

PubMed

Schlenker, Evelyn H

2016-06-15

Effects of microinjection of 2 doses of γ-aminobutyric acid (GABA)A receptor agonist, muscimol (M), into the hypothalamic arcuate nucleus on oxygen consumption and control of ventilation over time and body temperature (BT) at the end of the experiment were compared in adult male and female rats. Relative to cerebrospinal fluid (CSF, 0 nmol), BT was decreased only in male rats with both doses of M, while in female rats, the 5 nmol dose depressed oxygen consumption. Ventilation was depressed by 5 nmol M in male and 10 nmol M in female rats by decreasing tidal volume. M did not affect the ventilatory response of male or female rats to hypoxia, whereas in females 5 and 10 nmol M and in males 10 nmol M depressed the ventilatory response to hypercapnia. Thus, in rats GABAA receptors in the arcuate nucleus modulate BT, oxygen consumption, and ventilation in air and in response to hypercapnia in a sexually dimorphic manner. Copyright © 2016 Elsevier B.V. All rights reserved.

Temporary inhibition of dorsal or ventral hippocampus by muscimol: distinct effects on measures of innate anxiety on the elevated plus maze, but similar disruption of contextual fear conditioning.

PubMed

Zhang, Wei-Ning; Bast, Tobias; Xu, Yan; Feldon, Joram

2014-04-01

Studies in rats, involving hippocampal lesions and hippocampal drug infusions, have implicated the hippocampus in the modulation of anxiety-related behaviors and conditioned fear. The ventral hippocampus is considered to be more important for anxiety- and fear-related behaviors than the dorsal hippocampus. In the present study, we compared the role of dorsal and ventral hippocampus in innate anxiety and classical fear conditioning in Wistar rats, examining the effects of temporary pharmacological inhibition by the GABA-A agonist muscimol (0.5 ug/0.5 ul/side) in the elevated plus maze and on fear conditioning to a tone and the conditioning context. In the elevated plus maze, dorsal and ventral hippocampal muscimol caused distinct behavioral changes. The effects of ventral hippocampal muscimol were consistent with suppression of locomotion, possibly accompanied by anxiolytic effects, whereas the pattern of changes caused by dorsal hippocampal muscimol was consistent with anxiogenic effects. In contrast, dorsal and ventral hippocampal muscimol caused similar effects in the fear conditioning experiments, disrupting contextual, but not tone, fear conditioning. Copyright © 2013 Elsevier B.V. All rights reserved.

Activation of single heteromeric GABAA receptor ion channels by full and partial agonists

PubMed Central

Mortensen, Martin; Kristiansen, Uffe; Ebert, Bjarke; Frølund, Bente; Krogsgaard-Larsen, Povl; Smart, Trevor G

2004-01-01

The linkage between agonist binding and the activation of a GABAA receptor ion channel is yet to be resolved. This aspect was examined on human recombinant α1β2γ2S GABAA receptors expressed in human embryonic kidney cells using the following series of receptor agonists: GABA, isoguvacine, 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP), isonipecotic acid, piperidine-4-sulphonic acid (P4S), imidazole-4-acetic acid (IAA), 5-(4-piperidyl)-3-isothiazolol (thio-4-PIOL) and 5-(4-piperidyl)-3-isoxazolol (4-PIOL). Whole-cell concentration–response curves enabled the agonists to be categorized into four classes based upon their maximum responses. Single channel analyses revealed that the channel conductance of 25–27 pS was unaffected by the agonists. However, two open states were resolved from the open period distributions with mean open times reduced 5-fold by the weakest partial agonists. Using saturating agonist concentrations, estimates of the channel shutting rate, α, ranged from 200 to 600 s−1. The shut period distributions were described by three or four components and for the weakest partial agonists, the interburst shut periods increased whilst the mean burst durations and longest burst lengths were reduced relative to the full agonists. From the burst analyses, the opening rates for channel activation, β, and the total dissociation rates, k−1, for the agonists leaving the receptor were estimated. The agonist efficacies were larger for the full agonists (E ∼7−9) compared to the weak partial agonists (∼0.4–0.6). Overall, changes in agonist efficacy largely determined the different agonist profiles with contributions from the agonist affinities and the degree of receptor desensitization. From this we conclude that GABAA receptor activation does not occur in a switch-like manner since the agonist recognition sites are flexible, accommodating diverse agonist structures which differentially influence the opening and shutting rates of the ion

Developmental changes in expression of GABAA receptor-channels in rat intrinsic cardiac ganglion neurones

PubMed Central

Fischer, Harald; Harper, Alexander A; Anderson, Colin R; Adams, David J

2005-01-01

The effects of γ-aminobutyric acid (GABA) on the electrophysiological properties of intracardiac neurones were investigated in the intracardiac ganglion plexus in situ and in dissociated neurones from neonatal, juvenile and adult rat hearts. Focal application of GABA evoked a depolarizing, excitatory response in both intact and dissociated intracardiac ganglion neurones. Under voltage clamp, both GABA and muscimol elicited inward currents at −60 mV in a concentration-dependent manner. The fast, desensitizing currents were mimicked by the GABAA receptor agonists muscimol and taurine, and inhibited by the GABAA receptor antagonists, bicuculline and picrotoxin. The GABAA0 antagonist (1,2,5,6-tetrahydropyridin-4-yl)methyl phosphonic acid (TPMPA), had no effect on GABA-induced currents, suggesting that GABAA receptor-channels mediate the response. The GABA-evoked current amplitude recorded from dissociated neurones was age dependent whereby the peak current density measured at −100 mV was ∼ 20 times higher for intracardiac neurones obtained from neonatal rats (P2–5) compared with adult rats (P45–49). The decrease in GABA sensitivity occurred during the first two postnatal weeks and coincides with maturation of the sympathetic innervation of the rat heart. Immunohistochemical staining using antibodies against GABA demonstrate the presence of GABA in the intracardiac ganglion plexus of the neonatal rat heart. Taken together, these results suggest that GABA and taurine may act as modulators of neurotransmission and cardiac function in the developing mammalian intrinsic cardiac nervous system. PMID:15731187

Activation of GABA-A receptors during postnatal brain development increases anxiety- and depression-related behaviors in a time- and dose-dependent manner in adult mice.

PubMed

Salari, Ali-Akbar; Bakhtiari, Amir; Homberg, Judith R

2015-08-01

Disturbances of the gamma-amino butyric acid-ergic (GABAergic) system during postnatal development can have long-lasting consequences for later life behavior, like the individual's response to stress. However, it is unclear which postnatal windows of sensitivity to GABA-ergic modulations are associated with what later-life behavioral outcomes. Therefore, we sought to determine whether neonatal activation of the GABA-A receptor during two postnatal periods, an early window (postnatal day 3-5) and a late window (postnatal day 14-16), can affect anxiety- and depression-related behaviors in male mice in later life. To this end, mice were treated with either saline or muscimol (50, 100, 200, 300 and 500μg/kg) during the early and late postnatal periods. An additional group of mice was treated with the GABA-A receptor antagonist bicuculline+muscimol. When grown to adulthood male mice were exposed to behavioral tests to measure anxiety- and depression-related behaviors. Baseline and stress-induced corticosterone (CORT) levels were also measured. The results indicate that early postnatal and to a lesser extent later postnatal exposure to the GABA-A receptor agonist muscimol increased anxiety-like behavior and stress-induced CORT levels in adults. Moreover, the early postnatal treatment with muscimol increased depression-like behavior with increasing baseline CORT levels. The anxiogenic and depression-like later-life consequences could be antagonized by bicuculline. Our findings suggest that GABA-A receptor signaling during early-life can influence anxiety- and depression-related behaviors in a time- and dose-dependent manner in later life. Our findings help to increase insight in the developmental mechanisms contributing to stress-related disorders. Copyright © 2015 Elsevier B.V. and ECNP. All rights reserved.

Evidence for motivational effects elicited by activation of GABA-A or dopamine receptors in the nucleus accumbens shell

PubMed Central

Wirtshafter, David; Stratford, Thomas R.

2011-01-01

Microinjections of the inhibitory GABA-A receptor agonist muscimol into the shell region of the nucleus accumbens (AcbSh) have been reported to induce large increases in food intake, but the effect of these injections on motivational processes is less clear. In the current study, bilateral injections of saline, muscimol (50 ng/side) or D-amphetamine (10 μg/side) were made into the AcbSh of rats trained to lever press on a progressive ratio schedule for food reward. Injections of both muscimol and amphetamine were found to produce a large increase in the breaking point relative to saline injections. This result suggests that inactivation of the AcbSh does not simply drive ingestive behavior, but also affects motivational processes assessed by the progressive ratio schedule. Breaking points were also increased by injections of amphetamine into the AcbSh. PMID:20598739

Role of GABAA receptors in dorsal raphe nucleus in stress-induced reinstatement of morphine-conditioned place preference in rats.

PubMed

Li, Chen; Staub, Daniel R; Kirby, Lynn G

2013-12-01

The serotonin (5-hydroxytryptamine, 5-HT) system plays an important role in stress-related psychiatric disorders and substance abuse. Our data indicate that stress inhibits the dorsal raphe nucleus (DRN)-5-HT system via stimulation of GABA synaptic activity by the stress neurohormone corticotropin-releasing factor and, more recently, that morphine history sensitizes DRN-5-HT neurons to GABAergic inhibitory effects of stress. We tested the hypothesis that DRN GABAA receptors contribute to stress-induced reinstatement of morphine-conditioned place preference (CPP). First, we tested if activation of GABAA receptors in the DRN would reinstate morphine CPP. Second, we tested if blockade of GABAA receptors in the DRN would attenuate swim stress-induced reinstatement of morphine CPP. CPP was induced by morphine (5 mg/kg) in a 4-day conditioning phase followed by a conditioning test. Upon acquiring conditioning criteria, subjects underwent 4 days of extinction training followed by an extinction test. Upon acquiring extinction criteria, animals underwent a reinstatement test. For the first experiment, the GABAA receptor agonist muscimol (50 ng) or vehicle was injected into the DRN prior to the reinstatement test. For the second experiment, the GABAA receptor antagonist bicuculline (75 ng) or vehicle was injected into the DRN prior to a forced swim stress, and then, animals were tested for reinstatement of CPP. Intraraphe injection of muscimol reinstated morphine CPP, while intraraphe injection of bicuculline attenuated swim stress-induced reinstatement. These data provide evidence that GABAA receptor-mediated inhibition of the serotonergic DRN contributes to stress-induced reinstatement of morphine CPP.

GABAergic control of food intake in the meat-type chickens.

PubMed

Jonaidi, H; Babapour, V; Denbow, D M

2002-08-01

This study examined the effects of intracerebroventricular injections of gamma-aminobutyric acid (GABA) agonists on short-term food intake in meat-type cockerels. In Experiment 1, birds were injected with various doses of muscimol, a GABA(A) agonist. In Experiment 2, the birds received bicuculline, a GABA(A) antagonist, prior to injection of muscimol. In Experiment 3, the effect of varying doses of baclofen, a GABA(B) agonist, on food intake was determined. The intracerebroventricular injection of muscimol caused a dose-dependent increase in food intake. This effect was significantly attenuated by pretreatment with bicuculline. Food intake was not affected by the intracerebroventricular injection of baclofen. These results suggest that GABA acts within the brain of broilers at a GABA(A), but not GABA(B), receptor to increase voluntary food intake.

Evidence for motivational effects elicited by activation of GABA-A or dopamine receptors in the nucleus accumbens shell.

PubMed

Wirtshafter, David; Stratford, Thomas R

2010-09-01

Microinjections of the inhibitory GABA-A receptor agonist muscimol into the shell region of the nucleus accumbens (AcbSh) have been reported to induce large increases in food intake, but the effect of these injections on motivational processes is less clear. In the current study, bilateral injections of saline, muscimol (50ng/side) or d-amphetamine (10mug/side) were made into the AcbSh of rats trained to lever press on a progressive ratio schedule for food reward. Injections of both muscimol and amphetamine were found to produce a large increase in the breaking point relative to saline injections. This result suggests that inactivation of the AcbSh does not simply drive ingestive behavior, but also affects motivational processes assessed by the progressive ratio schedule. Breaking points were also increased by injections of amphetamine into the AcbSh. Copyright 2010 Elsevier Inc. All rights reserved.

Alterations in food intake elicited by GABA and opioid agonists and antagonists administered into the ventral tegmental area region of rats.

PubMed

Echo, Joyce A; Lamonte, Nicole; Ackerman, Tsippa F; Bodnar, Richard J

2002-05-01

Food intake is significantly increased following administration of mu-selective opioid agonists into the ventral tegmental area (VTA) region acting through multiple local opioid receptor subtypes. Since GABA receptor agonists in the VTA region are capable of eliciting feeding, the present study investigated whether feeding elicited by the mu-selective opioid agonist [D-Ala(2), NMe(4), Gly-ol(5)]-enkephalin (DAMGO) in the VTA region was altered by pretreatment into the same site with equimolar doses of either GABA(A) (bicuculline) or GABA(B) (saclofen) antagonists, and further, whether pretreatment with either general opioid or selective GABA receptor antagonists decreased feeding elicited by GABA(A) (muscimol) or GABA(B) (baclofen) agonists in the VTA region. DAMGO-induced feeding in the VTA region was dose-dependently decreased following pretreatment with either GABA(A) or GABA(B) antagonists in the absence of significant alterations in food intake by the antagonists per se. However, the presence of short-lived seizures following bicuculline in the VTA region suggests that this ingestive effect was caused by nonspecific actions. In contrast, GABA(B) receptors are involved in the full expression of mu-opioid agonist-induced feeding in this region since saclofen failed to elicit either seizure activity or a conditioned taste aversion. Pretreatment with naltrexone in the VTA region reduced intake elicited by baclofen, but not muscimol. Finally, baclofen-induced feeding was significantly reduced by saclofen, but not bicuculline, pretreatment in the VTA region. Therefore, possible coregulation between GABA(B) and opioid receptors in the VTA region, as suggested by immunocytochemical evidence, is supported by these behavioral effects upon ingestion.

AT1 receptor blockade in the central nucleus of the amygdala attenuates the effects of muscimol on sodium and water intake.

PubMed

Hu, B; Qiao, H; Sun, B; Jia, R; Fan, Y; Wang, N; Lu, B; Yan, J Q

2015-10-29

The blockade of the central nucleus of the amygdala (CeA) with the GABAA receptor agonist muscimol significantly reduces hypertonic NaCl and water intake by sodium-depleted rats. In the present study we investigated the effects of previous injection of losartan, an angiotensin II type-1 (AT1) receptor antagonist, into the CeA on 0.3M NaCl and water intake reduced by muscimol bilaterally injected into the same areas in rats submitted to water deprivation-partial rehydration (WD-PR) and in rats treated with the diuretic furosemide (FURO). Male Sprague-Dawley rats with stainless steel cannulas bilaterally implanted into the CeA were used. Bilateral injections of muscimol (0.2 nmol/0.5 μl, n=8 rats/group) into the CeA in WD-PR-treated rats reduced 0.3M NaCl intake and water intake, and pre-treatment of the CeA with losartan (50 μg/0.5 μl) reversed the inhibitory effect of muscimol. The negative effect of muscimol on sodium and water intake could also be blocked by pretreatment with losartan microinjected into the CeA in rats given FURO (n=8 rats/group). However, bilateral injections of losartan (50 μg/0.5 μl) alone into the CeA did not affect the NaCl or water intake. These results suggest that the deactivation of CeA facilitatory mechanisms by muscimol injection into the CeA is promoted by endogenous angiotensin II acting on AT1 receptors in the CeA, which prevents rats from ingesting large amounts of hypertonic NaCl and water. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

The role of GABA-A and mitochondrial diazepam-binding inhibitor receptors on the effects of neurosteroids on food intake in mice.

PubMed

Reddy, D S; Kulkarni, S K

1998-06-01

The present studies were undertaken to investigate the neuroactive steroidal modulation of feeding behavior and possible involvement of gamma-aminobutyric acid type-A (GABA-A) and mitochondrial diazepam binding inhibitor (DBI) receptors (MDR) in food-deprived male mice. Allopregnanolone (0.5-2 mg/kg), a neurosteroid, progesterone (1-10 mg/kg), a neurosteroid precursor, and 4'-chlordiazepam (0.25-1 mg/kg), a specific high affinity MDR agonist, produced a dose-dependent hyperphagic effects. In contrast, neurosteroids pregnenolone sulfate (PS) (1-10 mg/kg) and dehydroepiandrosterone sulfate (DHEAS) (1-10 mg/kg) produced a hypophagic effect, in a dose-dependent manner. The allopregnanolone-, progesterone- and 4'-chlordiazepam-induced hyperphagic effect was blocked by picrotoxin (1 mg/kg), a GABA-A chloride channel antagonist, but not by flumazenil (2 mg/kg), a benzodiazepine (BZD) antagonist. The 4'-chlordiazepam-induced hyperphagic effect was prevented by pretreatment with PK11195 (2 mg/kg), a selective partial MDR antagonist. The hypophagic effect of DHEAS (10 mg/kg) was reversed by dizocilpine (10 microg/kg), an NMDA receptor antagonist, but resistant to muscimol (0.1 mg/kg), a selective GABA-A receptor agonist. In contrast, the PS (10 mg/kg)-induced hypophagic response was resistant to dizocilpine, but sensitive to muscimol (0.1 mg/kg). Both the sulfated neurosteroids PS and DHEAS also reversed the hyperphagic effect of allopregnanolone. In addition, the BZD agonist triazolam (0.05-0.25 mg/kg) also produced a flumazenil- and picrotoxin-sensitive hyperphagic effects, thereby suggesting the changes in feeding behavior by neurosteroids represent GABA-A receptor mediated hyperphagic action. Although the possible antistress or anxiolytic actions of neurosteroids may confound the hyperphagia, behavioral effects observed were specific to food because the mice were adopted to the test environment and diet, and of a possible variation between various neurosteroids in the

Suppression of third ventricular NPY-elicited feeding following medullary reticular formation infusions of muscimol.

PubMed

Travers, Joseph B; Herman, Kenneth; Travers, Susan P

2010-04-01

The appetitive component of feeding is controlled by forebrain substrates, but the consummatory behaviors of licking, mastication, and swallowing are organized in the brainstem. The target of forebrain appetitive signals is unclear but likely includes regions of the medullary reticular formation (RF). This study was undertaken to determine the necessity of different RF regions for mastication induced by a descending appetitive signal. We measured solid food intake in response to third ventricular (3V) infusions of the orexigenic peptide neuropeptide Y 3-36 in awake, freely moving rats and determined whether focal RF infusions of the GABAA agonist muscimol suppressed eating. RF infusions were centered in either the lateral tegmental field, comprising the intermediate (IRt) and parvocellular (PCRt) RF, or in the nucleus gigantocellularis (Gi). Infusions of NPY 3-36 (5 microg/5 microl) into 3V significantly increased feeding of solid food over a 90-min period compared with the noninfused condition (4.3 g +/- 0.56 vs. 0.57 g +/- 0.57, p < .001). NPY 3-36-induced food intake was suppressed (1.7 g +/- 0.48) by simultaneous infusions of muscimol (0.6 mM/100 nl) into the IRt/PCRt (p < .01). Coincident with the decrease in feeding was a decrease in the amplitude of anterior digastric muscle contractions in response to intraoral sucrose infusions. In contrast, infusions of muscimol into Gi had no discernible effect on food intake or EMG amplitude. These data suggest that the IRt/PCRt is essential for forebrain-initiated mastication, but that the Gi is not a necessary link in this pathway.

The GABA[subscript A] Receptor Agonist Muscimol Induces an Age- and Region-Dependent Form of Long-Term Depression in the Mouse Striatum

ERIC Educational Resources Information Center

Zhang, Xiaoqun; Yao, Ning; Chergui, Karima

2016-01-01

Several forms of long-term depression (LTD) of glutamatergic synaptic transmission have been identified in the dorsal striatum and in the nucleus accumbens (NAc). Such experience-dependent synaptic plasticity might play important roles in reward-related learning. The GABA[subscript A] receptor agonist muscimol was recently found to trigger a…

Homocysteine alters cerebral microvascular integrity and causes remodeling by antagonizing GABA-A receptor*

PubMed Central

Lominadze, David; Tyagi, Neetu; Sen, Utpal; Ovechkin, Alexander; Tyagi, Suresh C.

2012-01-01

High levels of homocysteine (Hcy), known as hyperhomocysteinemia (HHcy), are associated with cerebrovascular diseases, such as vascualr dementia, stroke, and Alzheimer's disease. The -amino butyric acid (GABA) is a inhibitory neurotransmitter and a ligand of GABA-A receptor. By inhibiting excitatory response it may decrease complications associated with vascular dementia and stroke. Hcy specifically competes with the GABA-A receptors and acts as an excitotoxic neurotransmitter. Previously we have shown that Hcy increases levels of NADPH oxidase and reactive oxygen species (ROS), and decreases levels of thioredoxin and peroxiredoxin by antagonizing the GABA-A receptor. Hcy treatment leads to activation of matrix metalloproteinases (MMPs) in cerebral circualtion by inducing redox stress and ROS. The hypothesis is that Hcy induces MMPs and suppresses tissue inhibitors of metalloproteinase (TIMPs), in part, by inhibiting the GABA-A receptor. This leads to degradation of the matrix and disruption of the blood brain barrier. The brain cortex of transgenic mouse model of HHcy (cystathionine -synthase, CBS −/+) and GABA-A receptor null mice treated with and without muscimol (GABA-A receptor agonist) was analysed. The mRNA levels were measured by Q-RT-PCR. Levels of MMP-2, -9, -13, and TIMP-1, -2, -3, and -4 were evaluated by in situ labeling and PCR-gene arrays. Pial venular permeability to fluorescence-labeled albumin was assessed with intravital fluorescence microscopy. We found that Hcy increases metalloproteinase activity and decreases TIMP-4 by antagonizing the GABA-A receptor. The results demonstrate a novel mechanism in which brain microvascular permeability changes during HHcy and vascular dementias, and have therapeutic ramifications for microvascular disease in Alzheimer's patients. PMID:22886392

GABA(A) receptors mediate orexin-A induced stimulation of food intake.

PubMed

Kokare, Dadasaheb M; Patole, Angad M; Carta, Anna; Chopde, Chandrabhan T; Subhedar, Nishikant K

2006-01-01

Although the role of orexins in sleep/wake cycle and feeding behavior is well established, underlying mechanisms have not been fully understood. An attempt has been made to investigate the role of GABA(A) receptors and their benzodiazepine site on the orexin-A induced response to feeding. Different groups of rats were food deprived overnight and next day injected intracerebroventricularly (icv) with vehicle (artificial CSF; 5 microl/rat) or orexin-A (20-50 nM/rat) and the animals were given free access to food. Cumulative food intake was measured during light phase of light/dark cycle at 1-, 2-, 4- and 6-h post-injection time points. Orexin-A (30-50 nM/rat, icv) stimulated food intake at all the time points (P < 0.05). Prior administration of GABA(A) receptor agonists muscimol (25 ng/rat, icv) and diazepam (0.5 mg/kg, ip) at subeffective doses significantly potentiated the hyperphagic effect of orexin-A (30 nM/rat, icv). However, the effect was negated by the GABA(A) receptor antagonist bicuculline (1 mg/kg, ip). Interestingly, benzodiazepine receptor antagonist flumazenil (5 ng/rat, icv), augmented the orexin-A (30 nM/rat, icv) induced hyperphagia; the effect may be attributed to the intrinsic activity of the agent. The results suggest that the hyperphagic effect of orexin-A, at least in part, is mediated by enhanced GABA(A) receptor activity.

Recovery from ketamine-induced amnesia by blockade of GABA-A receptor in the medial prefrontal cortex of mice.

PubMed

Farahmandfar, Maryam; Akbarabadi, Ardeshir; Bakhtazad, Atefeh; Zarrindast, Mohammad-Reza

2017-03-06

Ketamine and other noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonists are known to induce deficits in learning and cognitive performance sensitive to prefrontal cortex (PFC) functions. The interaction of a glutamatergic and GABAergic systems is essential for many cognitive behaviors. In order to understand the effect of γ-aminobutyric acid (GABA)/glutamate interactions on learning and memory, we investigated the effects of intra medial prefrontal cortex (mPFC) injections of GABAergic agents on ketamine-induced amnesia using a one-trial passive avoidance task in mice. Pre-training systemic administration of ketamine (5, 10 and 15mg/kg, i.p.) dose-dependently decreased the memory acquisition of a one-trial passive avoidance task. Pre-training intra-mPFC injection of muscimol, GABAA receptor agonist (0.05, 0.1 and 0.2μg/mouse) and baclofen GABAB receptor agonist (0.05, 0.1, 0.5 and 1μg/mouse), impaired memory acquisition. However, co-pretreatment of different doses of muscimol and baclofen with a lower dose of ketamine (5mg/kg), which did not induce amnesia by itself, caused inhibition of memory formation. Our data showed that sole pre-training administration of bicuculline, GABA-A receptor antagonist and phaclofen GABA-B receptor antagonist into the mPFC, did not affect memory acquisition. In addition, the amnesia induced by pre-training ketamine (15mg/kg) was significantly decreased by the pretreatment of bicuculline (0.005, 0.1 and 0.5μg/mouse). It can be concluded that GABAergic system of the mPFC is involved in the ketamine-induced impairment of memory acquisition. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Nitric oxide in the dorsal hippocampal area is involved on muscimol state-dependent memory in the step-down passive avoidance test.

PubMed

Jafari-Sabet, Majid; Khodadadnejad, Mohammad-Amin; Ghoraba, Saeed; Ataee, Ramin

2014-02-01

In the present study, the effects of intra-dorsal hippocampal (intra-CA1) injections of nitric oxide (NO) agents on muscimol state-dependent memory were examined in mice. A single-trial step-down passive avoidance task was used for the assessment of memory retrieval in adult male NMRI mice. Post-training intra-CA1 administration of a GABAA receptor agonist, muscimol (0.05 and 0.1 μg/mouse) dose dependently induced impairment of memory retention. Pre-test injection of muscimol (0.05 and 0.1 μg/mouse) induced state-dependent retrieval of the memory acquired under post-training muscimol (0.1 μg/mouse, intra-CA1) influence. Pre-test injection of a NO precursor, L-arginine (1 and 2 μg/mouse, intra-CA1) improved memory retention, although the low dose of the drug (0.5 μg/mouse) did not affect memory retention. Pre-test injection of an inhibitor of NO-synthase, L-NAME (0.5 and 1 μg/mouse, intra-CA1) impaired memory retention, although the low dose of the drug (0.25 μg/mouse) did not affect memory retention. In other series of experiments, pre-test intra-CA1 injection of L-arginine (0.25 and 0.5 μg/mouse) 5 min before the administration of muscimol (0.1 μg/mouse, intra-CA1) dose dependently inhibited muscimol state-dependent memory. Pre-test intra-CA1 administration of L-arginine (0.125, 0.25 and 0.5 μg/mouse) by itself cannot affect memory retention. Pre-test intra-CA1 injection of L-NAME (0.25 μg/mouse, intra-CA1) reversed the memory impairment induced by post-training administration of muscimol (0.1 μg/mouse, intra-CA1). Moreover, pre-test administration of L-NAME (0.125 and 0.25 μg/mouse, intra-CA1) with an ineffective dose of muscimol (0.025 μg/mouse, intra-CA1) significantly restored the retrieval and induced muscimol state-dependent memory. Pre-test intra-CA1 administration of L-NAME (0.0625, 0.125 and 0.25 μg/mouse) by itself cannot affect memory retention. It may be suggested that the nitric oxide in the dorsal hippocampal area play an important

GABAA overactivation potentiates the effects of NMDA blockade during the brain growth spurt in eliciting locomotor hyperactivity in juvenile mice.

PubMed

Oliveira-Pinto, Juliana; Paes-Branco, Danielle; Cristina-Rodrigues, Fabiana; Krahe, Thomas E; Manhães, Alex C; Abreu-Villaça, Yael; Filgueiras, Cláudio C

2015-01-01

Both NMDA receptor blockade and GABAA receptor overactivation during the brain growth spurt may contribute to the hyperactivity phenotype reminiscent of attention-deficit/hyperactivity disorder. Here, we evaluated the effects of exposure to MK801 (a NMDA antagonist) and/or to muscimol (a GABAA agonist) during the brain growth spurt on locomotor activity of juvenile Swiss mice. This study was carried out in two separate experiments. In the first experiment, pups received a single i.p. injection of either saline solution (SAL), MK801 (MK, 0.1, 0.3 or 0.5 mg/kg) or muscimol (MU, 0.02, 0.1 or 0.5 mg/kg) at the second postnatal day (PND2), and PNDs 4, 6 and 8. In the second experiment, we investigated the effects of a combined injection of MK (0.1 mg/kg) and MU (doses: 0.02, 0.1 or 0.5 mg/kg) following the same injection schedule of the first experiment. In both experiments, locomotor activity was assessed for 15 min at PND25. While MK promoted a dose-dependent increase in locomotor activity, exposure to MU failed to elicit significant effects. The combined exposure to the highest dose of MU and the lowest dose of MK induced marked hyperactivity. Moreover, the combination of the low dose of MK and the high dose of MU resulted in a reduced activity in the center of the open field, suggesting an increased anxiety-like behavior. These findings suggest that, during the brain growth spurt, the blockade of NMDA receptors induces juvenile locomotor hyperactivity whereas hyperactivation of GABAA receptors does not. However, GABAA overactivation during this period potentiates the effects of NMDA blockade in inducing locomotor hyperactivity. Copyright © 2015 Elsevier Inc. All rights reserved.

Population patch-clamp electrophysiology analysis of recombinant GABAA alpha1beta3gamma2 channels expressed in HEK-293 cells.

PubMed

Hollands, Emma C; Dale, Tim J; Baxter, Andrew W; Meadows, Helen J; Powell, Andrew J; Clare, Jeff J; Trezise, Derek J

2009-08-01

Gamma-amino butyric acid (GABA)-activated Cl- channels are critical mediators of inhibitory postsynaptic potentials in the CNS. To date, rational design efforts to identify potent and selective GABA(A) subtype ligands have been hampered by the absence of suitable high-throughput screening approaches. The authors describe 384-well population patch-clamp (PPC) planar array electrophysiology methods for the study of GABA(A) receptor pharmacology. In HEK293 cells stably expressing human alpha1beta3gamma2 GABA(A) channels, GABA evoked outward currents at 0 mV of 1.05 +/- 0.08 nA, measured 8 s post GABA addition. The I(GABA) was linear and reversed close to the theoretical E(Cl) (-56 mV). Concentration-response curve analysis yielded a mean pEC(50) value of 5.4 and Hill slope of 1.5, and for a series of agonists, the rank order of potency was muscimol > GABA > isoguvacine. A range of known positive modulators, including diazepam and pentobarbital, produced concentration-dependent augmentation of the GABA EC( 20) response (1 microM). The competitive antagonists bicuculline and gabazine produced concentration-dependent, parallel, rightward displacement of GABA curves with pA(2) and slope values of 5.7 and 1.0 and 6.7 and 1.0, respectively. In contrast, picrotoxin (0.2-150 microM) depressed the maximal GABA response, implying a non-competitive antagonism. Overall, the pharmacology of human alpha1beta3gamma2 GABA(A) determined by PPC was highly similar to that obtained by conventional patch-clamp methods. In small-scale single-shot screens, Z' values of >0.5 were obtained in agonist, modulator, and antagonist formats with hit rates of 0% to 3%. The authors conclude that despite the inability of the method to resolve the peak agonist responses, PPC can rapidly and usefully quantify pharmacology for the alpha1beta3gamma2 GABA(A) isoform. These data suggest that PPC may be a valuable approach for a focused set and secondary screening of GABA(A) receptors and other slow ligand

The effects of agonists of ionotropic GABA(A) and metabotropic GABA(B) receptors on learning.

PubMed

Zyablitseva, Evgeniya A; Kositsyn, Nikolay S; Shul'gina, Galina I

2009-05-01

The research described here investigates the role played by inhibitory processes in the discriminations made by the nervous system of humans and animals between familiar and unfamiliar and significant and nonsignificant events. This research compared the effects of two inhibitory mediators of gamma-aminobutyric acid (GABA): 1) phenibut, a nonselective agonist of ionotropic GABA(A) and metabotropic GABA(B) receptors and 2) gaboxadol a selective agonist of ionotropic GABA(A) receptors on the process of developing active defensive and inhibitory conditioned reflexes in alert non-immobilized rabbits. It was found that phenibut, but not gaboxadol, accelerates the development of defensive reflexes at an early stage of conditioning. Both phenibut and gaboxadol facilitate the development of conditioned inhibition, but the effect of gaboxadol occurs at later stages of conditioning and is less stable than that of phenibut. The earlier and more stable effects of phenibut, as compared to gaboxadol, on storage in memory of the inhibitory significance of a stimulus may occur because GABA(B) receptors play the dominant role in the development of internal inhibition during an early stage of conditioning. On the other hand this may occur because the participation of both GABA(A) and GABA(B) receptors are essential to the process. We discuss the polyfunctionality of GABA receptors as a function of their structure and the positions of the relevant neurons in the brain as this factor can affect regulation of various types of psychological processes.

CO2-induced ocean acidification increases anxiety in Rockfish via alteration of GABAA receptor functioning

PubMed Central

Hamilton, Trevor James; Holcombe, Adam; Tresguerres, Martin

2014-01-01

The average surface pH of the ocean is dropping at a rapid rate due to the dissolution of anthropogenic CO2, raising concerns for marine life. Additionally, some coastal areas periodically experience upwelling of CO2-enriched water with reduced pH. Previous research has demonstrated ocean acidification (OA)-induced changes in behavioural and sensory systems including olfaction, which is due to altered function of neural gamma-aminobutyric acid type A (GABAA) receptors. Here, we used a camera-based tracking software system to examine whether OA-dependent changes in GABAA receptors affect anxiety in juvenile Californian rockfish (Sebastes diploproa). Anxiety was estimated using behavioural tests that measure light/dark preference (scototaxis) and proximity to an object. After one week in OA conditions projected for the next century in the California shore (1125 ± 100 µatm, pH 7.75), anxiety was significantly increased relative to controls (483 ± 40 µatm CO2, pH 8.1). The GABAA-receptor agonist muscimol, but not the antagonist gabazine, caused a significant increase in anxiety consistent with altered Cl− flux in OA-exposed fish. OA-exposed fish remained more anxious even after 7 days back in control seawater; however, they resumed their normal behaviour by day 12. These results show that OA could severely alter rockfish behaviour; however, this effect is reversible. PMID:24285203

CO2-induced ocean acidification increases anxiety in rockfish via alteration of GABAA receptor functioning.

PubMed

Hamilton, Trevor James; Holcombe, Adam; Tresguerres, Martin

2014-01-22

The average surface pH of the ocean is dropping at a rapid rate due to the dissolution of anthropogenic CO2, raising concerns for marine life. Additionally, some coastal areas periodically experience upwelling of CO2-enriched water with reduced pH. Previous research has demonstrated ocean acidification (OA)-induced changes in behavioural and sensory systems including olfaction, which is due to altered function of neural gamma-aminobutyric acid type A (GABAA) receptors. Here, we used a camera-based tracking software system to examine whether OA-dependent changes in GABAA receptors affect anxiety in juvenile Californian rockfish (Sebastes diploproa). Anxiety was estimated using behavioural tests that measure light/dark preference (scototaxis) and proximity to an object. After one week in OA conditions projected for the next century in the California shore (1125 ± 100 µatm, pH 7.75), anxiety was significantly increased relative to controls (483 ± 40 µatm CO2, pH 8.1). The GABAA-receptor agonist muscimol, but not the antagonist gabazine, caused a significant increase in anxiety consistent with altered Cl(-) flux in OA-exposed fish. OA-exposed fish remained more anxious even after 7 days back in control seawater; however, they resumed their normal behaviour by day 12. These results show that OA could severely alter rockfish behaviour; however, this effect is reversible.

Muscimol inactivation caudal to the interstitial nucleus of Cajal induces hemi-seesaw nystagmus.

PubMed

Das, Vallabh E; Leigh, R John; Swann, Michelle; Thurtell, Matthew J

2010-09-01

Hemi-seesaw nystagmus (hemi-SSN) is a jerk-waveform nystagmus with conjugate torsional and disjunctive vertical components. Halmagyi et al. in Brain 117(Pt 4):789-803 (1994), reported hemi-SSN in patients with unilateral lesions in the vicinity of the Interstitial Nucleus of Cajal (INC) and suggested that an imbalance in projections from the vestibular nuclei to the INC was the source of the nystagmus. However, this hypothesis was called into question by Helmchen et al. in Exp Brain Res 119(4):436-452 (1998), who inactivated INC in monkeys with muscimol (a GABA(A) agonist) and induced failure of vertical gaze-holding (neural integrator) function but not hemi-SSN. We injected 0.1-0.2 microl of 2% muscimol into the supraoculomotor area, 1-2 mm dorso-lateral to the right oculomotor nucleus and caudal to the right INC. A total of seven injections in two juvenile rhesus monkeys were performed. Hemi-SSN was noted within 5-10 min after injection for six of the injections. Around the time the hemi-SSN began, a small skew deviation also developed. However, there was no limitation of horizontal or vertical eye movements, suggesting that the nearby oculomotor nucleus was not initially compromised. Limitations in eye movement range developed about (1/2)-1 h following the injections. Clinical signs that were observed after the animal was released to his cage included a moderate to marked head tilt toward the left (contralesional) side, consistent with an ocular tilt reaction. We conclude that hemi-SSN can be caused by lesions just caudal to the INC, whereas lesions of the INC itself cause down-beat nystagmus and vertical gaze-holding failure, as demonstrated by Helmchen et al. Combined deficits may be encountered with lesions that involve several midbrain structures.

Muscimol inactivation caudal to the interstitial nucleus of Cajal induces hemi-seesaw nystagmus

PubMed Central

Leigh, R. John; Swann, Michelle; Thurtell, Matthew J.

2010-01-01

Hemi-seesaw nystagmus (hemi-SSN) is a jerk-waveform nystagmus with conjugate torsional and disjunctive vertical components. Halmagyi et al. in Brain 117(Pt 4):789–803 (1994), reported hemi-SSN in patients with unilateral lesions in the vicinity of the Interstitial Nucleus of Cajal (INC) and suggested that an imbalance in projections from the vestibular nuclei to the INC was the source of the nystagmus. However, this hypothesis was called into question by Helmchen et al. in Exp Brain Res 119(4):436–452 (1998), who inactivated INC in monkeys with muscimol (a GABAA agonist) and induced failure of vertical gaze-holding (neural integrator) function but not hemi-SSN. We injected 0.1–0.2 μl of 2% muscimol into the supraoculomotor area, 1–2 mm dorso-lateral to the right oculomotor nucleus and caudal to the right INC. A total of seven injections in two juvenile rhesus monkeys were performed. Hemi-SSN was noted within 5–10 min after injection for six of the injections. Around the time the hemi-SSN began, a small skew deviation also developed. However, there was no limitation of horizontal or vertical eye movements, suggesting that the nearby oculomotor nucleus was not initially compromised. Limitations in eye movement range developed about ½–1 h following the injections. Clinical signs that were observed after the animal was released to his cage included a moderate to marked head tilt toward the left (contralesional) side, consistent with an ocular tilt reaction. We conclude that hemi-SSN can be caused by lesions just caudal to the INC, whereas lesions of the INC itself cause down-beat nystagmus and vertical gaze-holding failure, as demonstrated by Helmchen et al. Combined deficits may be encountered with lesions that involve several midbrain structures. PMID:20686890

Effect of GABA receptor agonists or antagonists injected spinally on the blood glucose level in mice.

PubMed

Sim, Yun-Beom; Park, Soo-Hyun; Kang, Yu-Jung; Kim, Sung-Su; Kim, Chea-Ha; Kim, Su-Jin; Jung, Jun-Sub; Ryu, Ohk-Hyun; Choi, Moon-Gi; Suh, Hong-Won

2013-05-01

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.

Effects of buspirone, diazepam, and zolpidem on open field behavior, and brain [3H]muscimol binding after buspirone pretreatment.

PubMed

Siemiatkowski, M; Sienkiewicz-Jarosz, H; Członkowska, A I; Bidziński, A; Płaźnik, A

2000-07-01

The effects of 5-HT(1A) receptor agonist buspirone, a nonselective (diazepam), and a selective (zolpidem) GABA(A) receptor agonist were compared in the open field test of neophobia. Unhabituated rats were pretreated with the drugs once, prior to a first exposure to the open field, and their behavior was recorded both during this test and during a second trial 24 h later. It has been hypothesized that the decrease in exploratory activity observed during the second test session may be considered an adaptive reaction to the first day aversive experience (neophobia). If so, a selective modulation of 5-HT and GABA systems activity during the test could bring about significant changes in animal behavior on the retest. Buspirone at the lowest dose of 0.3 mg/kg revealed anxiolytic-like properties on the first day, whereas the action of diazepam and zolpidem was modulated by the dose-related sedative effect. At the dose of 2.4 mg/kg buspirone elicited delayed in time anxiolytic-like action, i.e., produced the antithigmotactic effect during the retrial 24 h later. Diazepam and zolpidem failed to exhibit similar profile of action. Autoradiography of [3H]muscimol binding after pretreatment of rats with buspirone showed a significant increase in the selective radioligand binding within the frontal cortex and a similar, near-significant tendency in the dentate gyrus of the hippocampus. The behavioral data validate buspirone as important drug for the treatment of anxiety disorders, devoid of disruptive influence on motor and cognitive processes. The open field test, as modified by us, appeared sensitive in distinguishing the behavioral profiles of action of different anxiolytic compounds, including 5-HT(1A) receptor agonist. The present results support the assumption that reduced turnover of 5-HT due to stimulation of 5-HT(1A) autoreceptors, may bring about changes in GABA(A) receptor system activity, in some brain structures, leading to the anxiolytic effect.

mGluR2/3 agonist LY379268 rescues NMDA and GABAA receptor level deficits induced in a two-hit mouse model of schizophrenia.

PubMed

Engel, Martin; Snikeris, Peta; Matosin, Natalie; Newell, Kelly Anne; Huang, Xu-Feng; Frank, Elisabeth

2016-04-01

An imbalance of excitatory and inhibitory neurotransmission underlies the glutamate hypothesis of schizophrenia. Agonists of group II metabotropic glutamate receptors, mGluR2/3, have been proposed as novel therapeutic agents to correct this imbalance. However, the influence of mGluR2/3 activity on excitatory and inhibitory neurotransmitter receptors has not been explored. We aimed to investigate the ability of a novel mGluR2/3 agonist, LY379268, to modulate the availability of the excitatory N-methyl-D-aspartate receptor (NMDA-R) and the inhibitory gamma-aminobutyrate-A receptor (GABAA-R), in a two-hit mouse model of schizophrenia. Wild type (WT) and heterozygous neuregulin 1 transmembrane domain mutant mice (NRG1 HET) were treated daily with phencyclidine (10 mg/kg ip) or saline for 14 days. After a 14-day washout, an acute dose of the mGluR2/3 agonist LY379268 (3 mg/kg), olanzapine (antipsychotic drug comparison, 1.5 mg/kg), or saline was administered. NMDA-R and GABAA-R binding densities were examined by receptor autoradiography in several schizophrenia-relevant brain regions. In both WT and NRG1 HET mice, phencyclidine treatment significantly reduced NMDA-R and GABAA-R binding density in the prefrontal cortex, hippocampus, and nucleus accumbens. Acute treatment with LY379268 restored NMDA-R and GABAA-R levels in the two-hit mouse model comparable to olanzapine. We demonstrate that the mGluR2/3 agonist LY379268 restores excitatory and inhibitory deficits with similar efficiency as olanzapine in our two-hit schizophrenia mouse model. This study significantly contributes to our understanding of the mechanisms underlying the therapeutic effects of LY379268 and supports the use of agents aimed at mGluR2/3.

GABAA- and glycine-mediated inhibitory modulation of the cough reflex in the caudal nucleus tractus solitarii of the rabbit.

PubMed

Cinelli, Elenia; Iovino, Ludovica; Bongianni, Fulvia; Pantaleo, Tito; Mutolo, Donatella

2016-09-01

Cough-related sensory inputs from rapidly adapting receptors (RARs) and C fibers are processed by second-order neurons mainly located in the caudal nucleus tractus solitarii (NTS). Both GABAA and glycine receptors have been proven to be involved in the inhibitory control of second-order cells receiving RAR projections. We investigated the role of these receptors within the caudal NTS in the modulation of the cough reflex induced by either mechanical or chemical stimulation of the tracheobronchial tree in pentobarbital sodium-anesthetized, spontaneously breathing rabbits. Bilateral microinjections (30-50 nl) of the receptor antagonists bicuculline and strychnine as well as of the receptor agonists muscimol and glycine were performed. Bicuculline (0.1 mM) and strychnine (1 mM) caused decreases in peak abdominal activity and marked increases in respiratory frequency due to decreases in both inspiratory time (Ti) and expiratory time (Te), without concomitant changes in arterial blood pressure. Noticeably, these microinjections induced potentiation of the cough reflex consisting of increases in the cough number associated with decreases either in cough-related Ti after bicuculline or in both cough-related Ti and Te after strychnine. The effects caused by muscimol (0.1 mM) and glycine (10 mM) were in the opposite direction to those produced by the corresponding antagonists. The results show that both GABAA and glycine receptors within the caudal NTS mediate a potent inhibitory modulation of the pattern of breathing and cough reflex responses. They strongly suggest that disinhibition is one important mechanism underlying cough regulation and possibly provide new hints for novel effective antitussive strategies. Copyright © 2016 the American Physiological Society.

PWZ-029, A COMPOUND WITH MODERATE INVERSE AGONIST FUNCTIONAL SELECTIVITY AT GABAA RECEPTORS CONTAINING α5 SUBUNITS, IMPROVES PASSIVE, BUT NOT ACTIVE, AVOIDANCE LEARNING IN RATS

PubMed Central

Savić, Miroslav M.; Clayton, Terry; Furtmüller, Roman; Gavrilović, Ivana; Samardžić, Janko; Savić, Snežana; Huck, Sigismund; Sieghart, Werner; Cook, James M.

2008-01-01

Benzodiazepine (BZ) site ligands affect vigilance, anxiety, memory processes, muscle tone and epileptogenic propensity through modulation of neurotransmission at GABAA receptors containing α1, α2, α3 or α5 subunits, and may have numerous experimental and clinical applications. The ability of nonselective BZ site inverse agonists to enhance cognition, documented in animal models and human studies, is clinically not feasible due to potentially unacceptable psychomotor effects. Most investigations to date have proposed the α1 and/or α5 subunit-containing GABAA receptors as comprising the memory-modulating population of these receptors. The novel ligand PWZ-029, which we synthesised and characterized electrophysiologically, possesses in vitro binding selectivity and moderate inverse agonist functional selectivity at α5-containing GABAA receptors. This ligand has also been examined in rats in the passive and active avoidance, spontaneous locomotor activity, elevated plus maze and grip strength tests, primarily predictive of the effects on the memory acquisition, basal locomotor activity, anxiety level and muscle tone, respectively. The improvement of task learning was detected at the dose of 5 mg/kg in the passive, but not active avoidance test. The inverse agonist PWZ-029 had no effect on anxiety or muscle tone, whereas at higher doses (10 and 20 mg/kg) it decreased locomotor activity. This effect was antagonized by flumazenil and also by the lower (but not the higher) dose of an agonist (SH-053-R-CH3-2’F) selective for GABAA receptors containing the α5 subunit. The hypolocomotor effect of PWZ-029 was not antagonized by the antagonist β-CCt exhibiting a preferential affinity for α1-subunit containing receptors. These data suggest that moderate negative modulation at GABAA receptors containing the α5 subunit is a sufficient condition for eliciting enhanced encoding/consolidation of declarative memory, while the influence of higher doses of modulators at

Low concentrations of ethanol do not affect radioligand binding to the delta-subunit-containing GABAA receptors in the rat brain.

PubMed

Mehta, Ashok K; Marutha Ravindran, C R; Ticku, Maharaj K

2007-08-24

In the present study, we investigated the co-localization pattern of the delta subunit with other subunits of GABA(A) receptors in the rat brain using immunoprecipitation and Western blotting techniques. Furthermore, we investigated whether low concentrations of ethanol affect the delta-subunit-containing GABA(A) receptor assemblies in the rat brain using radioligand binding to the rat brain membrane homogenates as well as to the immunoprecipitated receptor assemblies. Our results revealed that delta subunit is not co-localized with gamma(2) subunit but it is associated with the alpha(1), alpha(4) or alpha(6), beta(2) and/or beta(3) subunit(s) of GABA(A) receptors in the rat brain. Ethanol (1-50 mM) neither affected [(3)H]muscimol (3 nM) binding nor diazepam-insensitive [(3)H]Ro 15-4513 (2 nM) binding in the rat cerebellum and cerebral cortex membranes. However, a higher concentration of ethanol (500 mM) inhibited the binding of these radioligands to the GABA(A) receptors partially in the rat cerebellum and cerebral cortex. Similarly, ethanol (up to 50 mM) did not affect [(3)H]muscimol (15 nM) binding to the immunoprecipitated delta-subunit-containing GABA(A) receptor assemblies in the rat cerebellum and hippocampus but it inhibited the binding partially at a higher concentration (500 mM). These results suggest that the native delta-subunit-containing GABA(A) receptors do not play a major role in the pharmacology of clinically relevant low concentrations of ethanol.

The vigilance promoting drug modafinil increases extracellular glutamate levels in the medial preoptic area and the posterior hypothalamus of the conscious rat: prevention by local GABAA receptor blockade.

PubMed

Ferraro, L; Antonelli, T; Tanganelli, S; O'Connor, W T; Perez de la Mora, M; Mendez-Franco, J; Rambert, F A; Fuxe, K

1999-04-01

The effects of modafinil on glutamatergic and GABAergic transmission in the rat medial preoptic area (MPA) and posterior hypothalamus (PH), are analysed. Modafinil (30-300 mg/kg) increased glutamate and decreased GABA levels in the MPA and PH. Local perfusion with the GABAA agonist muscimol (10 microM), reduced, while the GABAA antagonist bicuculline (1 microM and 10 microM) increased glutamate levels. The modafinil (100 mg/kg)-induced increase of glutamate levels was antagonized by local perfusion with bicuculline (1 microM). When glutamate levels were increased by the local perfusion with the glutamate uptake inhibitor L-trans-PDC (0.5 mM), modafinil produced an additional enhancement of glutamate levels. Modafinil (1-33 microM) failed to affect [3H]glutamate uptake in hypothalamic synaptosomes and slices. These findings show that modafinil increases glutamate and decreases GABA levels in MPA and PH. The evidence that bicuculline counteracts the modafinil-induced increase of glutamate levels strengthens the evidence for an inhibitory GABA/glutamate interaction in the above regions controlling the sleep-wakefulness cycle.

Lack of effect of mossy fiber-released zinc on granule cell GABA(A) receptors in the pilocarpine model of epilepsy.

PubMed

Molnár, P; Nadler, J V

2001-05-01

The recurrent mossy fiber pathway of the dentate gyrus expands dramatically in the epileptic brain and serves as a mechanism for synchronization of granule cell epileptiform activity. It has been suggested that this pathway also promotes epileptiform activity by inhibiting GABA(A) receptor function through release of zinc. Hippocampal slices from pilocarpine-treated rats were used to evaluate this hypothesis. The rats had developed status epilepticus after pilocarpine administration, followed by robust recurrent mossy fiber growth. The ability of exogenously applied zinc to depress GABA(A) receptor function in dentate granule cells depended on removal of polyvalent anions from the superfusion medium. Under these conditions, 200 microM zinc reduced the amplitude of the current evoked by applying muscimol to the proximal portion of the granule cell dendrite (23%). It also reduced the mean amplitude (31%) and frequency (36%) of miniature inhibitory postsynaptic currents. Nevertheless, repetitive mossy fiber stimulation (10 Hz for 1 s, 100 Hz for 1 s, or 10 Hz for 5 min) at maximal intensity did not affect GABA(A) receptor-mediated currents evoked by photorelease of GABA onto the proximal portion of the dendrite, where recurrent mossy fiber synapses were located. These results could not be explained by stimulation-induced depletion of zinc from the recurrent mossy fiber boutons. Negative results were obtained even during exposure to conditions that promoted transmitter release and synchronized granule cell activity (6 mM [K(+)](o), nominally Mg(2+)-free medium, 33 degrees C). These results suggest that zinc released from the recurrent mossy fiber pathway did not reach a concentration at postsynaptic GABA(A) receptors sufficient to inhibit agonist-evoked activation.

Microinjection of muscimol into caudal periaqueductal gray lowers body temperature and attenuates increases in temperature and activity evoked from the dorsomedial hypothalamus.

PubMed

de Menezes, Rodrigo C A; Zaretsky, Dmitry V; Fontes, Marco A P; DiMicco, Joseph A

2006-05-30

Microinjection of the neuronal inhibitor muscimol into the midbrain lateral/dorsolateral periaqueductal gray (l/dlPAG) suppresses increases in heart rate (HR) and mean arterial pressure (MAP) evoked by microinjection of the GABA(A) receptor antagonist bicuculline methiodide (BMI) into the dorsomedial hypothalamus (DMH) in rats. Injection of BMI into the DMH also increases body temperature (Tco) and motor activity. Here, our goal was to extend previous findings by examining the effect of microinjection of muscimol into the PAG on these thermogenic and behavioral responses in conscious freely moving rats. Microinjection of muscimol (300 pmol and 1 nmol) alone into the l/dlPAG reduced baseline Tco without affecting activity, HR, or MAP. Similar injection of a dose that failed to alter baseline Tco (100 pmol) suppressed the increases in Tco evoked from the DMH and significantly attenuated DMH-induced increases in locomotor activity. Whereas microinjection of 1 nmol muscimol into the ldlPAG abolished the increases in Tco evoked from the DMH and in fact lowered body temperature to a degree similar to that seen after this dose of muscimol alone, 1 nmol muscimol at adjacent sites outside the targeted region of the PAG had no significant effect on DMH-induced increases in Tco or any other parameter. These results indicate a role for neuronal activity in the l/dlPAG in (1) the temperature and behavioral responses to disinhibition of neurons in the DMH, and (2) the maintenance of basal body temperature in conscious freely moving rats.

The Basolateral Amygdala Is Necessary for the Encoding and the Expression of Odor Memory

ERIC Educational Resources Information Center

Sevelinges, Yannick; Desgranges, Bertrand; Ferreira, Guillaume

2009-01-01

Conditioned odor avoidance (COA) results from the association between a novel odor and a delayed visceral illness. The present experiments investigated the role of the basolateral amygdala (BLA) in acquisition and retrieval of COA memory. To address this, we used the GABAA agonist muscimol to temporarily inactivate the BLA during COA acquisition…

GABAA Receptor Regulation of Voluntary Ethanol Drinking Requires PKCε

PubMed Central

Besheer, Joyce; Lepoutre, Veronique; Mole, Beth; Hodge, Clyde W.

2010-01-01

Protein kinase C (PKC) regulates a variety of neural functions, including ion channel activity, neurotransmitter release, receptor desensitization and differentiation. We have shown previously that mice lacking the ε-isoform of PKC (PKCε) self-administer 75% less ethanol and exhibit supersensitivity to acute ethanol and allosteric positive modulators of GABAA receptors when compared with wild-type controls. The purpose of the present study was to examine involvement of PKCε in GABAA receptor regulation of voluntary ethanol drinking. To address this question, PKCε null-mutant and wild-type control mice were allowed to drink ethanol (10% v/v) vs. water on a two-bottle continuous access protocol. The effects of diazepam (nonselective GABAA BZ positive modulator), zolpidem (GABAA α1 agonist), L-655,708 (BZ-sensitive GABAA α5 inverse agonist), and flumazenil (BZ antagonist) were then tested on ethanol drinking. Ethanol intake (grams/kg/day) by wild-type mice decreased significantly after diazepam or zolpidem but increased after L-655,708 administration. Flumazenil antagonized diazepam-induced reductions in ethanol drinking in wild-type mice. However, ethanol intake by PKCε null mice was not altered by any of the GABAergic compounds even though effects were seen on water drinking in these mice. Increased acute sensitivity to ethanol and diazepam, which was previously reported, was confirmed in PKCε null mice. Thus, results of the present study show that PKCε null mice do not respond to doses of GABAA BZ receptor ligands that regulate ethanol drinking by wild-type control mice. This suggests that PKCε may be required for GABAA receptor regulation of chronic ethanol drinking. PMID:16881070

GABAA receptor subunit gene expression in human prefrontal cortex: comparison of schizophrenics and controls

NASA Technical Reports Server (NTRS)

Akbarian, S.; Huntsman, M. M.; Kim, J. J.; Tafazzoli, A.; Potkin, S. G.; Bunney, W. E. Jr; Jones, E. G.; Bloom, F. E. (Principal Investigator)

1995-01-01

The prefrontal cortex of schizophrenics is hypoactive and displays changes related to inhibitory, GABAergic neurons, and GABAergic synapses. These changes include decreased levels of glutamic acid decarboxylase (GAD), the enzyme for GABA synthesis, upregulation of muscimol binding, and downregulation of benzodiazepine binding to GABAA receptors. Studies in the visual cortex of nonhuman primates have demonstrated that gene expression for GAD and for several GABAA receptor subunit polypeptides is under control of neuronal activity, raising the possibility that similar mechanisms in the hypoactive prefrontal cortex of schizophrenics may explain the abnormalities in GAD and in GABAA receptor regulation. In the present study, which is the first of its type on human cerebral cortex, levels of mRNAs for six GABAA receptor subunits (alpha 1, alpha 2, alpha 5, beta 1, beta 2, gamma 2) and their laminar expression patterns were analyzed in the prefrontal cortex of schizophrenics and matched controls, using in situ hybridization histochemistry and densitometry. Three types of laminar expression pattern were observed: mRNAs for the alpha 1, beta 2, and gamma 2 subunits, which are the predominant receptor subunits expressed in the mature cortex, were expressed at comparatively high levels by cells of all six cortical layers, but most intensely by cells in lower layer III and layer IV. mRNAs for the alpha 2, alpha 5, and beta 1 subunits were expressed at lower levels; alpha 2 and beta 1 were expressed predominantly by cells in layers II, III, and IV; alpha 5 was expressed predominantly in layers IV, V, and VI. There were no significant changes in overall mRNA levels for any of the receptor subunits in the prefrontal cortex of schizophrenics, and the laminar expression pattern of all six receptor subunit mRNAs did not differ between schizophrenics and controls. Because gene expression for GABAA receptor subunits is not consistently altered in the prefrontal cortex of

GABAA-current rundown of temporal lobe epilepsy is associated with repetitive activation of GABAA “phasic” receptors

PubMed Central

Palma, Eleonora; Roseti, Cristina; Maiolino, Francesca; Fucile, Sergio; Martinello, Katiuscia; Mazzuferi, Manuela; Aronica, Eleonora; Manfredi, Mario; Esposito, Vincenzo; Cantore, Gianpaolo; Miledi, Ricardo; Simonato, Michele; Eusebi, Fabrizio

2007-01-01

A study was made of the “rundown” of GABAA receptors, microtransplanted to Xenopus oocytes from surgically resected brain tissues of patients afflicted with drug-resistant human mesial temporal lobe epilepsy (mTLE). Cell membranes, isolated from mTLE neocortex specimens, were injected into frog oocytes that rapidly incorporated functional GABAA receptors. Upon repetitive activation with GABA (1 mM), “epileptic” GABAA receptors exhibited a GABAA-current (IGABA) rundown that was significantly enhanced by Zn2+ (≤250 μM), and practically abolished by the high-affinity GABAA receptor inverse agonist SR95531 (gabazine; 2.5–25 μM). Conversely, IGABA generated by “control” GABAA receptors microtransplanted from nonepileptic temporal lobe, lesional TLE, or authoptic disease-free tissues remained stable during repetitive stimulation, even in oocytes treated with Zn2+. We conclude that rundown of mTLE epileptic receptors depends on the presence of “phasic GABAA receptors” that have low sensitivity to antagonism by Zn2+. Additionally, we found that GABAA receptors, microtransplanted from the cerebral cortex of adult rats exhibiting recurrent seizures, caused by pilocarpine-induced status epilepticus, showed greater rundown than control tissue, an event also occurring in patch-clamped rat pyramidal neurons. Rundown of epileptic rat receptors resembled that of human mTLE receptors, being enhanced by Zn2+ (40 μM) and sensitive to the antiepileptic agent levetiracetam, the neurotrophin brain-derived neurotrophic factor, and the phosphatase blocker okadaic acid. Our findings point to the rundown of GABAA receptors as a hallmark of TLE and suggest that modulating tonic and phasic mTLE GABAA receptor activity may represent a useful therapeutic approach to the disease. PMID:18083839

The lateral septum and anterior hypothalamus act in tandem to regulate burying in the shock-probe test but not open-arm avoidance in the elevated plus-maze.

PubMed

Lamontagne, Steven J; Olmstead, Mary C; Menard, Janet L

2016-11-01

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. Copyright © 2016 Elsevier B.V. All rights reserved.

5-HT1A receptor blockade reverses GABAA receptor α3 subunit-mediated anxiolytic effects on stress-induced hyperthermia

PubMed Central

van Oorschot, Ruud; Korte, S. Mechiel; Olivier, Berend; Groenink, Lucianne

2010-01-01

Rationale Stress-related disorders are associated with dysfunction of both serotonergic and GABAergic pathways, and clinically effective anxiolytics act via both neurotransmitter systems. As there is evidence that the GABAA and the serotonin receptor system interact, a serotonergic component in the anxiolytic actions of benzodiazepines could be present. Objectives The main aim of the present study was to investigate whether the anxiolytic effects of (non-)selective α subunit GABAA receptor agonists could be reversed with 5-HT1A receptor blockade using the stress-induced hyperthermia (SIH) paradigm. Results The 5-HT1A receptor antagonist WAY-100635 (0.1–1 mg/kg) reversed the SIH-reducing effects of the non-α-subunit selective GABAA receptor agonist diazepam (1–4 mg/kg) and the GABAA receptor α3-subunit selective agonist TP003 (1 mg/kg), whereas WAY-100635 alone was without effect on the SIH response or basal body temperature. At the same time, co-administration of WAY-100635 with diazepam or TP003 reduced basal body temperature. WAY-100635 did not affect the SIH response when combined with the preferential α1-subunit GABAA receptor agonist zolpidem (10 mg/kg), although zolpidem markedly reduced basal body temperature. Conclusions The present study suggests an interaction between GABAA receptor α-subunits and 5-HT1A receptor activation in the SIH response. Specifically, our data indicate that benzodiazepines affect serotonergic signaling via GABAA receptor α3-subunits. Further understanding of the interactions between the GABAA and serotonin system in reaction to stress may be valuable in the search for novel anxiolytic drugs. PMID:20535452

Additive effect of harmane and muscimol for memory consolidation impairment in inhibitory avoidance task.

PubMed

Nasehi, Mohammad; Morteza-Zadeh, Parastoo; Khakpai, Fatemeh; Zarrindast, Mohammad-Reza

2016-12-17

In the current study, we examined the effect of bilateral intra-dorsal hippocampal (intra-CA1) microinjections of GABA A receptor agents on amnesia induced by a β-carboline alkaloid, harmane in mice. We used a single-trial step-down passive avoidance task to assess memory retention and then, open-field test to assess locomotor activity. The results indicated that post-training intra-CA1 injections of bicuculline - a GABA A receptor antagonist - had no significant effect, while muscimol (0.01 and 0.1μg/mouse) - a GABA A receptor agonist - impaired memory consolidation. Post-training intra-peritoneal (i.p.) infusion of harmane (3 and 5mg/kg) decreased memory consolidation. Furthermore, post-training intra-CA1 administration of sub-threshold dose of bicuculline (0.001μg/mouse) restored, whereas muscimol (0.001μg/mouse) potentiated impairment of memory consolidation induced by harmane. The isobologram analysis revealed that there is an additive effect between harmane and muscimol on impairment of memory consolidation. Moreover, all above doses of drugs did not alter locomotor activity. These findings suggest that GABA A receptors of the CA1 area, at least partly, play a role in modulating the effect of harmane on memory consolidation. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

MmTX1 and MmTX2 from coral snake venom potently modulate GABAA receptor activity.

PubMed

Rosso, Jean-Pierre; Schwarz, Jürgen R; Diaz-Bustamante, Marcelo; Céard, Brigitte; Gutiérrez, José M; Kneussel, Matthias; Pongs, Olaf; Bosmans, Frank; Bougis, Pierre E

2015-02-24

GABAA receptors shape synaptic transmission by modulating Cl(-) conductance across the cell membrane. Remarkably, animal toxins that specifically target GABAA receptors have not been identified. Here, we report the discovery of micrurotoxin1 (MmTX1) and MmTX2, two toxins present in Costa Rican coral snake venom that tightly bind to GABAA receptors at subnanomolar concentrations. Studies with recombinant and synthetic toxin variants on hippocampal neurons and cells expressing common receptor compositions suggest that MmTX1 and MmTX2 allosterically increase GABAA receptor susceptibility to agonist, thereby potentiating receptor opening as well as desensitization, possibly by interacting with the α(+)/β(-) interface. Moreover, hippocampal neuron excitability measurements reveal toxin-induced transitory network inhibition, followed by an increase in spontaneous activity. In concert, toxin injections into mouse brain result in reduced basal activity between intense seizures. Altogether, we characterized two animal toxins that enhance GABAA receptor sensitivity to agonist, thereby establishing a previously unidentified class of tools to study this receptor family.

Unsaturated free fatty acids increase benzodiazepine receptor agonist binding depending on the subunit composition of the GABAA receptor complex.

PubMed

Witt, M R; Westh-Hansen, S E; Rasmussen, P B; Hastrup, S; Nielsen, M

1996-11-01

It has been shown previously that unsaturated free fatty acids (FFAs) strongly enhance the binding of agonist benzodiazepine receptor ligands and GABAA receptor ligands in the CNS in vitro. To investigate the selectivity of this effect, recombinant human GABAA/benzodiazepine receptor complexes formed by different subunit compositions (alpha x beta y gamma 2, x = 1, 2, 3, and 5; y = 1, 2, and 3) were expressed using the baculovirus-transfected Sf9 insect cell system. At 10(-4) M, unsaturated FFAs, particularly arachidonic (20:4) and docosahexaenoic (22:6) acids, strongly stimulated (> 200% of control values) the binding of [3H]flunitrazepam ([3H]FNM) to the alpha 3 beta 2 gamma 2 receptor combination in whole cell preparations. No effect or small increases in levels of unsaturated FFAs on [3H]FNM binding to alpha 1 beta x gamma 2 and alpha 2 beta x gamma 2 receptor combinations were observed, and weak effects (130% of control values) were detected using the alpha 5 beta 2 gamma 2 receptor combination. The saturated FFAs, stearic and palmitic acids, were without effect on [3H]FNM binding to any combination of receptor complexes. The hydroxylated unsaturated FFAs, ricinoleic and ricinelaidic acids, were shown to decrease the binding of [3H]FNM only if an alpha 1 beta 2 gamma 2 receptor combination was used. Given the heterogeneity of the GABAA/ benzodiazepine receptor subunit distribution in the CNS, the effects of FFAs on the benzodiazepine receptor can be assumed to vary at both cellular and regional levels.

Muscimol inactivation of caudal fastigial nucleus and posterior interposed nucleus in monkeys with strabismus.

PubMed

Joshi, Anand C; Das, Vallabh E

2013-10-01

Previously, we showed that neurons in the supraoculomotor area (SOA), known to encode vergence angle in normal monkeys, encode the horizontal eye misalignment in strabismic monkeys. The SOA receives afferent projections from the caudal fastigial nucleus (cFN) and the posterior interposed nucleus (PIN) in the cerebellum. The objectives of the present study were to investigate the potential roles of the cFN and PIN in 1) conjugate eye movements and 2) binocular eye alignment in strabismic monkeys. We used unilateral injections of the GABAA agonist muscimol to reversibly inactivate the cFN (4 injections in exotropic monkey S1 with ≈ 4° of exotropia; 5 injections in esotropic monkey S2 with ≈ 34° of esotropia) and the PIN (3 injections in monkey S1). cFN inactivation induced horizontal saccade dysmetria in all experiments (mean 39% increase in ipsilesional saccade gain and 26% decrease in contralesional gain). Also, mean contralesional smooth-pursuit gain was decreased by 31%. cFN inactivation induced a divergent change in eye alignment in both monkeys, with exotropia increasing by an average of 9.8° in monkey S1 and esotropia decreasing by an average of 11.2° in monkey S2 (P < 0.001). Unilateral PIN inactivation in monkey S1 resulted in a mean increase in the gain of upward saccades by 13% and also induced a convergent change in eye alignment, reducing exotropia by an average of 2.7° (P < 0.001). We conclude that cFN/PIN influences on conjugate eye movements in strabismic monkeys are similar to those postulated in normal monkeys and cFN/PIN play important and complementary roles in maintaining the steady-state misalignment in strabismus.

Noni as an anxiolytic and sedative: a mechanism involving its gamma-aminobutyric acidergic effects.

PubMed

Deng, S; West, B J; Palu, A K; Zhou, B-N; Jensen, C J

2007-08-01

Noni (Morinda citrifolia) is increasing in worldwide popularity as a food or dietary supplement with versatile health benefits. The aim of this study was to investigate the effects of Noni fruit on anxiety symptoms in vitro. To this end, a competitive GABAa receptor-binding assay was developed. Our preliminary study indicates that the methanol crude extract of Noni fruit showed significant affinity to the gamma-aminobutyric acid A (GABAa) inhibitory neurotransmitter receptors, and displayed 75% binding inhibition of the agonist radioligand [3H] muscimol at a concentration of 100 microg/ml. Further experiments demonstrated that the MeOH extract, and its BuOH and H2O partitions, exhibited IC50 values of 22.8, 27.2, and 17.1 microg/ml, respectively, in the GABAa-binding assay. Experimental results with Noni fruit indicate the presence of competitive ligand(s), which may bind to the GABAa receptor as an agonist, and thus induce its anxiolytic and sedative effects. The study provides an in vitro rationale for one of Noni's versatile and traditional uses. In addition, an HPLC fingerprint profile of the methanolic extract of Noni fruit has been established for quality control purpose.

Contribution of ventral tegmental GABA receptors to cocaine self-administration in rats.

PubMed

Backes, E N; Hemby, S E

2008-03-01

Recent evidence has suggested that compounds affecting GABAergic transmission may provide useful pharmacological tools for the treatment of cocaine addiction. Using a rat model of self-administration, the present study examined the effects of GABA agonists and antagonists injected directly into the ventral tegmental area (VTA) on cocaine intake in rats trained to self-administer cocaine (0, 125, 250 and 500 microg/infusion) under an FR5 schedule of reinforcement. Separate groups of rats received bilateral intra-VTA injections of the GABA-A antagonist picrotoxin (34 ng/side, n = 7; 68 ng/side, n = 8), GABA-A agonist muscimol (14 ng/side, n = 8), GABA-B agonist baclofen (56 ng/side, n = 7; 100 ng/side, n = 6), picrotoxin (68 ng/side) co-injected with the GABA-B antagonist 2-hydroxysaclofen (100 ng/side, n = 7; 2 microg/side, n = 8) or artificial cerebrospinal fluid (aCSF, n = 6) to assess the effects of the various compounds on the cocaine self-administration dose-response curve. Both picrotoxin and baclofen reduced responding maintained by cocaine, whereas muscimol had no effect on responding. In contrast, neither picrotoxin (n = 6) nor baclofen (n = 8) affected responding maintained by food. Interestingly, 2-hydroxysaclofen effectively blocked the suppression of responding produced by picrotoxin, suggesting that both picrotoxin and baclofen exert their effects via activation of GABA-B receptors. Additionally, these effects appear to be specific to cocaine reinforcement, supporting current investigation of baclofen as a treatment for cocaine addiction.

Contribution of ventral tegmental GABA receptors to cocaine self-administration in rats

PubMed Central

Backes, E.N.; Hemby, S.E.

2008-01-01

Recent evidence has suggested that compounds affecting GABAergic transmission may provide useful pharmacological tools for the treatment of cocaine addiction. Using a rat model of self-administration, the present study examined the effects of GABA agonists and antagonists injected directly into the ventral tegmental area (VTA) on cocaine intake in rats trained to self-administer cocaine (0, 125, 250 and 500 µg/infusion) under an FR5 schedule of reinforcement. Separate groups of rats received bilateral intra-VTA injections of the GABA-A antagonist picrotoxin (34 ng/side, n=7; 68 ng/side, n=8), GABA-A agonist muscimol (14 ng/side, n=8), GABA-B agonist baclofen (56 ng/side, n=7; 100 ng/side, n=6), picrotoxin (68 ng/side) co-injected with the GABA-B antagonist 2-hydroxysaclofen (100 ng/side, n=7; 2 µg/side, n=8) or artificial cerebrospinal fluid (aCSF, n=6) to assess the effects of the various compounds on the cocaine self-administration dose-response curve. Both picrotoxin and baclofen reduced responding maintained by cocaine, whereas muscimol had no effect on responding. In contrast, neither picrotoxin (n=6) nor baclofen (n=8) affected responding maintained by food. Interestingly, 2-hydroxysaclofen effectively blocked the suppression of responding produced by picrotoxin, suggesting that both picrotoxin and baclofen exert their effects via activation of GABA-B receptors. Additionally, these effects appear to be specific to cocaine reinforcement, supporting current investigation of baclofen as a treatment for cocaine addiction. PMID:17943439

Comparison of Cell Expression Formats for the Characterization of GABAA Channels Using a Microfluidic Patch Clamp System

PubMed Central

Chen, Qin; Yim, Peter D.; Yuan, Nina; Johnson, Juliette; Cook, James M.; Smith, Steve; Ionescu-Zanetti, Cristian; Wang, Zhi-Jian; Arnold, Leggy A.

2012-01-01

Abstract Ensemble recording and microfluidic perfusion are recently introduced techniques aimed at removing the laborious nature and low recording success rates of manual patch clamp. Here, we present assay characteristics for these features integrated into one automated electrophysiology platform as applied to the study of GABAA channels. A variety of cell types and methods of GABAA channel expression were successfully studied (defined as IGABA>500 pA), including stably transfected human embryonic kidney (HEK) cells expressing α1β3γ2 GABAA channels, frozen ready-to-assay (RTA) HEK cells expressing α1β3γ2 or α3β3γ2 GABAA channels, transiently transfected HEK293T cells expressing α1β3γ2 GABAA channels, and immortalized cultures of human airway smooth muscle cells endogenously expressing GABAA channels. Current measurements were successfully studied in multiple cell types with multiple modes of channel expression in response to several classic GABAA channel agonists, antagonists, and allosteric modulators. We obtained success rates above 95% for transiently or stably transfected HEK cells and frozen RTA HEK cells expressing GABAA channels. Tissue-derived immortalized cultures of airway smooth muscle cells exhibited a slightly lower recording success rate of 75% using automated patch, which was much higher than the 5% success rate using manual patch clamp technique by the same research group. Responses to agonists, antagonists, and allosteric modulators compared well to previously reported manual patch results. The data demonstrate that both the biophysics and pharmacologic characterization of GABAA channels in a wide variety of cell formats can be performed using this automated patch clamp system. PMID:22574655

α5-GABAA receptors negatively regulate MYC-amplified medulloblastoma growth

PubMed Central

Sengupta, Soma; Weeraratne, Shyamal Dilhan; Sun, Hongyu; Phallen, Jillian; Rallapalli, Sundari K.; Teider, Natalia; Kosaras, Bela; Amani, Vladimir; Pierre-Francois, Jessica; Tang, Yujie; Nguyen, Brian; Yu, Furong; Schubert, Simone; Balansay, Brianna; Mathios, Dimitris; Lechpammer, Mirna; Archer, Tenley C.; Tran, Phuoc; Reimer, Richard J.; Cook, James M.; Lim, Michael; Jensen, Frances E.; Pomeroy, Scott L.; Cho, Yoon-Jae

2013-01-01

Neural tumors often express neurotransmitter receptors as markers of their developmental lineage. Although these receptors have been well characterized in electrophysiological, developmental and pharmacological settings, their importance in the maintenance and progression of brain tumors, and importantly, the effect of their targeting in brain cancers remains obscure. Here, we demonstrate high levels of GABR5, which encodes the α-subunit of the GABAA receptor complex, in aggressive MYC-driven, “Group 3” medulloblastomas. We hypothesized that modulation of α-GABAA receptors alters medulloblastoma cell survival and monitored biological and electrophysiological responses of GABR5-expressing medulloblastoma cells upon pharmacological targeting of the GABAA receptor. While antagonists, inverse agonists and non-specific positive allosteric modulators had limited effects on medulloblastoma cells, a highly specific and potent α5-GABAA receptor agonist, QHii066, resulted in marked membrane depolarization and a significant decrease in cell survival. This effect was GABR5 dependent and mediated through the induction of apoptosis as well as accumulation of cells in S and G2 phases of the cell cycle. Chemical genomic profiling of QHii066-treated medulloblastoma cells confirmed inhibition of MYC-related transcriptional activity and revealed an enrichment of HOX5 target gene expression. siRNA-mediated knockdown of HOX5 markedly blunted the response of medulloblastoma cells to QHii066. Furthermore, QHii066 sensitized GABR5 positive medulloblastoma cells to radiation and chemotherapy consistent with the role of HOX5 in directly regulating p53 expression and inducing apoptosis. Thus, our results provide novel insights into the synthetic lethal nature of α5-GABAA receptor activation in MYC-driven/Group 3 medulloblastomas and propose its targeting as a novel strategy for the management of this highly aggressive tumor. PMID:24196163

GABA receptors in the region of the dorsomedial hypothalamus of rats regulate anxiety in the elevated plus-maze test. II. Physiological measures.

PubMed

Shekhar, A; Sims, L S; Bowsher, R R

1993-11-05

In the previous report, we had shown that blockade and enhancement of GABAA receptors in the DMH of rats increased or decreased the level of anxiety, respectively, as measured by the elevated plus-maze test. The present study was conducted to assess the effects of enhancing GABAA neurotransmission in the DMH of rats on the physiological concomitants of anxiety such as increases in heart rate (HR), blood pressure (BP) and plasma norepinephrine (NE) levels while the animals were placed on the elevated plus-maze. Male Sprague-Dawley rats were equipped with arterial and venous catheters and stereotaxically implanted with microinjection cannulae in the cardiostimulatory region of the DMH where injection of bicuculline methiodide (BMI) elicited increases in heart rate under anesthesia. After recovery, rats were injected with either saline or the GABAA agonist muscimol and their HR, BP and plasma NE responses were measured when confined in the open or the closed arm of the elevated plus-maze. Injection of muscimol into the DMH reduced the increases seen in HR, BP and plasma NE when the rats were confined to either the closed or the open arms in addition to decreasing 'anxiety' in the plus-maze. Injection of muscimol into the areas of the hypothalamus surrounding the DMH did not significantly affect the changes in HR, BP and plasma NE in the plus-maze. Blocking the changes in HR and BP elicited by microinjecting GABAergic drugs into the DMH of rats, with systemic injections of a combination of atropine and the beta-blocker atenolol, did not block the behavioral effects of the GABAergic drugs in the plus-maze test.

Effects of GABA ligands injected into the nucleus accumbens shell on fear/anxiety-like and feeding behaviours in food-deprived rats.

PubMed

Lopes, Ana Paula Fraga; Ganzer, Laís; Borges, Aline Caon; Kochenborger, Larissa; Januário, Ana Cláudia; Faria, Moacir Serralvo; Marino-Neto, José; Paschoalini, Marta Aparecida

2012-03-01

In an attempt to establish a relationship between food intake and fear/anxiety-related behaviours, the goal of this study was to investigate the effect of bilateral injections of GABAA (Muscimol, MUS, doses 25 and 50ng/side) and GABAB (Baclofen, BAC, doses 32 and 64ng/side) receptor agonists in the nucleus accumbens shell (AcbSh) on the level of fear/anxiety-like and feeding behaviours in 24h food-deprived rats. The antagonists of GABAA (Bicuculline, BIC, doses 75 and 150ng/side) and GABAB (Saclofen, SAC, doses 1.5 and 3μg/side) were also tested. The results indicated that the total number of risk assessment behaviour decreased after the injection of both doses of GABAA agonist (MUS) into the AcbSh of 24h food-deprived rats exposed to elevated plus maze. Similar results were obtained after treatment with both doses of GABAB (BAC) agonist in the AcbSh. These data indicated that the activation of both GABAA and GABAB receptors within the AcbSh caused anxiolysis in 24h food-deprived rats. In addition, feeding behaviour (food intake, feeding latency and feeding duration) remained unchanged after treatment with both GABA agonists. In contrast, both food intake and feeding duration decreased after injections of both doses of BIC (GABAA antagonist), while the feeding latency remained unchanged after treatment with both GABA antagonists in the AcbSh of 24h food-deprived rats. The treatment with SAC (GABAB antagonist) did not affect feeding behaviour. Collectively, these data suggest that emotional changes evoked by pharmacological manipulation of the GABA neurotransmission in the AcbSh are not linked with changes in food intake. Copyright © 2011 Elsevier Inc. All rights reserved.

Prefrontal gamma-aminobutyric acid type A receptor insertion controls cue-induced relapse to nicotine seeking.

PubMed

Lubbers, Bart R; van Mourik, Yvar; Schetters, Dustin; Smit, August B; De Vries, Taco J; Spijker, Sabine

2014-11-01

Current smoking cessation therapies offer limited success, as relapse rates remain high. Nicotine, which is the major component of tobacco smoke, is thought to be primarily responsible for the addictive properties of tobacco. However, little is known about the molecular mechanisms underlying nicotine relapse, hampering development of more effective therapies. The objective of this study was to elucidate the role of medial prefrontal cortex (mPFC) glutamatergic and gamma-aminobutyric acid (GABA)ergic receptors in controlling relapse to nicotine seeking. Using an intravenous self-administration model, we studied glutamate and gamma-aminobutyric acid receptor regulation in the synaptic membrane fraction of the rat mPFC following extinction and cue-induced relapse to nicotine seeking. Subsequently, we locally intervened at the level of GABAergic signaling by using a mimetic peptide of the GABA receptor associated protein-interacting domain of GABA type A (GABAA) receptor subunit γ2 (TAT-GABAγ2) and muscimol, a GABAA receptor agonist. Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid and N-methyl-D-aspartate receptors were not regulated after the 30-min relapse test. However, GABAA receptor subunits α1 and γ2 were upregulated, and interference with GABAA receptor insertion in the cell membrane using the TAT-GABAγ2 peptide in the dorsal mPFC, but not the ventral mPFC, significantly increased responding during relapse. Increasing GABAA transmission with muscimol in the dorsal and ventral mPFC attenuated relapse. These data indicate that cue-induced relapse entails a GABAergic plasticity mechanism that limits nicotine seeking by restoring inhibitory control in the dorsal mPFC. GABAA receptor-mediated neurotransmission in the dorsal mPFC constitutes a possible future therapeutic target for maintaining smoking abstinence. Copyright © 2014 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

PubMed

Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

2017-11-15

The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABA A receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABA A receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABA A receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABA A receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABA A receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABA A receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

Nucleus accumbens opioid, GABaergic, and dopaminergic modulation of palatable food motivation: contrasting effects revealed by a progressive ratio study in the rat.

PubMed

Zhang, Min; Balmadrid, Christian; Kelley, Ann E

2003-04-01

The current studies were designed to evaluate whether incentive motivation for palatable food is altered after manipulations of opioid, GABAergic, and dopaminergic transmission within the nucleus accumbens. A progressive ratio schedule was used to measure lever-pressing for sugar pellets after microinfusion of drugs into the nucleus accumbens in non-food-deprived rats. The mu opioid agonist D-Ala2, NMe-Phe4, Glyo15-enkephalin and the indirect dopamine agonist amphetamine induced a marked increase in break point and correct lever-presses; the GABA(A) agonist muscimol did not affect breakpoint or lever-presses. The data suggest that opioid, dopaminergic, and GABAergic systems within the accumbens differentially modulate food-seeking behavior through mechanisms related to hedonic evaluation of food, incentive salience, and control of motor feeding circuits, respectively.

Role of gamma-aminobutyric acid type A (GABAA) receptor subtypes in acute benzodiazepine physical dependence-like effects: evidence from squirrel monkeys responding under a schedule of food presentation

PubMed Central

Fischer, Bradford D.; Teixeira, Laura P.; van Linn, Michael L.; Namjoshi, Ojas A.; Cook, James M.; Rowlett, James K.

2013-01-01

Rationale Assays of schedule-controlled responding can be used to characterize the pharmacology of benzodiazepines and other GABAA receptor modulators, and are sensitive to changes in drug effects that are related to physical dependence. Objective The present study used this approach to investigate the role of GABAA receptor subtypes in mediating dependence-like effects following benzodiazepine administration. Methods Squirrel monkeys (n=6) were trained on a fixed-ratio schedule of food reinforcement. Initially, the response rate-decreasing effects of chlordiazepoxide (0.1–10 mg/kg; nonselective GABAA receptor agonist), zolpidem (0.032–1.0 mg/kg; α1 subunit-containing GABAA subtype-preferring agonist) and HZ-166 (0.1–10 mg/kg; functionally selective α2 and α3 subunit-containing GABAA receptor agonist) were assessed. Next, acute dependence-like effects following single injections of chlordiazepoxide, zolpidem and HZ-166 were assessed with flumazenil (0.1–3.2 mg/kg; nonselective GABAA receptor antagonist). Finally, acute dependence-like effects following zolpidem administration were assessed with βCCt and 3-PBC (0.1–3.2 mg/kg and 0.32–10 mg/kg, respectively; α1 subunit-containing GABAA receptor antagonists). Results Chlordiazepoxide, zolpidem and HZ-166 produced dose- and time-dependent decreases in response rates, whereas flumazenil, βCCt and 3-PBC were ineffective. After the drug effects waned, flumazenil produced dose-dependent decreases in response rates following administration of 10 mg/kg chlordiazepoxide and 1.0 mg/kg zolpidem, but not following any dose of HZ-166. Further, both βCCt and 3-PBC produced dose-dependent decreases in response rates when administered after 1.0 mg/kg zolpidem. Conclusions These data raise the possibility that α1 subunit-containing GABAA receptors play a major role in physical dependence-related behaviors following a single injection of a benzodiazepine. PMID:23354533

Role of gamma-aminobutyric acid type A (GABAA) receptor subtypes in acute benzodiazepine physical dependence-like effects: evidence from squirrel monkeys responding under a schedule of food presentation.

PubMed

Fischer, Bradford D; Teixeira, Laura P; van Linn, Michael L; Namjoshi, Ojas A; Cook, James M; Rowlett, James K

2013-05-01

Assays of schedule-controlled responding can be used to characterize the pharmacology of benzodiazepines and other GABAA receptor modulators, and are sensitive to changes in drug effects that are related to physical dependence. The present study used this approach to investigate the role of GABAA receptor subtypes in mediating dependence-like effects following benzodiazepine administration. Squirrel monkeys (n = 6) were trained on a fixed-ratio schedule of food reinforcement. Initially, the response rate-decreasing effects of chlordiazepoxide (0.1-10 mg/kg; nonselective GABAA receptor agonist), zolpidem (0.032-1.0 mg/kg; α1 subunit-containing GABAA subtype-preferring agonist), and HZ-166 (0.1-10 mg/kg; functionally selective α2 and α3 subunit-containing GABAA receptor agonist) were assessed. Next, acute dependence-like effects following single injections of chlordiazepoxide, zolpidem, and HZ-166 were assessed with flumazenil (0.1-3.2 mg/kg; nonselective GABAA receptor antagonist). Finally, acute dependence-like effects following zolpidem administration were assessed with βCCt and 3-PBC (0.1-3.2 mg/kg and 0.32-10 mg/kg, respectively; α1 subunit-containing GABAA receptor antagonists). Chlordiazepoxide, zolpidem, and HZ-166 produced dose- and time-dependent decreases in response rates, whereas flumazenil, βCCT, and 3-PBC were ineffective. After the drug effects waned, flumazenil produced dose-dependent decreases in response rates following administration of 10 mg/kg chlordiazepoxide and 1.0 mg/kg zolpidem, but not following any dose of HZ-166. Further, both βCCT and 3-PBC produced dose-dependent decreases in response rates when administered after 1.0 mg/kg zolpidem. These data raise the possibility that α1 subunit-containing GABAA receptors play a major role in physical dependence-related behaviors following a single injection of a benzodiazepine.

Acutely increasing δGABAA receptor activity impairs memory and inhibits synaptic plasticity in the hippocampus

PubMed Central

Whissell, Paul D.; Eng, Dave; Lecker, Irene; Martin, Loren J.; Wang, Dian-Shi; Orser, Beverley A.

2013-01-01

Extrasynaptic γ-aminobutyric acid type A (GABAA) receptors that contain the δ subunit (δGABAA receptors) are expressed in several brain regions including the dentate gyrus (DG) and CA1 subfields of the hippocampus. Drugs that increase δGABAA receptor activity have been proposed as treatments for a variety of disorders including insomnia, epilepsy and chronic pain. Also, long-term pretreatment with the δGABAA receptor–preferring agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) enhances discrimination memory and increases neurogenesis in the DG. Despite the potential therapeutic benefits of such treatments, the effects of acutely increasing δGABAA receptor activity on memory behaviors remain unknown. Here, we studied the effects of THIP (4 mg/kg, i.p.) on memory performance in wild-type (WT) and δGABAA receptor null mutant (Gabrd−/−) mice. Additionally, the effects of THIP on long-term potentiation (LTP), a molecular correlate of memory, were studied within the DG and CA1 subfields of the hippocampus using electrophysiological recordings of field potentials in hippocampal slices. The results showed that THIP impaired performance in the Morris water maze, contextual fear conditioning and object recognition tasks in WT mice but not Gabrd−/− mice. Furthermore, THIP inhibited LTP in hippocampal slices from WT but not Gabrd−/− mice, an effect that was blocked by GABAA receptor antagonist bicuculline. Thus, acutely increasing δGABAA receptor activity impairs memory behaviors and inhibits synaptic plasticity. These results have important implications for the development of therapies aimed at increasing δGABAA receptor activity. PMID:24062648

Menthol enhances phasic and tonic GABAA receptor-mediated currents in midbrain periaqueductal grey neurons

PubMed Central

Lau, Benjamin K; Karim, Shafinaz; Goodchild, Ann K; Vaughan, Christopher W; Drew, Geoffrey M

2014-01-01

Background and Purpose Menthol, a naturally occurring compound in the essential oil of mint leaves, is used for its medicinal, sensory and fragrant properties. Menthol acts via transient receptor potential (TRPM8 and TRPA1) channels and as a positive allosteric modulator of recombinant GABAA receptors. Here, we examined the actions of menthol on GABAA receptor-mediated currents in intact midbrain slices. Experimental Approach Whole-cell voltage-clamp recordings were made from periaqueductal grey (PAG) neurons in midbrain slices from rats to determine the effects of menthol on GABAA receptor-mediated phasic IPSCs and tonic currents. Key Results Menthol (150–750 μM) produced a concentration-dependent prolongation of spontaneous GABAA receptor-mediated IPSCs, but not non-NMDA receptor-mediated EPSCs throughout the PAG. Menthol actions were unaffected by TRPM8 and TRPA1 antagonists, tetrodotoxin and the benzodiazepine antagonist, flumazenil. Menthol also enhanced a tonic current, which was sensitive to the GABAA receptor antagonists, picrotoxin (100 μM), bicuculline (30 μM) and Zn2+ (100 μM), but unaffected by gabazine (10 μM) and a GABAC receptor antagonist, 1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid hydrate (TPMPA; 50 μM). In addition, menthol potentiated currents induced by the extrasynaptic GABAA receptor agonist THIP/gaboxadol (10 μM). Conclusions and Implications These results suggest that menthol positively modulates both synaptic and extrasynaptic populations of GABAA receptors in native PAG neurons. The development of agents that potentiate GABAA-mediated tonic currents and phasic IPSCs in a manner similar to menthol could provide a basis for novel GABAA-related pharmacotherapies. PMID:24460753

Role of the anterior cingulate cortex in the retrieval of novel object recognition memory after a long delay

PubMed Central

Pezze, Marie A.; Marshall, Hayley J.; Fone, Kevin CF.; Cassaday, Helen J.

2017-01-01

Previous in vivo electrophysiological studies suggest that the anterior cingulate cortex (ACgx) is an important substrate of novel object recognition (NOR) memory. However, intervention studies are needed to confirm this conclusion and permanent lesion studies cannot distinguish effects on encoding and retrieval. The interval between encoding and retrieval tests may also be a critical determinant of the role of the ACgx. The current series of experiments used micro-infusion of the GABAA receptor agonist, muscimol, into ACgx to reversibly inactivate the area and distinguish its role in encoding and retrieval. ACgx infusions of muscimol, before encoding did not alter NOR assessed after a delay of 20 min or 24 h. However, when infused into the ACgx before retrieval muscimol impaired NOR assessed after a delay of 24 h, but not after a 20-min retention test. Together these findings suggest that the ACgx plays a time-dependent role in the retrieval, but not the encoding, of NOR memory, neuronal activation being required for the retrieval of remote (24 h old), but not recent (20 min old) visual memory. PMID:28620078

Pharmacodynamic and pharmacokinetic effects of MK-0343, a GABA(A) alpha2,3 subtype selective agonist, compared to lorazepam and placebo in healthy male volunteers.

PubMed

de Haas, S L; de Visser, S J; van der Post, J P; Schoemaker, R C; van Dyck, K; Murphy, M G; de Smet, M; Vessey, L K; Ramakrishnan, R; Xue, L; Cohen, A F; van Gerven, J M A

2008-01-01

The use of non-selective gamma-aminobutyric acid (GABA) enhancers, such as benzodiazepines in the treatment of anxiety disorders is still widespread but hampered by unfavourable side effects. some of these may be associated with binding properties to certain subtypes of the GABA(A) receptor that are unnecessary for therapeutic effects. MK-0343 was designed to be a less sedating anxiolytic, based on reduced efficacy at the alpha1 subtype and significant efficacy at alpha2 and alpha3 subtypes of the GABA(A) receptor. This paper is a double-blind, four-way cross-over (n = 12) study to investigate the effects of MK-0343 (0.25 and 0.75 mg) in comparison to placebo and an anxiolytic dose (2 mg) of the non-selective agonist lorazepam. Effects were measured by eye movements, body sway, Visual Analogue scales (VAS) and memory tests. Lorazepam impaired saccadic peak velocity (SPV), VAs alertness scores, postural stability and memory and increased saccadic latency and inaccuracy. MK-0343 0.75 mg was equipotent with lorazepam as indicated by SPV (-42.4 deg/s), saccadic latency (0.02 s) and VAS alertness scores (1.50 ln mm), while effects on memory and postural stability were smaller. MK-0343 0.25 mg only affected postural stability to a similar extent as MK-0343 0.75 mg. The effect profile of MK-0343 0.75 mg is different from the full agonist lorazepam, which could reflect the selective actions of this compound. Although less effect on VAS alertness was expected, diminished effects on memory and postural stability were present. Clinical studies in anxiety patients should show whether this dose of MK-0343 is therapeutically effective with a different side-effect profile.

Hypothalamic GABAergic influences on treadmill exercise responses in rats.

PubMed

Overton, J M; Redding, M W; Yancey, S L; Stremel, R W

1994-01-01

Microinjection of GABAergic antagonists in the posterior hypothalamus (PH) produces exercise-like adjustments in cardiovascular function. To test the hypothesis that a hypothalamic GABAergic mechanism within the PH modulates the cardiovascular adjustments to dynamic exercise in conscious animals, Sprague-Dawley rats (n = 10) were instrumented with bilateral guide cannula directed at the pH, an arterial cannula, and Doppler flow probes on the iliac and mesenteric arteries. Saline (100 nl) or the GABAA receptor agonist muscimol (125 ng.100 nl-1) was bilaterally injected into the PH during treadmill exercise (20 m.min-1). Microinjection of saline had no effect on mean arterial pressure (MAP), heart rate (HR), mesenteric vascular resistance (MR), or iliac vascular resistance (IR) during exercise. Microinjection of muscimol during exercise produced no significant changes in MAP (mean change +/- SE; +0 +/- 1 mmHg), HR (+17 +/- 12 b.min-1), or MR (+7 +/- 13%). However, microinjection of muscimol produced a significant increase in IR during exercise (16 +/- 6%). In addition, muscimol significantly decreased treadmill run time (saline = 19.6 +/- 0.4 min; muscimol = 17.8 +/- 0.6 min) and produced behavioral effects (including mild sedation) that were most evident after exercise. The results of these experiments suggest that while the posterior hypothalamic GABAergic system may modulate iliac blood flow during exercise in rats, this system does not modulate HR and MR responses to dynamic exercise.

Relative positioning of classical benzodiazepines to the γ2-subunit of GABAA receptors.

PubMed

Middendorp, Simon J; Hurni, Evelyn; Schönberger, Matthias; Stein, Marco; Pangerl, Michael; Trauner, Dirk; Sigel, Erwin

2014-08-15

GABAA receptors are the major inhibitory neurotransmitter receptors in the brain. Benzodiazepine exert their action via a high affinity-binding site at the α/γ subunit interface on some of these receptors. Diazepam has sedative, hypnotic, anxiolytic, muscle relaxant, and anticonvulsant effects. It acts by potentiating the current evoked by the agonist GABA. Understanding specific interaction of benzodiazepines in the binding pocket of different GABAA receptor isoforms might help to separate these divergent effects. As a first step, we characterized the interaction between diazepam and the major GABAA receptor isoform α1β2γ2. We mutated several amino acid residues on the γ2-subunit assumed to be located near or in the benzodiazepine binding pocket individually to cysteine and studied the interaction with three ligands that are modified with a cysteine-reactive isothiocyanate group (-NCS). When the reactive NCS group is in apposition to the cysteine residue this leads to a covalent reaction. In this way, three amino acid residues, γ2Tyr58, γ2Asn60, and γ2Val190 were located relative to classical benzodiazepines in their binding pocket on GABAA receptors.

Sex dimorphism in seizure-controlling networks.

PubMed

Giorgi, Fillippo Sean; Galanopoulou, Aristea S; Moshé, Solomon L

2014-12-01

Males and females show a different predisposition to certain types of seizures in clinical studies. Animal studies have provided growing evidence for sexual dimorphism of certain brain regions, including those that control seizures. Seizures are modulated by networks involving subcortical structures, including thalamus, reticular formation nuclei, and structures belonging to the basal ganglia. In animal models, the substantia nigra pars reticulata (SNR) is the best studied of these areas, given its relevant role in the expression and control of seizures throughout development in the rat. Studies with bilateral infusions of the GABA(A) receptor agonist muscimol have identified distinct roles of the anterior or posterior rat SNR in flurothyl seizure control, that follow sex-specific maturational patterns during development. These studies indicate that (a) the regional functional compartmentalization of the SNR appears only after the third week of life, (b) only the male SNR exhibits muscimol-sensitive proconvulsant effects which, in older animals, is confined to the posterior SNR, and (c) the expression of the muscimol-sensitive anticonvulsant effects become apparent earlier in females than in males. The first three postnatal days are crucial in determining the expression of the muscimol-sensitive proconvulsant effects of the immature male SNR, depending on the gonadal hormone setting. Activation of the androgen receptors during this early period seems to be important for the formation of this proconvulsant SNR region. We describe molecular/anatomical candidates underlying these age- and sex-related differences, as derived from in vitro and in vivo experiments, as well as by [(14)C]2-deoxyglucose autoradiography. These involve sex-specific patterns in the developmental changes in the structure or physiology or GABA(A) receptors or of other subcortical structures (e.g., locus coeruleus, hippocampus) that may affect the function of seizure-controlling networks

Role of ventral medial prefrontal cortex in incubation of cocaine craving

PubMed Central

Koya, Eisuke; Uejima, Jamie L.; Wihbey, Kristina A.; Bossert, Jennifer M.; Hope, Bruce T.; Shaham, Yavin

2009-01-01

Cue-induced drug-seeking in rodents progressively increases after withdrawal from cocaine, suggesting that cue-induced cocaine craving incubates over time. Here, we explored the role of the medial prefrontal cortex (mPFC, a brain area previously implicated in cue-induced cocaine seeking) in this incubation. We trained rats to self-administer cocaine for 10 d (6 h/d, infusions were paired with a tone-light cue), and then assessed after 1 or 30 withdrawal days the effect of exposure to cocaine cues on lever presses in extinction tests. We found that cue-induced cocaine-seeking in the extinction tests was higher after 30 withdrawal days than after 1 day. The time-dependent increases in extinction responding were associated with large (ventral mPFC) or modest (dorsal mPFC) increases in ERK phosphorylation (a measure of ERK activity and an index of neuronal activation). After 30 withdrawal days, ventral but not dorsal injections of muscimol+baclofen (GABAa+GABAb receptor agonists that inhibit neuronal activity) decreased extinction responding. After 1 withdrawal day, ventral but not dorsal mPFC injections of bicuculline+saclofen (GABAa+GABAb receptor antagonists that increase neuronal activity) strongly increased extinction responding. Finally, muscimol+baclofen had minimal effect on extinction responding after 1 day, and in cocaine-experienced rats, ventral mPFC injections of muscimol+baclofen or bicuculline+saclofen had no effect on lever presses for an oral sucrose solution. The present results indicate that ventral mPFC neuronal activity plays an important role in the incubation of cocaine craving. PMID:18565549

Medial prefrontal cortex serotonergic and GABAergic mechanisms modulate the expression of contextual fear: intratelencephalic pathways and differential involvement of cortical subregions.

PubMed

Almada, R C; Coimbra, N C; Brandão, M L

2015-01-22

Several lines of evidence indicate that the dorsal hippocampus (dH) and medial prefrontal cortex (mPFC) regulate contextual fear conditioning. The prelimbic (PrL), infralimbic (IL) and the anterior cingulate cortex (ACC) subregions of the mPFC likely play distinct roles in the expression of fear. Moreover, studies have highlighted the role of serotonin (5-hydroxytryptamine, 5-HT)- and γ-aminobutyric acid (GABA)-mediated mechanisms in the modulation of innate fear in the mPFC. The present study characterized dH-mPFC pathways and investigated the role of serotonergic and GABAergic mechanisms of the PrL, IL and ACC-area 1 (Cg1) in the elaboration of contextual fear conditioning using fear-potentiated startle (FPS) and freezing behavior in Rattus norvegicus. The results of neurotracing with microinjections of biotinylated dextran amine into the dH revealed a neural link of the dH with the PrL and ACC. Intra-PrL injections of the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) and the GABAA receptor-selective agonist muscimol reduced contextual FPS and freezing responses. Intra-Cg1 injections of muscimol but not 8-OH-DPAT decreased FPS and freezing responses. However, neither intra-IL injections of a 5-HT1A agonist nor of a GABAA agonist affected these defensive responses. Labeled neuronal fibers from the dH reached the superficial layers of the PrL cortex and spread to the inner layers of PrL and Cg1 cortices, supporting the pharmacological findings. The present results confirmed the involvement of PrL and Cg1 in the expression of FPS and freezing responses to aversive conditions. In addition, PrL serotoninergic mechanisms play a key role in contextual fear conditioning. This study suggests that PrL, IL and Cg1 distinctively contribute to the modulation of contextual fear conditioning. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

Analysis of responses to valerian root extract in the feline pulmonary vascular bed.

PubMed

Fields, Aaron M; Richards, Todd A; Felton, Jason A; Felton, Shaili K; Bayer, Erin Z; Ibrahim, Ikhlass N; Kaye, Alan David

2003-12-01

This study was undertaken to investigate pulmonary vascular response to valerian (Valeriana officinalis) in the feline pulmonary vasculature under constant flow conditions. In separate experiments, the effects of NG-L-nitro-L-arginine methyl ester (L-NIO), a nitric oxide synthase inhibitor, glibenclamide, an adenosine triphosphate (ATP)-sensitive potassium (K+) channel blocker, meclofenamate, a nonselective cyclooxygenase (COX) inhibitor, bicuculline, a GABA(A) receptor antagonist, and saclofen, a GABA(B) antagonist, were investigated on pulmonary arterial responses to various agonists in the feline pulmonary vascular bed. These agonists included valerian, muscimol, a GABA(A) agonist, SKF-97541 a GABA(B) agonist, acetylcholine (ACh), and bradykinin, both inducers of nitric oxide synthase, arachidonic acid, a COX substrate, and pinacidil, an ATP-sensitive K+ channel activator, during increased tone conditions induced by the thromboxane A2 mimic, U46619. Laboratory investigation. Mongrel cats of either gender. Injections of the abovementioned agonists and antagonists were given. Baseline pulmonary tone, responses to the agonists, and responses to the agonists after injections of antagonists were all measured via a pulmonary catheter transducer and recorded. Valerian root extract is a potent smooth muscle dilator in the feline pulmonary vascular bed. The vasodilatory effects of valerian root extract were unchanged after the administration of L-NIO, glibenclamide, and meclofenamate. These effects were ablated, however, by both saclofen and bicuculline. The ability of saclofen and bicuculline to modulate the dilatory effects of valerian root extract was not statistically different. The vasodilatory effects of valerian root extract are mediated by a nonselective GABA mechanism.

Synergistic effect between D-AP5 and muscimol in the nucleus accumbens shell on memory consolidation deficit in adult male Wistar rats: An isobologram analysis.

PubMed

Nasehi, Mohammad; Ostadi, Elaheh; Khakpai, Fatemeh; Ebrahimi-Ghiri, Mohaddeseh; Zarrindast, Mohammad-Reza

2017-05-01

The nucleus accumbens (NAc) glutamatergic and GABAergic systems are involved in memory processes. This study was investigated the involvement of NAc shell GABAergic system on D-AP5 induced memory consolidation deficit. The elevated plus-maze (EPM) test-retest paradigm was employed to assess memory in adult male Wistar rats. The results indicated that post-training intra-NAc shell injection of bicuculline (GABA A receptor antagonist) did not alter emotional memory consolidation. However, post-training intra-NAc shell microinjection of muscimol (GABA A receptor agonist, 0.1μg/rat) and D-AP5 (a competitive NMDA receptor antagonist, 4μg/rat) decreased emotional memory consolidation, suggesting the drugs induced amnesia. Moreover, a sub-threshold dose of muscimol (0.05μg/rat) potentiated the D-AP5 (2μg/rat) response on memory consolidation impairment. On the other hand, the middle dose of bicuculline (0.25μg/rat) reversed memory impairment induced by D-AP5 at the higher dose. Interestingly, there is a synergistic effect between D-AP5 and muscimol on impairment of emotional memory consolidation. None of the above doses changed the locomotor activity. Our results suggest that the glutamatergic and GABAergic neurons of the NAc shell interact with each other for modulation of emotional memory consolidation. Copyright © 2017 Elsevier Inc. All rights reserved.

Modulation of GABAergic receptor binding by activation of calcium and calmodulin-dependent kinase II membrane phosphorylation.

PubMed

Churn, S B; DeLorenzo, R J

1998-10-26

gamma-Aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the central nervous system (CNS). Because of the important role that GABA plays in the CNS, alteration of GABAA receptor function would significantly affect neuronal excitability. Protein phosphorylation is a major mechanism for regulating receptor function in the brain and has been implicated in modulating GABAA receptor function. Therefore, this study was initiated to determine the role of calmodulin-dependent kinase II (CaM kinase II) membrane phosphorylation on GABAA receptor binding. Synaptosomal membrane fractions were tested for CaM kinase II activity towards endogenous substrates. In addition, muscimol binding was evaluated under equilibrium conditions in synaptosomal membrane fractions subjected to either basal (Mg2+ alone) or maximal CaM kinase II-dependent phosphorylation. Activation of endogenous CaM kinase II-dependent phosphorylation resulted in a significant enhancement of the apparent Bmax for muscimol binding without significantly altering the apparent binding affinity. The enhanced muscimol binding could be increased further by the addition of exogenous CaM kinase II to synaptosomal membrane fractions. Co-incubation with inhibitors of kinase activity during the phosphorylation reactions blocked the CaM kinase II-dependent increase in muscimol binding. The data support the hypothesis that activation of CaM kinase II-dependent phosphorylation caused an increased GABAA receptor binding and may play an important role in modulating the function of this inhibitory receptor/chloride ion channel complex. Copyright 1998 Elsevier Science B.V.

The effects of GABAA and NMDA receptors in the shell-accumbens on spatial memory of METH-treated rats.

PubMed

Heysieattalab, Soomaayeh; Naghdi, Nasser; Zarrindast, Mohammad-Reza; Haghparast, Abbas; Mehr, Shahram Ejtemaei; Khoshbouei, Habibeh

2016-03-01

Methamphetamine (METH) is a highly addictive and neurotoxic psychostimulant. Its use in humans is often associated with neurocognitive impairment and deficits in hippocampal plasticity. Striatal dopamine system is one of the main targets of METH. The dopamine neurons in the striatum directly or indirectly regulate the GABA and glutamatergic signaling in this region and thus their outputs. This is consistent with previous reports showing modification of neuronal activity in the striatum modulates the expression of hippocampal LTP and hippocampal-dependent memory tasks such as Morris water maze (MWM). Therefore, reversing or preventing METH-induced synaptic modifications via pharmacological manipulations of the shell-nucleus accumbens (shell-NAc) may introduce a viable therapeutic target to attenuate the METH-induced memory deficits. This study is designed to investigate the role of intra-shell NAc manipulation of GABAA and NMDA receptors and their interaction with METH on memory performance in MWM task. Pharmacological manipulations were performed in rats received METH or saline. We found systemic saline plus intra-shell NAc infusions of muscimol dose-dependently impaired performance, while bicuculline had no effect. Surprisingly, the intra-NAc infusions of 0.005μg/rat muscimol that has no effect on memory performance (ineffective dose) prevented METH-induced memory impairment. In the contrary, the intra-NAc infusions of bicuculline (0.2μg/rat) increased METH-induced memory impairment. However, pre-training intra-NAc infusions of D-AP5 dose-dependently impaired performance, while NMDA had no effect in rats received systemic saline (control group). The intra-NAc infusions with an ineffective dose of NMDA (0.1μg/rat) increased METH-induced memory impairment. Furthermore, intra-NAc infusions of D-AP5 with an ineffective dose (0.1μg/rat) prevented METH-induced memory impairment. Our result is consistent with the interpretation that METH-mediated learning deficit

NOVEL POSITIVE ALLOSTERIC MODULATORS OF GABAA RECEPTORS: DO SUBTLE DIFFERENCES IN ACTIVITY AT α1 PLUS α5 VERSUS α2 PLUS α3 SUBUNITS ACCOUNT FOR DISSIMILARITIES IN BEHAVIORAL EFFECTS IN RATS?

PubMed Central

Savić, Miroslav M.; Majumder, Samarpan; Huang, Shengming; Edwankar, Rahul V.; Furtmüller, Roman; Joksimović, Srđan; Clayton, Terry; Ramerstorfer, Joachim; Milinković, Marija M.; Roth, Bryan L.; Sieghart, Werner; Cook, James M.

2010-01-01

Over the last years, genetic studies have greatly improved our knowledge on the receptor subtypes mediating various pharmacological effects of positive allosteric modulators at GABAA receptors. This stimulated the development of new benzodiazepine (BZ)-like ligands, especially those inactive/low-active at GABAA receptors containing the α1 subunit, with the aim of generating more selective drugs. Hereby, the affinity and efficacy of four recently-synthesized BZ site ligands: SH-053-2’N, SH-053-S-CH3-2’F, SH-053-R-CH3-2’F and JY-XHe-053 were assessed. They were also studied in behavioral tests of spontaneous locomotor activity, elevated plus maze, and water maze in rats, which are considered predictive of, respectively, the sedative, anxiolytic, and amnesic influence of BZs. The novel ligands had moderately low to low affinity and mild to partial agonistic efficacy at GABAA receptors containing the α1 subunit, with variable, but more pronounced efficacy at other BZ-sensitive binding sites. While presumably α1 receptor-mediated sedative effects of GABAA modulation were not fully eliminated with any of the ligands tested, only SH-053-2’N and SH-053-S-CH3-2’F, both dosed at 30 mg/kg, exerted anxiolytic effects. The lack of clear anxiolytic-like activity of JY-XHe-053, despite its efficacy at α2- and α3-GABAA receptors, may have been partly connected with its preferential affinity at α5-GABAA receptors coupled with weak agonist activity at α1-containing subtypes. The memory impairment in water-maze experiments, generally reported with BZ site agonists, was completely circumvented with all four ligands. The results suggest that a substantial amount of activity at α1 GABAA receptors is needed for effecting spatial learning and memory impairments, while much weaker activity at α1- and α5-GABAA receptors is sufficient for eliciting sedation. PMID:20074611

A Review of the Updated Pharmacophore for the Alpha 5 GABA(A) Benzodiazepine Receptor Model

PubMed Central

Clayton, Terry; Poe, Michael M.; Rallapalli, Sundari; Biawat, Poonam; Savić, Miroslav M.; Rowlett, James K.; Gallos, George; Emala, Charles W.; Kaczorowski, Catherine C.; Stafford, Douglas C.; Arnold, Leggy A.; Cook, James M.

2015-01-01

An updated model of the GABA(A) benzodiazepine receptor pharmacophore of the α5-BzR/GABA(A) subtype has been constructed prompted by the synthesis of subtype selective ligands in light of the recent developments in both ligand synthesis, behavioral studies, and molecular modeling studies of the binding site itself. A number of BzR/GABA(A) α5 subtype selective compounds were synthesized, notably α5-subtype selective inverse agonist PWZ-029 (1) which is active in enhancing cognition in both rodents and primates. In addition, a chiral positive allosteric modulator (PAM), SH-053-2′F-R-CH3 (2), has been shown to reverse the deleterious effects in the MAM-model of schizophrenia as well as alleviate constriction in airway smooth muscle. Presented here is an updated model of the pharmacophore for α5β2γ2 Bz/GABA(A) receptors, including a rendering of PWZ-029 docked within the α5-binding pocket showing specific interactions of the molecule with the receptor. Differences in the included volume as compared to α1β2γ2, α2β2γ2, and α3β2γ2 will be illustrated for clarity. These new models enhance the ability to understand structural characteristics of ligands which act as agonists, antagonists, or inverse agonists at the Bz BS of GABA(A) receptors. PMID:26682068

A novel positive allosteric modulator of the GABAA receptor: the action of (+)-ROD188

PubMed Central

Thomet, Urs; Baur, Roland; Razet, Rodolphe; Dodd, Robert H; Furtmüller, Roman; Sieghart, Werner; Sigel, Erwin

2000-01-01

(+)-ROD188 was synthesized in the search for novel ligands of the GABA binding site. It shares some structural similarity with bicuculline. (+)-ROD188 failed to displace [3H]-muscimol in binding studies and failed to induce channel opening in recombinant rat α1β2γ2 GABAA receptors functionally expressed in Xenopus oocytes. (+)-ROD188 allosterically stimulated GABA induced currents. Displacement of [3H]-Ro15-1788 indicated a low affinity action at the benzodiazepine binding site. In functional studies, stimulation by (+)-ROD188 was little sensitive to the presence of 1 μM of the benzodiazepine antagonist Ro 15-1788, and (+)-ROD188 also stimulated currents mediated by α1β2, indicating a major mechanism of action different from that of benzodiazepines. Allosteric stimulation by (+)-ROD188 was similar in α1β2N265S as in unmutated α1β2, while that by loreclezole was strongly reduced. (+)-ROD188 also strongly stimulated currents elicited by either pentobarbital or 5α-pregnan-3α-ol-20-one (3α-OH-DHP), in line with a mode of action different from that of barbiturates or neurosteroids as channel agonists. Stimulation by (+)-ROD188 was largest in α6β2γ2 (α6β2γ2>>α1β2γ2=α5β2γ2>α2β2γ2= α3β2γ2), indicating a unique subunit isoform specificity. Miniature inhibitory postsynaptic currents (mIPSC) in cultures of rat hippocampal neurons, caused by spontaneous release of GABA showed a prolonged decay time in the presence of 30 μM (+)-ROD188, indicating an enhanced synaptic inhibitory transmission. PMID:11030736

Dual Modulators of GABA-A and Alpha7 Nicotinic Receptors for Treating Autism

DTIC Science & Technology

2014-08-01

Synthesis of 2-261, AVL-3288 & GRN-529. This task was accomplished in December, 2013. The task was accomplished one month later than predicted in the...approved SOW because of the need to synthesize some of the starting materials that were commercially unavailable for the synthesis of the compounds...Interestingly recent studies with the benzodiazepine agonist clonazepam , a non-selective GABAA receptor PAM, resulted in a bell-shaped dose response

Endogenous gamma-aminobutyric acid modulates tonic guinea pig airway tone and propofol-induced airway smooth muscle relaxation.

PubMed

Gallos, George; Gleason, Neil R; Virag, Laszlo; Zhang, Yi; Mizuta, Kentaro; Whittington, Robert A; Emala, Charles W

2009-04-01

Emerging evidence indicates that an endogenous autocrine/paracrine system involving gamma-aminobutyric acid (GABA) is present in airways. GABAA channels, GABAB receptors, and the enzyme that synthesizes GABA have been identified in airway epithelium and smooth muscle. However, the endogenous ligand itself, GABA, has not been measured in airway tissues. The authors sought to demonstrate that GABA is released in response to contractile agonists and tonically contributes a prorelaxant component to contracted airway smooth muscle. The amount and cellular localization of GABA in upper guinea pig airways under resting and contracted tone was determined by high pressure liquid chromatography and immunohistochemistry, respectively. The contribution that endogenous GABA imparts on the maintenance of airway smooth muscle acetylcholine-induced contraction was assessed in intact guinea pig airway tracheal rings using selective GABAA antagonism (gabazine) under resting or acetylcholine-contracted conditions. The ability of an allosteric agent (propofol) to relax a substance P-induced relaxation in an endogenous GABA-dependent manner was assessed. GABA levels increased and localized to airway smooth muscle after contractile stimuli in guinea pig upper airways. Acetylcholine-contracted guinea pig tracheal rings exhibited an increase in contracted force upon addition of the GABAA antagonist gabazine that was subsequently reversed by the addition of the GABAA agonist muscimol. Propofol dose-dependently relaxed a substance P contraction that was blocked by gabazine. These studies demonstrate that GABA is endogenously present and increases after contractile stimuli in guinea pig upper airways and that endogenous GABA contributes a tonic prorelaxant component in the maintenance of airway smooth muscle tone.

Endogenous γ-aminobutyric Acid Modulates Tonic Guinea Pig Airway Tone and Propofol-induced Airway Smooth Muscle Relaxation

PubMed Central

Gallos, George; Gleason, Neil R.; Virag, Laszlo; Zhang, Yi; Mizuta, Kentauro; Whittington, Robert A.; Emala, Charles W.

2009-01-01

Background Emerging evidence indicates that an endogenous autocrine/paracrine system involving γ-aminobutyric acid (GABA) is present in airways. GABAA channels, GABAB receptors and the enzyme that synthesizes GABA have been identified in airway epithelium and smooth muscle. However, the endogenous ligand itself, GABA, has not been measured in airway tissues. We sought to demonstrate that GABA is released in response to contractile agonists and tonically contributes a pro-relaxant component to contracted airway smooth muscle. Methods The amount and cellular localization of GABA in upper guinea pig airways under resting and contracted tone was determined by high pressure liquid chromatography and immunohistochemistry, respectively. The contribution that endogenous GABA imparts on the maintenance of airway smooth muscle acetylcholine-induced contraction was assessed in intact guinea pig airway tracheal rings using selective GABAA antagonism (gabazine) under resting or acetylcholine-contracted conditions. The ability of an allosteric agent (propofol) to relax a substance P-induced relaxation in an endogenous GABA-dependent manner was assessed. Results GABA levels increased and localized to airway smooth muscle following contractile stimuli in guinea pig upper airways. Acetylcholine-contracted guinea pig tracheal rings exhibited an increase in contracted force upon addition of the GABAA antagonist gabazine which was subsequently reversed by the addition of the GABAA agonist muscimol. Propofol dose-dependently relaxed a substance P contraction that was blocked by gabazine. Conclusion These studies demonstrate that GABA is endogenously present and increases following contractile stimuli in guinea pig upper airways and that endogenous GABA contributes a tonic pro-relaxant component in the maintenance of airway smooth muscle tone. PMID:19322939

GABA-independent GABAA Receptor Openings Maintain Tonic Currents

PubMed Central

Wlodarczyk, Agnieszka I.; Sylantyev, Sergiy; Herd, Murray B.; Kersanté, Flavie; Lambert, Jeremy J.; Rusakov, Dmitri A.; Linthorst, Astrid C.E.; Semyanov, Alexey; Belelli, Delia; Pavlov, Ivan; Walker, Matthew C.

2013-01-01

Activation of GABAA receptors (GABAARs) produces two forms of inhibition: ‘phasic’ inhibition generated by the rapid, transient activation of synaptic GABAARs by presynaptic GABA release, and tonic inhibition generated by the persistent activation of peri- or extrasynaptic GABAARs which can detect extracellular GABA. Such tonic GABAAR-mediated currents are particularly evident in dentate granule cells in which they play a major role in regulating cell excitability. Here we show that in rat dentate granule cells in ex-vivo hippocampal slices, tonic currents are predominantly generated by GABA-independent GABAA receptor openings. This tonic GABAAR conductance is resistant to the competitive GABAAR antagonist SR95531, which at high concentrations acts as a partial agonist, but can be blocked by an open channel blocker picrotoxin. When slices are perfused with 200 nM GABA, a concentration that is comparable to cerebrospinal fluid concentrations but is twice that measured by us in the hippocampus in vivo using zero-net-flux microdialysis, negligible GABA is detected by dentate granule cells. Spontaneously opening GABAARs, therefore, maintain dentate granule cell tonic currents in the face of low extracellular GABA concentrations. PMID:23447601

Pharmacodynamic and pharmacokinetic effects of TPA023, a GABA(A) alpha(2,3) subtype-selective agonist, compared to lorazepam and placebo in healthy volunteers.

PubMed

de Haas, S L; de Visser, S J; van der Post, J P; de Smet, M; Schoemaker, R C; Rijnbeek, B; Cohen, A F; Vega, J M; Agrawal, N G B; Goel, T V; Simpson, R C; Pearson, L K; Li, S; Hesney, M; Murphy, M G; van Gerven, J M A

2007-06-01

TPA023, a GABA(A) alpha2,3 alphasubtype-selective partial agonist, is expected to have comparable anxiolytic efficacy as benzodiazepines with reduced sedating effects. The compound lacks efficacy at the alpha1 subtype, which is believed to mediate these effects. This study investigated the effects of 0.5 and 1.5 mg TPA023 and compared them with placebo and lorazepam 2 mg (therapeutic anxiolytic dose). Twelve healthy male volunteers participated in this placebo-controlled, double-blind, double-dummy, four-way, cross-over study. Saccadic eye movements and visual analogue scales (VAS) were used to assess the sedative properties of TPA023. The effects on posturaL stability and cognition were assessed using body sway and a standardized battery of neurophysiological memory tests. Lorazepam caused a significant reduction in saccadic peak velocity, the VAS alertness score and impairment of memory and body sway. TPA023 had significant dose dependent effects on saccadic peak velocity (85 deg/sec maximum reduction at the higher dose) that approximated the effects of lorazepam. In contrast to lorazepam, TPA023 had no detectabLe effects on saccadic latency or inaccuracy. Also unlike lorazepam, TPA023 did not affect VAS alertness, memory or body sway. These results show that the effect profile of TPA023 differs markedly from that of lorazepam, at doses that were equipotent with regard to effects on saccadic peak veLocity. Contrary to lorazepam, TPA023 caused no detectable memory impairment or postural imbalance. These differences reflect the selectivity of TPA023 for different GABA(A) receptor subtypes.

Theoretical and experimental NMR studies on muscimol from fly agaric mushroom (Amanita muscaria)

NASA Astrophysics Data System (ADS)

Kupka, Teobald; Wieczorek, Piotr P.

2016-01-01

In this article we report results of combined theoretical and experimental NMR studies on muscimol, the bioactive alkaloid from fly agaric mushroom (Amanita muscaria). The assignment of 1H and 13C NMR spectra of muscimol in DMSO-d6 was supported by additional two-dimensional heteronuclear correlated spectra (2D NMR) and gauge independent atomic orbital (GIAO) NMR calculations using density functional theory (DFT). The effect of solvent in theoretical calculations was included via polarized continuum model (PCM) and the hybrid three-parameter B3LYP density functional in combination with 6-311++G(3df,2pd) basis set enabled calculation of reliable structures of non-ionized (neutral) molecule and its NH and zwitterionic forms in the gas phase, chloroform, DMSO and water. GIAO NMR calculations, using equilibrium and rovibrationally averaged geometry, at B3LYP/6-31G* and B3LYP/aug-cc-pVTZ-J levels of theory provided muscimol nuclear magnetic shieldings. The theoretical proton and carbon chemical shifts were critically compared with experimental NMR spectra measured in DMSO. Our results provide useful information on its structure in solution. We believe that such data could improve the understanding of basic features of muscimol at atomistic level and provide another tool in studies related to GABA analogs.

Functional characterization of GABAA receptor-mediated modulation of cortical neuron network activity in microelectrode array recordings.

PubMed

Bader, Benjamin M; Steder, Anne; Klein, Anders Bue; Frølund, Bente; Schroeder, Olaf H U; Jensen, Anders A

2017-01-01

The numerous γ-aminobutyric acid type A receptor (GABAAR) subtypes are differentially expressed and mediate distinct functions at neuronal level. In this study we have investigated GABAAR-mediated modulation of the spontaneous activity patterns of primary neuronal networks from murine frontal cortex by characterizing the effects induced by a wide selection of pharmacological tools at a plethora of activity parameters in microelectrode array (MEA) recordings. The basic characteristics of the primary cortical neurons used in the recordings were studied in some detail, and the expression levels of various GABAAR subunits were investigated by western blotting and RT-qPCR. In the MEA recordings, the pan-GABAAR agonist muscimol and the GABABR agonist baclofen were observed to mediate phenotypically distinct changes in cortical network activity. Selective augmentation of αβγ GABAAR signaling by diazepam and of δ-containing GABAAR (δ-GABAAR) signaling by DS1 produced pronounced changes in the majority of the activity parameters, both drugs mediating similar patterns of activity changes as muscimol. The apparent importance of δ-GABAAR signaling for network activity was largely corroborated by the effects induced by the functionally selective δ-GABAAR agonists THIP and Thio-THIP, whereas the δ-GABAAR selective potentiator DS2 only mediated modest effects on network activity, even when co-applied with low THIP concentrations. Interestingly, diazepam exhibited dramatically right-shifted concentration-response relationships at many of the activity parameters when co-applied with a trace concentration of DS1 compared to when applied alone. In contrast, the potencies and efficacies displayed by DS1 at the networks were not substantially altered by the concomitant presence of diazepam. In conclusion, the holistic nature of the information extractable from the MEA recordings offers interesting insights into the contributions of various GABAAR subtypes/subgroups to cortical

Too Little and Too Much: Hypoactivation and Disinhibition of Medial Prefrontal Cortex Cause Attentional Deficits

PubMed Central

McGarrity, Stephanie; Mason, Rob; Fone, Kevin C.

2014-01-01

Attentional deficits are core symptoms of schizophrenia, contributing strongly to disability. Prefrontal dysfunction has emerged as a candidate mechanism, with clinical evidence for prefrontal hypoactivation and disinhibition (reduced GABAergic inhibition), possibly reflecting different patient subpopulations. Here, we tested in rats whether imbalanced prefrontal neural activity impairs attention. To induce prefrontal hypoactivation or disinhibition, we microinfused the GABA-A receptor agonist muscimol (C4H6N2O2; 62.5, 125, 250 ng/side) or antagonist picrotoxin (C30H34O13; 75, 150, 300 ng/side), respectively, into the medial prefrontal cortex. Using the five-choice serial reaction time (5CSRT) test, we showed that both muscimol and picrotoxin impaired attention (reduced accuracy, increased omissions). Muscimol also impaired response control (increased premature responses). In addition, muscimol dose dependently reduced open-field locomotor activity, whereas 300 ng of picrotoxin caused locomotor hyperactivity; sensorimotor gating (startle prepulse inhibition) was unaffected. Therefore, infusion effects on the 5CSRT test can be dissociated from sensorimotor effects. Combining microinfusions with in vivo electrophysiology, we showed that muscimol inhibited prefrontal firing, whereas picrotoxin increased firing, mainly within bursts. Muscimol reduced and picrotoxin enhanced bursting and both drugs changed the temporal pattern of bursting. Picrotoxin also markedly enhanced prefrontal LFP power. Therefore, prefrontal hypoactivation and disinhibition both cause attentional deficits. Considering the electrophysiological findings, this suggests that attention requires appropriately tuned prefrontal activity. Apart from attentional deficits, prefrontal disinhibition caused additional neurobehavioral changes that may be relevant to schizophrenia pathophysiology, including enhanced prefrontal bursting and locomotor hyperactivity, which have been linked to psychosis

Insights into GABA receptor signalling in TM3 Leydig cells.

PubMed

Doepner, Richard F G; Geigerseder, Christof; Frungieri, Monica B; Gonzalez-Calvar, Silvia I; Calandra, Ricardo S; Raemsch, Romi; Fohr, Karl; Kunz, Lars; Mayerhofer, Artur

2005-01-01

Gamma-aminobutyric acid (GABA) is an emerging signalling molecule in endocrine organs, since it is produced by endocrine cells and acts via GABA(A) receptors in a paracrine/autocrine fashion. Testicular Leydig cells are producers and targets for GABA. These cells express GABA(A) receptor subunits and in the murine Leydig cell line TM3 pharmacological activation leads to increased proliferation. The signalling pathway of GABA in these cells is not known in this study. We therefore attempted to elucidate details of GABA(A) signalling in TM3 and adult mouse Leydig cells using several experimental approaches. TM3 cells not only express GABA(A )receptor subunits, but also bind the GABA agonist [(3)H]muscimol with a binding affinity in the range reported for other endocrine cells (K(d) = 2.740 +/- 0.721 nM). However, they exhibit a low B(max) value of 28.08 fmol/mg protein. Typical GABA(A) receptor-associated events, including Cl(-) currents, changes in resting membrane potential, intracellular Ca(2+) or cAMP, were not measurable with the methods employed in TM3 cells, or, as studied in part, in primary mouse Leydig cells. GABA or GABA(A) agonist isoguvacine treatment resulted in increased or decreased levels of several mRNAs, including transcription factors (c-fos, hsf-1, egr-1) and cell cycle-associated genes (Cdk2, cyclin D1). In an attempt to verify the cDNA array results and because egr-1 was recently implied in Leydig cell development, we further studied this factor. RT-PCR and Western blotting confirmed a time-dependent regulation of egr-1 in TM3. In the postnatal testis egr-1 was seen in cytoplasmic and nuclear locations of developing Leydig cells, which bear GABA(A) receptors and correspond well to TM3 cells. Thus, GABA acts via an atypical novel signalling pathway in TM3 cells. Further details of this pathway remain to be elucidated. Copyright (c) 2005 S. Karger AG, Basel.

Characteristics of concatemeric GABAA receptors containing α4/δ subunits expressed in Xenopus oocytes

PubMed Central

Shu, Hong-Jin; Bracamontes, John; Taylor, Amanda; Wu, Kyle; Eaton, Megan M; Akk, Gustav; Manion, Brad; Evers, Alex S; Krishnan, Kathiresan; Covey, Douglas F; Zorumski, Charles F; Steinbach, Joe Henry; Mennerick, Steven

2012-01-01

BACKGROUND AND PURPOSE GABAA receptors mediate both synaptic and extrasynaptic actions of GABA. In several neuronal populations, α4 and δ subunits are key components of extrasynaptic GABAA receptors that strongly influence neuronal excitability and could mediate the effects of neuroactive agents including neurosteroids and ethanol. However, these receptors can be difficult to study in native cells and recombinant δ subunits can be difficult to express in heterologous systems. EXPERIMENTAL APPROACH We engineered concatemeric (fused) subunits to ensure δ and α4 subunit expression. We tested the pharmacology of the concatemeric receptors, compared with a common synaptic-like receptor subunit combination (α1 +β2 +γ2L), and with free-subunit α4/δ receptors, expressed in Xenopus oocytes. KEY RESULTS δ-β2 −α4 +β2-α4 cRNA co-injected into Xenopus oocytes resulted in GABA-gated currents with the expected pharmacological properties of α4/δ-containing receptors. Criteria included sensitivity to agonists of different efficacy, sensitivity to the allosteric activator pentobarbital, and modulation of agonist responses by DS2 (4-chloro-N-[2-(2-thienyl)imidazo[1,2-a]pyridine-3-yl benzamide; a δ-selective positive modulator), furosemide, and Zn2+. We used the concatemers to examine neurosteroid sensitivity of extrasynaptic-like, δ-containing receptors. We found no qualitative differences between extrasynaptic-like receptors and synaptic-like receptors in the actions of either negative or positive neurosteroid modulators of receptor function. Quantitative differences were explained by the partial agonist effects of the natural agonist GABA and by a mildly increased sensitivity to low steroid concentrations. CONCLUSIONS AND IMPLICATIONS The neurosteroid structure-activity profile for α4/δ-containing extrasynaptic receptors is unlikely to differ from that of synaptic-like receptors such as α1/β2/γ2-containing receptors. PMID:21950777

Alpha1- and alpha2-containing GABAA receptor modulation is not necessary for benzodiazepine-induced hyperphagia.

PubMed

Morris, H V; Nilsson, S; Dixon, C I; Stephens, D N; Clifton, P G

2009-06-01

Benzodiazepines increase food intake, an effect attributed to their ability to enhance palatability. We investigated which GABA(A) receptor subtypes may be involved in mediating benzodiazepine-induced hyperphagia. The role of the alpha2 subtype was investigated by observing the effects of midazolam, on the behavioural satiety sequence in mice with targeted deletion of the alpha2 gene (alpha2 knockout). Midazolam (0.125, 0.25 and 0.5mg/kg) increased food intake and the amount of time spent feeding in alpha2 knockout mice, suggesting that BZ-induced hyperphagia does not involve alpha2-containing GABA(A) receptors. We further investigated the roles of alpha1- and alpha3-containing GABA(A) receptors in mediating BZ-induced hyperphagia. We treated alpha2(H101R) mice, in which alpha2-containing receptors are rendered benzodiazepine insensitive, with L-838417, a compound which acts as a partial agonist at alpha2-, alpha3- and alpha5-receptors but is inactive at alpha1-containing receptors. L-838417 (10 and 30 mg/kg) increased food intake and the time spent feeding in both wildtype and alpha2(H101R) mice, demonstrating that benzodiazepine-induced hyperphagia does not require alpha1- and alpha2-containing GABA(A) receptors. These observations, together with evidence against the involvement of alpha5-containing GABA(A) receptors, suggest that alpha3-containing receptors mediate BZ-induced hyperphagia in the mouse.

Ipsilateral feeding-specific circuits between the nucleus accumbens shell and the lateral hypothalamus: regulation by glutamate and GABA receptor subtypes.

PubMed

Urstadt, Kevin R; Kally, Peter; Zaidi, Sana F; Stanley, B Glenn

2013-04-01

The nucleus accumbens shell (AcbSh) and the lateral hypothalamus (LH) are both involved in the control of food intake. Activation of GABA(A) receptors or blockade of AMPA and kainate receptors within the AcbSh induces feeding, as does blockade of GABA(A) receptors or activation of NMDA receptors in the LH. Further, evidence suggests that feeding induced via the AcbSh can be suppressed by LH inhibition. However, it is unclear if this suppression is specific to feeding. Adult male Sprague-Dawley rats with 3 intracranial guide cannulas, one unilaterally into the AcbSh and two bilaterally into the LH, were used to explore this issue. DNQX (1.25 μg) or muscimol (100 ng) infused into the AcbSh unilaterally elicited feeding, and this elicited intake was suppressed by bilateral LH injection of d-AP5 (2 μg) or muscimol (25 ng). The effectiveness of d-AP5 or muscimol infusion into either the LH site ipsilateral or contralateral to the AcbSh injection was compared. Ipsilateral LH injection of d-AP5 or muscimol was significantly more effective than contralateral injection in suppressing food intake initiated by AcbSh injection of DNQX or muscimol. These results add to the prior evidence that inhibition of the LH through pharmacological modulation of NMDA or GABA(A) receptors specifically suppresses feeding initiated by AcbSh inhibition, and that these two regions communicate via an ipsilateral circuit to specifically regulate feeding. Copyright © 2012 Elsevier Ltd. All rights reserved.

Taurine-induced attenuation of MPP+ neurotoxicity in vitro: a possible role for the GABA(A) subclass of GABA receptors.

PubMed

O'Byrne, M B; Tipton, K F

2000-05-01

Taurine is a sulphur-containing beta-amino acid found in high (millimolar) concentrations in excitable tissues such as brain and heart. Its suggested roles include osmoregulator, thermoregulator, neuromodulator, and potential neurotransmitter. This amino acid has also been shown to be released in large concentrations during ischaemia and excitotoxin-induced neuronal damage. Here we report a protective effect of taurine against MPP(+)-induced neurotoxicity in coronal slices from rat brain. Significant protective effects were observed at taurine concentrations of 20 and 1 mM, suggesting a potential role for taurine in cases of neuronal insult. Studies with the synthetic taurine analogues taurine phosphonate, guanidinoethane sulphonate, and trimethyltaurine suggested the observed effect to be mediated via an extracellular mechanism. The use of GABA receptor ligands muscimol and bicuculline indicated the effect to be mediated through activation of GABA(A) receptors.

Modulation of neuronal and recombinant GABAA receptors by redox reagents

PubMed Central

Amato, Alessandra; Connolly, Christopher N; Moss, Stephen J; Smart, Trevor G

1999-01-01

The functional role played by the postulated disulphide bridge in γ-aminobutyric acid type A (GABAA) receptors and its susceptibility to oxidation and reduction were studied using recombinant (murine receptor subunits expressed in human embryonic kidney cells) and rat neuronal GABAA receptors in conjunction with whole-cell and single channel patch-clamp techniques. The reducing agent dithiothreitol (DTT) reversibly potentiated GABA-activated responses (IGABA) of α1β1 or α1β2 receptors while the oxidizing reagent 5,5′-dithio-bis-(2-nitrobenzoic acid) (DTNB) caused inhibition. Redox modulation of IGABA was independent of GABA concentration, membrane potential and the receptor agonist and did not affect the GABA EC50 or Hill coefficient. The endogenous antioxidant reduced glutathione (GSH) also potentiated IGABA in α1β2 receptors, while both the oxidized form of DTT and glutathione (GSSG) caused small inhibitory effects. Recombinant receptors composed of α1β1γ2S or α1β2γ2S were considerably less sensitive to DTT and DTNB. For neuronal GABAA receptors, IGABA was enhanced by flurazepam and relatively unaffected by redox reagents. However, in cultured sympathetic neurones, nicotinic acetylcholine-activated responses were inhibited by DTT whilst in cerebellar granule neurones, NMDA-activated currents were potentiated by DTT and inhibited by DTNB. Single GABA-activated ion channel currents exhibited a conductance of 16 pS for α1β1 constructs. DTT did not affect the conductance or individual open time constants determined from dwell time histograms, but increased the mean open time by affecting the channel open probability without increasing the number of cell surface receptors. A kinetic model of the effects of DTT and DTNB suggested that the receptor existed in equilibrium between oxidized and reduced forms. DTT increased the rate of entry into reduced receptor forms and also into desensitized states. DTNB reversed these kinetic effects. Our results

Modulation of acetylcholine release from rat striatal slices by the GABA/benzodiazepine receptor complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

Supavilai, P.; Karobath, M.

1985-02-04

GABA, THIP and muscimol enhance spontaneous and inhibit electrically induced release of tritium labelled compounds from rat striatal slices which have been pre-labelled with /sup 3/H-choline. Baclofen is inactive in this model. Muscimol can inhibit electrically induced release of tritiated material by approximately 75% with half maximal effects at 2 ..mu..M. The response to muscimol can be blocked by the GABA antagonists bicuculline methobromide, picrotoxin, anisatin, R 5135 and CPTBO (cyclopentylbicyclophosphate). Drugs which act on the benzodiazepine receptor (BR) require the presence of muscimol to be effective and they modulate the effects of muscimol in a bidirectional manner. Thus BRmore » agonists enhance and inverse BR agonists attenuate the inhibitory effects of muscimol on electrically induced release. Ro15-1788, a BR antagonist, does not modulate the inhibitory effects of muscimol but antagonizes the actions of clonazepam, a BR agonist, and of DMCM, an inverse BR agonist. These results demonstrate that a GABA/benzodiazepine receptor complex can modulate acetylcholine release from rat striatal slices in vitro. 24 references, 3 figures, 5 table.« less

Non-neuronal, slow GABA signalling in the ventrobasal thalamus targets δ-subunit-containing GABAA receptors

PubMed Central

Jiménez-González, Cristina; Pirttimaki, Tiina; Cope, David W; Parri, H R

2011-01-01

The rodent ventrobasal (VB) thalamus contains a relatively uniform population of thalamocortical (TC) neurons that receive glutamatergic input from the vibrissae and the somatosensory cortex, and inhibitory input from the nucleus reticularis thalami (nRT). In this study we describe γ-aminobutyric acid (GABA)A receptor-dependent slow outward currents (SOCs) in TC neurons that are distinct from fast inhibitory postsynaptic currents (IPSCs) and tonic currents. SOCs occurred spontaneously or could be evoked by hypo-osmotic stimulus, and were not blocked by tetrodotoxin, removal of extracellular Ca2+ or bafilomycin A1, indicating a non-synaptic, non-vesicular GABA origin. SOCs were more common in TC neurons of the VB compared with the dorsal lateral geniculate nucleus, and were rarely observed in nRT neurons, whilst SOC frequency in the VB increased with age. Application of THIP, a selective agonist at δ-subunit-containing GABAA receptors, occluded SOCs, whereas the benzodiazepine site inverse agonist β-CCB had no effect, but did inhibit spontaneous and evoked IPSCs. In addition, the occurrence of SOCs was reduced in mice lacking the δ-subunit, and their kinetics were also altered. The anti-epileptic drug vigabatrin increased SOC frequency in a time-dependent manner, but this effect was not due to reversal of GABA transporters. Together, these data indicate that SOCs in TC neurons arise from astrocytic GABA release, and are mediated by δ-subunit-containing GABAA receptors. Furthermore, these findings suggest that the therapeutic action of vigabatrin may occur through the augmentation of this astrocyte–neuron interaction, and highlight the importance of glial cells in CNS (patho) physiology. PMID:21395866

GABAergic excitation of spider mechanoreceptors increases information capacity by increasing entropy rather than decreasing jitter.

PubMed

Pfeiffer, Keram; French, Andrew S

2009-09-02

Neurotransmitter chemicals excite or inhibit a range of sensory afferents and sensory pathways. These changes in firing rate or static sensitivity can also be associated with changes in dynamic sensitivity or membrane noise and thus action potential timing. We measured action potential firing produced by random mechanical stimulation of spider mechanoreceptor neurons during long-duration excitation by the GABAA agonist muscimol. Information capacity was estimated from signal-to-noise ratio by averaging responses to repeated identical stimulation sequences. Information capacity was also estimated from the coherence function between input and output signals. Entropy rate was estimated by a data compression algorithm and maximum entropy rate from the firing rate. Action potential timing variability, or jitter, was measured as normalized interspike interval distance. Muscimol increased firing rate, information capacity, and entropy rate, but jitter was unchanged. We compared these data with the effects of increasing firing rate by current injection. Our results indicate that the major increase in information capacity by neurotransmitter action arose from the increased entropy rate produced by increased firing rate, not from reduction in membrane noise and action potential jitter.

Neurosteroid Modulators of GABAA Receptors Differentially Modulate Ethanol Intake Patterns in Male C57BL/6J Mice

PubMed Central

Ford, Matthew M.; Nickel, Jeffrey D.; Phillips, Tamara J.; Finn, Deborah A.

2006-01-01

Background Allopregnanolone (ALLO) and structurally related endogenous neurosteroids are potent modulators of GABAA receptor function at physiologically relevant concentrations. Accumulating evidence implicates a modulatory role for ALLO in behavioral processes underlying ethanol self-administration, discrimination and reinstatement. The purpose of this study was to evaluate the impact of exogenous neurosteroid challenges with the agonist ALLO and the partial agonist/antagonist epipregnanolone (EPI) on the microarchitecture of ethanol drinking patterns. Methods Male C57BL/6J mice were initiated to consume an unsweetened 10% v/v ethanol solution (10E) by a saccharin fading procedure during daily 2-hour limited access sessions beginning 1 hour after dark phase onset. Cumulative lick responses were recorded for 10E and water using lickometer circuits. After establishing 10E intake baselines, mice were habituated to vehicle injection (VEH; 20% w/v β-cyclodextrin; i.p.), and then were treated with either VEH or neurosteroid immediately prior to the drinking session. Each mouse received a series of ALLO doses (3.2, 10, 17 and 24 mg/kg) alone and EPI doses (0.15, 1, 3 and 10 mg/kg) alone in a counterbalanced within-group design. Results The GABAA receptor positive modulator, ALLO, dose-dependently modulated overall ethanol intake throughout the 2-hr session with the 3.2 mg/kg dose eliciting a significant increase whereas the 24 mg/kg dose produced a significant suppression of ethanol intake versus vehicle pretreatment. ALLO-evoked alterations in intake corresponded with a significant, dose-dependent alterations in bout frequency and inter-bout interval. ALLO also elicited robust, dose-dependent elevations in 10E licks during the initial 5-minutes of access, but subsequently resulted in a dose-dependent suppression of 10E licks during session minutes 20–80. In contrast, the partial agonist/antagonist neurosteroid, EPI, exhibited no influence on any consumption parameter

Characterization of GABAA receptor ligands with automated patch-clamp using human neurons derived from pluripotent stem cells

PubMed Central

Yuan, Nina Y.; Poe, Michael M.; Witzigmann, Christopher; Cook, James M.; Stafford, Douglas; Arnold, Leggy A.

2016-01-01

Introduction Automated patch clamp is a recent but widely used technology to assess pre-clinical drug safety. With the availability of human neurons derived from pluripotent stem cells, this technology can be extended to determine CNS effects of drug candidates, especially those acting on the GABAA receptor. Methods iCell Neurons (Cellular Dynamics International, A Fujifilm Company) were cultured for ten days and analyzed by patch clamp in the presence of agonist GABA or in combination with positive allosteric GABAA receptor modulators. Both efficacy and affinity were determined. In addition, mRNA of GABAA receptor subunits were quantified by qRT-PCR. Results We have shown that iCell Neurons are compatible with the IonFlux microfluidic system of the automated patch clamp instrument. Resistance ranging from 15-25 MΩ was achieved for each trap channel of patch clamped cells in a 96-well plate format. GABA induced a robust change of current with an EC50 of 0.43 μM. Positive GABAA receptor modulators diazepam, HZ166, and CW-04-020 exhibited EC50 values of 0.42 μM, 1.56 μM, and 0.23 μM, respectively. The α2/α3/α5 selective compound HZ166-induced the highest potentiation (efficacy) of 810% of the current induced by 100 nM GABA. Quantification of GABAA receptor mRNA in iCell Neurons revealed high levels of α5 and β3 subunits and low levels of α1, which is similar to the configuration in human neonatal brain. Discussion iCell Neurons represent a new cellular model to characterize GABAergic compounds using automated patch clamp. These cells have excellent representation of cellular GABAA receptor distribution that enable determination of total small molecule efficacy and affinity as measured by cell membrane current change. PMID:27544543

Women with PTSD have a changed sensitivity to GABA-A receptor active substances.

PubMed

Möller, Anna Tiihonen; Bäckström, Torbjörn; Nyberg, Sigrid; Söndergaard, Hans Peter; Helström, Lotti

2016-06-01

The use of benzodiazepines in treating anxiety symptoms in patients with posttraumatic stress disorder (PTSD) has been debated. Studies on other anxiety disorders have indicated changed sensitivity to GABA-A receptor active substances. In the present study, we investigated the GABA receptor sensitivity in PTSD patients. Injections of allopreganolone, diazepam, and flumazenil were carried out, each on separate occasions, in 10 drug naïve patients with PTSD compared to 10 healthy controls. Effects were measured in saccadic eye velocity (SEV) and in subjective ratings of sedation. The PTSD patients were less sensitive to allopregnanolone compared with healthy controls. This was seen as a significant difference in SEV between the groups (p = 0.047). Further, the patients were less sensitive to diazepam, with a significant less increase in sedation compared to controls (p = 0.027). After flumazenil injection, both patients and controls had a significant agonistic effect on SEV, leading to decreased SEV after injection. The patients also responded with an increase in sedation after flumazenil injection, while this was not seen in the controls. Patients with PTSD have a changed sensitivity to GABA-A receptor active substances. As a consequence of this, benzodiazepines and other GABA-A receptor active compounds such as sleeping pills will be less useful for this group of patients.

Chronic administration of aripiprazole activates GSK3β-dependent signalling pathways, and up-regulates GABAA receptor expression and CREB1 activity in rats.

PubMed

Pan, Bo; Huang, Xu-Feng; Deng, Chao

2016-07-20

Aripiprazole is a D2-like receptor (D2R) partial agonist with a favourable clinical profile. Previous investigations indicated that acute and short-term administration of aripiprazole had effects on PKA activity, GSK3β-dependent pathways, GABAA receptors, NMDA receptor and CREB1 in the brain. Since antipsychotics are used chronically in clinics, the present study investigated the long-term effects of chronic oral aripiprazole treatment on these cellular signalling pathways, in comparison with haloperidol (a D2R antagonist) and bifeprunox (a potent D2R partial agonist). We found that the Akt-GSK3β pathway was activated by aripiprazole and bifeprunox in the prefrontal cortex; NMDA NR2A levels were reduced by aripiprazole and haloperidol. In the nucleus accumbens, all three drugs increased Akt-GSK3β signalling; in addition, both aripiprazole and haloperidol, but not bifeprunox, increased the expression of Dvl-3, β-catenin and GABAA receptors, NMDA receptor subunits, as well as CREB1 phosphorylation levels. The results suggest that chronic oral administration of aripiprazole affects schizophrenia-related cellular signalling pathways and markers (including Akt-GSK3β signalling, Dvl-GSK3β-β-catenin signalling, GABAA receptor, NMDA receptor and CREB1) in a brain-region-dependent manner; the selective effects of aripiprazole on these signalling pathways might be associated with its unique clinical effects.

Chronic administration of aripiprazole activates GSK3β-dependent signalling pathways, and up-regulates GABAA receptor expression and CREB1 activity in rats

PubMed Central

Pan, Bo; Huang, Xu-Feng; Deng, Chao

2016-01-01

Aripiprazole is a D2-like receptor (D2R) partial agonist with a favourable clinical profile. Previous investigations indicated that acute and short-term administration of aripiprazole had effects on PKA activity, GSK3β-dependent pathways, GABAA receptors, NMDA receptor and CREB1 in the brain. Since antipsychotics are used chronically in clinics, the present study investigated the long-term effects of chronic oral aripiprazole treatment on these cellular signalling pathways, in comparison with haloperidol (a D2R antagonist) and bifeprunox (a potent D2R partial agonist). We found that the Akt-GSK3β pathway was activated by aripiprazole and bifeprunox in the prefrontal cortex; NMDA NR2A levels were reduced by aripiprazole and haloperidol. In the nucleus accumbens, all three drugs increased Akt-GSK3β signalling; in addition, both aripiprazole and haloperidol, but not bifeprunox, increased the expression of Dvl-3, β-catenin and GABAA receptors, NMDA receptor subunits, as well as CREB1 phosphorylation levels. The results suggest that chronic oral administration of aripiprazole affects schizophrenia-related cellular signalling pathways and markers (including Akt-GSK3β signalling, Dvl-GSK3β-β-catenin signalling, GABAA receptor, NMDA receptor and CREB1) in a brain-region-dependent manner; the selective effects of aripiprazole on these signalling pathways might be associated with its unique clinical effects. PMID:27435909

Complexity measures of the central respiratory networks during wakefulness and sleep

NASA Astrophysics Data System (ADS)

Dragomir, Andrei; Akay, Yasemin; Curran, Aidan K.; Akay, Metin

2008-06-01

Since sleep is known to influence respiratory activity we studied whether the sleep state would affect the complexity value of the respiratory network output. Specifically, we tested the hypothesis that the complexity values of the diaphragm EMG (EMGdia) activity would be lower during REM compared to NREM. Furthermore, since REM is primarily generated by a homogeneous population of neurons in the medulla, the possibility that REM-related respiratory output would be less complex than that of the awake state was also considered. Additionally, in order to examine the influence of neuron vulnerabilities within the rostral ventral medulla (RVM) on the complexity of the respiratory network output, we inhibited respiratory neurons in the RVM by microdialysis of GABAA receptor agonist muscimol. Diaphragm EMG, nuchal EMG, EEG, EOG as well as other physiological signals (tracheal pressure, blood pressure and respiratory volume) were recorded from five unanesthetized chronically instrumented intact piglets (3-10 days old). Complexity of the diaphragm EMG (EMGdia) signal during wakefulness, NREM and REM was evaluated using the approximate entropy method (ApEn). ApEn values of the EMGdia during NREM and REM sleep were found significantly (p < 0.05 and p < 0.001, respectively) lower than those of awake EMGdia after muscimol inhibition. In the absence of muscimol, only the differences between REM and wakefulness ApEn values were found to be significantly different.

Ventral tegmental area GABA neurons and opiate motivation

PubMed Central

Vargas-Perez, Hector; Mabey, Jennifer K.; Shin, Samuel I.; Steffensen, Scott C.; van der Kooy, Derek

2013-01-01

Rational Past research has demonstrated that when an animal changes from a previously drug-naive to an opiate-dependent and withdrawn state, morphine’s motivational effects are switched from a tegmental pedunculopontine nucleus (TPP)-dependent to a dopamine-dependent pathway. Interestingly, a corresponding change is observed in ventral tegmental area (VTA) GABAA receptors, which change from mediating hyperpolarization of VTA GABA neurons to mediating depolarization. Objectives The present study investigated whether pharmacological manipulation of VTA GABAA receptor activity could directly influence the mechanisms underlying opiate motivation. Results Using an unbiased place conditioning procedure, we demonstrated that in Wistar rats, intra-VTA administration of furosemide, a Cl− cotransporter inhibitor, was able to promote a switch in the mechanisms underlying morphine’s motivational properties, one which is normally observed only after chronic opiate exposure. This behavioral switch was prevented by intra-VTA administration of acetazolamide, an inhibitor of the bicarbonate ion-producing carbonic anhydrase enzyme. Electrophysiological recordings of mouse VTA showed that furosemide reduced the sensitivity of VTA GABA neurons to inhibition by the GABAA receptor agonist muscimol, instead increasing the firing rate of a significant subset of these GABA neurons. Conclusion Our results suggest that the carbonic anhydrase enzyme may constitute part of a common VTA GABA neuron-based biological pathway responsible for controlling the mechanisms underlying opiate motivation, supporting the hypothesis that VTA GABAA receptor hyperpolarization or depolarization is responsible for selecting TPP- or dopamine-dependent motivational outputs, respectively. PMID:23392354

Enhancement of GABAergic transmission by zolpidem, an imidazopyridine with preferential affinity for type I benzodiazepine receptors.

PubMed

Biggio, G; Concas, A; Corda, M G; Serra, M

1989-02-28

The effect of zolpidem, an imidazopyridine derivative with high affinity at the type I benzodiazepine recognition site, on the function of the GABAA/ionophore receptor complex was studied in vitro. Zolpidem, mimicking the action of diazepam, increased [3H]GABA binding, enhanced muscimol-stimulated 36Cl- uptake and reduced [35S]TBPS binding in rat cortical membrane preparations. Zolpidem was less effective than diazepam on the above parameters. Zolpidem induced a lower increase of [3H]GABA binding (23 vs. 35%) and muscimol-stimulated 36Cl- uptake (22 vs. 40%) and a smaller decrease of [35S]TBPS binding (47 vs. 77%) than diazepam. The finding that zolpidem enhanced the function of GABAergic synapses with an efficacy qualitatively and quantitatively different from that of diazepam suggests that this compound is a partial agonist at the benzodiazepine recognition site. Thus, our results are consistent with the view that the biochemical and pharmacological profile of a benzodiazepine recognition site ligand reflects its efficacy to enhance GABAergic transmission. Whether the preferential affinity of zolpidem at the type I site is involved in its atypical biochemical and pharmacological profile remains to be clarified.

Contribution of the lateral lemniscus to the control of swallowing in decerebrate cats.

PubMed

Ota, R; Takakusaki, K; Katada, A; Harada, H; Nonaka, S; Harabuchi, Y

2013-12-19

Lateral lemniscus, a relay nucleus of auditory sensation, is involved in the control of phonatory movements such as human speech and vocalization of animals. The present study was designed to test whether neurons in the lateral lemniscus contributed to the control of swallowing, one of non-phonic oro-pharyngolaryngeal movements. In acutely decerebrated cats (n=15), swallowing was induced by electrical stimulation (20-80μA at 10Hz for 20s with rectangular pulses of 0.2ms duration) delivered to the superior laryngeal nerve (SLN). Repetitive electrical stimulation (30-50μA at 50Hz for 10-20s) applied to the dorsal nucleus of the lateral lemniscus (LLD) increased the number and reduced the latency to the onset of the SLN-induced swallowing. On the other hand, stimulation of the ventral nucleus of the lateral lemniscus and the paralemniscal area, corresponding to the ventrolateral part of the parabrachial nucleus and the Kölliker-Fuse nucleus, often suppressed the SLN-induced swallowing. Microinjection of NMDA (0.1-0.15μl, 5.0-10mM) into the LLD through a stereotaxically placed glass micropipette facilitated the SLN-induced swallowing, i.e., the number was increased and the latency of swallowing was reduced. We also injected muscimol (a gamma amino-butyric acid (GABA)A receptor agonist), bicuculline (a GABAA receptor antagonist) and baclofen (a GABAB receptor agonist) into the LLD (0.1-0.15μl and 5.0mM for each substance). It was observed that an injection of muscimol suppressed the SLN-induced swallowing. However, an injection of bicuculline facilitated the swallowing. An injection of baclofen did not alter the swallowing. These results suggest the presence of functional topography in the lateral lemniscus and the paralemniscal area in relation to the control of swallowing. The facilitatory LLD-effects on swallowing are modulated by glutamatergic and GABAergic receptors on neurons in the LLD. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

Nootropic α7 nicotinic receptor allosteric modulator derived from GABAA receptor modulators

PubMed Central

Ng, Herman J.; Whittemore, Edward R.; Tran, Minhtam B.; Hogenkamp, Derk J.; Broide, Ron S.; Johnstone, Timothy B.; Zheng, Lijun; Stevens, Karen E.; Gee, Kelvin W.

2007-01-01

Activation of brain α7 nicotinic acetylcholine receptors (α7 nAChRs) has broad therapeutic potential in CNS diseases related to cognitive dysfunction, including Alzheimer's disease and schizophrenia. In contrast to direct agonist activation, positive allosteric modulation of α7 nAChRs would deliver the clinically validated benefits of allosterism to these indications. We have generated a selective α7 nAChR-positive allosteric modulator (PAM) from a library of GABAA receptor PAMs. Compound 6 (N-(4-chlorophenyl)-α-[[(4-chloro-phenyl)amino]methylene]-3-methyl-5-isoxazoleacet-amide) evokes robust positive modulation of agonist-induced currents at α7 nAChRs, while preserving the rapid native characteristics of desensitization, and has little to no efficacy at other ligand-gated ion channels. In rodent models, it corrects sensory-gating deficits and improves working memory, effects consistent with cognitive enhancement. Compound 6 represents a chemotype for allosteric activation of α7 nAChRs, with therapeutic potential in CNS diseases with cognitive dysfunction. PMID:17470817

Functional expression of the GABAA receptor α2 and α3 subunits at synapses between intercalated medial paracapsular neurons of mouse amygdala

PubMed Central

Geracitano, Raffaella; Fischer, David; Kasugai, Yu; Ferraguti, Francesco; Capogna, Marco

2012-01-01

In the amygdala, GABAergic neurons in the intercalated medial paracapsular cluster (Imp) have been suggested to play a key role in fear learning and extinction. These neurons project to the central (CE) amygdaloid nucleus and to other areas within and outside the amygdala. In addition, they give rise to local collaterals that innervate other neurons in the Imp. Several drugs, including benzodiazepines (BZ), are allosteric modulators of GABAA receptors. BZ has both anxiolytic and sedative actions, which are mediated through GABAA receptors containing α2/α3 and α1 subunits, respectively. To establish whether α1 or α2/α3 subunits are expressed at Imp cell synapses, we used paired recordings of anatomically identified Imp neurons and high resolution immunocytochemistry in the mouse. We observed that a selective α3 subunit agonist, TP003 (100 nM), significantly increased the decay time constant of the unitary IPSCs. A similar effect was also induced by zolpidem (10 μM) or by diazepam (1 μM). In contrast, lower doses of zolpidem (0.1–1 μM) did not significantly alter the kinetics of the unitary IPSCs. Accordingly, immunocytochemical experiments established that the α2 and α3, but not the α1 subunits of the GABAA receptors, were present at Imp cell synapses of the mouse amygdala. These results define, for the first time, some of the functional GABAA receptor subunits expressed at synapses of Imp cells. The data also provide an additional rationale to prompt the search of GABAA receptor α3 selective ligands as improved anxiolytic drugs. PMID:22666188

Actions of Agonists, Fipronil and Ivermectin on the Predominant In Vivo Splice and Edit Variant (RDLbd, I/V) of the Drosophila GABA Receptor Expressed in Xenopus laevis Oocytes

PubMed Central

Suwanmanee, Siros; Buckingham, Steven David; Biggin, Philip; Sattelle, David

2014-01-01

Ionotropic GABA receptors are the targets for several classes of insecticides. One of the most widely-studied insect GABA receptors is RDL (resistance to dieldrin), originally isolated from Drosophila melanogaster. RDL undergoes alternative splicing and RNA editing, which influence the potency of GABA. Most work has focussed on minority isoforms. Here, we report the first characterisation of the predominant native splice variant and RNA edit, combining functional characterisation with molecular modelling of the agonist-binding region. The relative order of agonist potency is GABA> muscimol> TACA> β-alanine. The I/V edit does not alter the potency of GABA compared to RDLbd. Docking calculations suggest that these agonists bind and activate RDLbdI/V through a similar binding mode. TACA and β-alanine are predicted to bind with lower affinity than GABA, potentially explaining their lower potency, whereas the lower potency of muscimol and isoguvacine cannot be explained structurally from the docking calculations. The A301S (resistance to dieldrin) mutation reduced the potency of antagonists picrotoxin, fipronil and pyrafluprole but the I/V edit had no measurable effect. Ivermectin suppressed responses to GABA of RDLbdI/V, RDLbd and RDLbdI/VA301S. The dieldrin resistant variant also showed reduced sensitivity to Ivermectin. This study of a highly abundant insect GABA receptor isoform will help the design of new insecticides. PMID:24823815

Short-term memory impairment after isoflurane in mice is prevented by the α5 γ-aminobutyric acid type A receptor inverse agonist L-655,708.

PubMed

Saab, Bechara J; Maclean, Ashley J B; Kanisek, Marijana; Zurek, Agnieszka A; Martin, Loren J; Roder, John C; Orser, Beverley A

2010-11-01

Memory blockade is an essential component of the anesthetic state. However, postanesthesia memory deficits represent an undesirable and poorly understood adverse effect. Inhibitory α5 subunit-containing γ-aminobutyric acid subtype A receptors (α5GABAA) are known to play a critical role in memory processes and are highly sensitive to positive modulation by anesthetics. We postulated that inhibiting the activity of α5GABAA receptors during isoflurane anesthesia would prevent memory deficits in the early postanesthesia period. Mice were pretreated with L-655,708, an α5GABAA receptor-selective inverse agonist, or vehicle. They were then exposed to isoflurane for 1 h (1.3%, or 1 minimum alveolar concentration, or air-oxygen control). Then, either 1 or 24 h later, mice were conditioned in fear-associated contextual and cued learning paradigms. In addition, the effect of L-655,708 on the immobilizing dose of isoflurane was studied. Motor coordination, sedation, anxiety, and the concentration of isoflurane in the brain at 5 min, 1 h, and 24 h after isoflurane were also examined. Motor and sensory function recovered within minutes after termination of isoflurane administration. In contrast, a robust deficit in contextual fear memory persisted for at least 24 h. The α5GABAA receptor inverse agonist, L-655,708, completely prevented memory deficits without changing the immobilizing dose of isoflurane. Trace concentrations of isoflurane were measured in the brain 24 h after treatment. Memory deficits occurred long after the sedative, analgesic, and anxiolytic effects of isoflurane subsided. L-655,708 prevented memory deficit, suggesting that an isoflurane interaction at α5GABAA receptors contributes to memory impairment during the early postanesthesia period.

Evidence of two populations of GABA(A) receptors in cerebellar granule cells in culture: different desensitization kinetics, pharmacology, serine/threonine kinase sensitivity, and localization.

PubMed

Robello, M; Amico, C; Cupello, A

1999-12-20

GABA(A) receptors of rat cerebellar granule cells in culture have been studied by the whole cell patch clamp technique. The biphasic desensitization kinetic observed could be due either to different desensitization mechanisms of a single receptor population or to different receptor populations. The overall data indicate that the latter hypothesis is most probably the correct one. In fact, the fast desensitizing component was selectively potentiated by a benzodiazepine agonist and preferentially down-regulated by activation of the protein serine/threonine kinases A and G, as a consequence of the latter characteristic that receptor population was preferentially down-regulated by previous activation of N-methyl-d-aspartate glutamate receptors, via production of nitric oxide and PKG activation, most probably in dendrites. The other population is benzodiazepine insensitive and not influenced by activation of PKA or PKG. This slowly desensitizing population may correspond to the extrasynaptic delta subunit containing GABA(A) receptors described by other authors. Instead, the rapidly desensitizing population appears to represent dendritic synaptic GABA(A) receptors. Copyright 1999 Academic Press.

LINKING GABAA RECEPTOR SUBUNITS TO ALCOHOL-INDUCED CONDITIONED TASTE AVERSION AND RECOVERY FROM ACUTE ALCOHOL INTOXICATION

PubMed Central

Blednov, Y.A.; Benavidez, J.M.; Black, M.; Chandra, D.; Homanics, G.E.; Rudolph, U.; Harris, R.A.

2012-01-01

GABA type A receptors (GABAA-R) are important for ethanol actions and it is of interest to link individual subunits with specific ethanol behaviors. We studied null mutant mice for six different GABAA-R subunits (α1, α2, α3, α4, α5 and δ). Only mice lacking the α2 subunit showed reduction of conditioned taste aversion (CTA) to ethanol. These results are in agreement with data from knock-in mice with mutation of the ethanol-sensitive site in the α2-subunit (Blednov et al., 2011) and indicate this aversive property of ethanol is dependent on ethanol action on α2-containing GABAA-R. Deletion of the α2-subunit led to faster recovery whereas absence of the α3-subunit slowed recovery from ethanol-induced incoordination (rotarod). Deletion of the other four subunits did not affect this behavior. Similar changes in this behavior for the α2 and α3 null mutants were found for flurazepam motor-incoordination. However, no differences in recovery were found in motor-incoordinating effects of an α1-selective modulator (zolpidem) or an α4-selective agonist (gaboxadol). Therefore, recovery of rotarod incoordination is under control of two GABAA-R subunits: α2 and α3. For motor activity, α3 null mice demonstrated higher activation by ethanol (1 g/kg) whereas both α2 and α3 (-/-) knockout mice were less sensitive to ethanol-induced reduction of motor activity (1.5 g/kg). These studies demonstrate that the effects of ethanol at GABAergic synapses containing α2 subunit are important for specific behavioral effects of ethanol which may be relevant to the genetic linkage of the α2 subunit with human alcoholism. PMID:23147414

The Role of GABAA Receptors in the Development of Alcoholism

PubMed Central

Enoch, Mary-Anne

2008-01-01

Alcoholism is a common, heritable, chronic relapsing disorder. GABAA receptors undergo allosteric modulation by ethanol, anesthetics, benzodiazepines and neurosteroids and have been implicated in the acute as well as the chronic effects of ethanol including tolerance, dependence and withdrawal. Medications targeting GABAA receptors ameliorate the symptoms of acute withdrawal. Ethanol induces plasticity in GABAA receptors: tolerance is associated with generally decreased GABAA receptor activation and differentially altered subunit expression. The dopamine (DA) mesolimbic reward pathway originating in the ventral tegmental area (VTA), and interacting stress circuitry play an important role in the development of addiction. VTA GABAergic interneurons are the primary inhibitory regulators of DA neurons and a subset of VTA GABAA receptors may be implicated in the switch from heavy drinking to dependence. GABAA receptors modulate anxiety and response to stress; important elements of sustained drinking and relapse. The GABAA receptor subunit genes clustered on chromosome 4 are highly expressed in the reward pathway. Several recent studies have provided strong evidence that one of these genes, GABRA2, is implicated in alcoholism in humans. The influence of the interaction between ethanol and GABAA receptors in the reward pathway on the development of alcoholism together with genetic and epigenetic vulnerabilities will be explored in this review. PMID:18440057

Increases in food intake or food-seeking behavior induced by GABAergic, opioid, or dopaminergic stimulation of the nucleus accumbens: is it hunger?

PubMed

Hanlon, Erin C; Baldo, Brian A; Sadeghian, Ken; Kelley, Ann E

2004-03-01

Previous work has shown that stimulation of GABAergic, opioid, or dopaminergic systems within the nucleus accumbens modulates food intake and food-seeking behavior. However, it is not known whether such stimulation mimics a motivational state of food deprivation that commonly enables animals to learn a new operant response to obtain food. In order to address this question, acquisition of lever pressing for food in hungry animals was compared with acquisition in non-food-deprived rats subjected to various nucleus accumbens drug treatments. All animals were given the opportunity to learn an instrumental response (a lever press) to obtain a food pellet. Prior to training, ad lib-fed rats were infused with the gamma-aminobutyric acid (GABA)A agonist muscimol (100 ng/0.5 microl per side) or the mu-opioid receptor agonist D-Ala2, N-me-Phe4, Gly-ol5-enkephalin (DAMGO, 0.25 microg/0.5 microl per side), or saline into the nucleus accumbens shell (AcbSh). The indirect dopamine agonist amphetamine (10 microg/0.5 microl per side) was infused into the AcbSh or nucleus accumbens core (AcbC) of ad lib-fed rats. An additional group was food deprived and infused with saline in the AcbSh. Chow and sugar pellet intake responses after drug treatments were also evaluated in free-feeding tests. Muscimol, DAMGO, or amphetamine did not facilitate acquisition of lever pressing for food, despite clearly increasing food intake in free-feeding tests. In contrast, food-deprived animals rapidly learned the task. These findings suggest that pharmacological stimulation of any of these neurochemical systems in isolation is insufficient to enable acquisition of a food-reinforced operant task. Thus, these selective processes, while likely involved in control of food intake and food-seeking behavior, appear unable to recapitulate the conditions necessary to mimic the state of negative energy balance.

The Central Amygdala Nucleus is Critical for Incubation of Methamphetamine Craving

PubMed Central

Li, Xuan; Zeric, Tamara; Kambhampati, Sarita; Bossert, Jennifer M; Shaham, Yavin

2015-01-01

Cue-induced methamphetamine seeking progressively increases after withdrawal but mechanisms underlying this ‘incubation of methamphetamine craving' are unknown. Here we studied the role of central amygdala (CeA), ventral medial prefrontal cortex (vmPFC), and orbitofrontal cortex (OFC), brain regions implicated in incubation of cocaine and heroin craving, in incubation of methamphetamine craving. We also assessed the role of basolateral amygdala (BLA) and dorsal medial prefrontal cortex (dmPFC). We trained rats to self-administer methamphetamine (10 days; 9 h/day, 0.1 mg/kg/infusion) and tested them for cue-induced methamphetamine seeking under extinction conditions during early (2 days) or late (4–5 weeks) withdrawal. We first confirmed that ‘incubation of methamphetamine craving' occurs under our experimental conditions. Next, we assessed the effect of reversible inactivation of CeA or BLA by GABAA+GABAB receptor agonists (muscimol+baclofen, 0.03+0.3 nmol) on cue-induced methamphetamine seeking during early and late withdrawal. We also assessed the effect of muscimol+baclofen reversible inactivation of vmPFC, dmPFC, and OFC on ‘incubated' cue-induced methamphetamine seeking during late withdrawal. Lever presses in the cue-induced methamphetamine extinction tests were higher during late withdrawal than during early withdrawal (incubation of methamphetamine craving). Muscimol+baclofen injections into CeA but not BLA decreased cue-induced methamphetamine seeking during late but not early withdrawal. Muscimol+baclofen injections into dmPFC, vmPFC, or OFC during late withdrawal had no effect on incubated cue-induced methamphetamine seeking. Together with previous studies, results indicate that the CeA has a critical role in incubation of both drug and non-drug reward craving and demonstrate an unexpected dissociation in mechanisms of incubation of methamphetamine vs cocaine craving. PMID:25475163

Selective GABA(A) α5 positive allosteric modulators improve cognitive function in aged rats with memory impairment.

PubMed

Koh, Ming Teng; Rosenzweig-Lipson, Sharon; Gallagher, Michela

2013-01-01

A condition of excess activity in the hippocampal formation is observed in the aging brain and in conditions that confer additional risk during aging for Alzheimer's disease. Compounds that act as positive allosteric modulators at GABA(A) α5 receptors might be useful in targeting this condition because GABA(A) α5 receptors mediate tonic inhibition of principal neurons in the affected network. While agents to improve cognitive function in the past focused on inverse agonists, which are negative allosteric modulators at GABA(A) α5 receptors, research supporting that approach used only young animals and predated current evidence for excessive hippocampal activity in age-related conditions of cognitive impairment. Here, we used two compounds, Compound 44 [6,6-dimethyl-3-(3-hydroxypropyl)thio-1-(thiazol-2-yl)-6,7-dihydro-2-benzothiophen-4(5H)-one] and Compound 6 [methyl 3,5-diphenylpyridazine-4-carboxylate], with functional activity as potentiators of γ-aminobutyric acid at GABA(A) α5 receptors, to test their ability to improve hippocampal-dependent memory in aged rats with identified cognitive impairment. Improvement was obtained in aged rats across protocols differing in motivational and performance demands and across varying retention intervals. Significant memory improvement occurred after either intracereboventricular infusion with Compound 44 (100 μg) in a water maze task or systemic administration with Compound 6 (3 mg/kg) in a radial arm maze task. Furthermore, systemic administration improved behavioral performance at dosing shown to provide drug exposure in the brain and in vivo receptor occupancy in the hippocampus. These data suggest a novel approach to improve neural network function in clinical conditions of excess hippocampal activity. This article is part of a Special Issue entitled 'Cognitive Enhancers'. Copyright © 2012 Elsevier Ltd. All rights reserved.

Independent of 5-HT1A receptors, neurons in the paraventricular hypothalamus mediate ACTH responses from MDMA

PubMed Central

Zaretsky, Dmitry V.; Zaretskaia, Maria V.; DiMicco, Joseph A.; Durant, Pamela J.; Ross, Christian T.; Rusyniak, Daniel E.

2013-01-01

Acute and chronic complications from the substituted amphetamine 3,4-methylenedioxymethamphetamine (MDMA) are linked to activation of the hypothalamic-pituitary-adrenal (HPA) axis. How MDMA activates the HPA axis is not known. HPA responses to stress are known to be mediated through the paraventricular (PVH) hypothalamus and to involve serotonin-1a (5-HT1A) receptors. We sought to determine if the PVH and 5-HT1A receptors were also involved in mediating HPA responses to MDMA. Rats were pretreated with either saline or a 5-HT1A antagonist, WAY-100635 (WAY), followed by a systemic dose of MDMA (7.5 mg/kg i.v.). Animals pretreated with WAY had significantly lower plasma ACTH concentrations after MDMA. To determine if neurons in the PVH were involved, and if their involvement was mediated by 5-HT1A receptors, rats implanted with guide cannulas targeting the PVH were microinjected with the GABAA receptor agonist muscimol, aCSF, or WAY followed by MDMA. Compared to aCSF microinjections of muscimol significantly attenuated the MDMA-induced rise in plasma ACTH (126 vs. 588 pg/ml, P=<0.01). WAY had no effect. Our data demonstrates that neurons in the PVH, independent of 5-HT1A receptors, mediate ACTH responses to MDMA. PMID:23933156

Evidence That GABA Mediates Dopaminergic and Serotonergic Pathways Associated with Locomotor Activity in Juvenile Chinook Salmon (Oncorhynchus tshawytscha)

USGS Publications Warehouse

Clements, S.; Schreck, C.B.

2004-01-01

The authors examined the control of locomotor activity in juvenile salmon (Oncorhynchus tshawytscha) by manipulating 3 neurotransmitter systems-gamma-amino-n-butyric acid (GABA), dopamine, and serotonin-as well as the neuropeptide corticotropin releasing hormone (CRH). Intracerebroventricular (ICV) injections of CRH and the GABAAagonist muscimol stimulated locomotor activity. The effect of muscimol was attenuated by administration of a dopamine receptor antagonist, haloperidol. Conversely, the administration of a dopamine uptake inhibitor (4???,4??? -difluoro-3-alpha-[diphenylmethoxy] tropane hydrochloride [DUI]) potentiated the effect of muscimol. They found no evidence that CRH-induced hyperactivity is mediated by dopaminergic systems following concurrent injections of haloperidol or DUI with CRH. Administration of muscimol either had no effect or attenuated the locomotor response to concurrent injections of CRH and fluoxetine, whereas the GABAA antagonist bicuculline methiodide potentiated the effect of CRH and fluoxetine.

The role of GABA(A) receptors in the development of alcoholism.

PubMed

Enoch, Mary-Anne

2008-07-01

Alcoholism is a common, heritable, chronic relapsing disorder. GABA(A) receptors undergo allosteric modulation by ethanol, anesthetics, benzodiazepines and neurosteroids and have been implicated in the acute as well as the chronic effects of ethanol including tolerance, dependence and withdrawal. Medications targeting GABA(A) receptors ameliorate the symptoms of acute withdrawal. Ethanol induces plasticity in GABA(A) receptors: tolerance is associated with generally decreased GABA(A) receptor activation and differentially altered subunit expression. The dopamine (DA) mesolimbic reward pathway originating in the ventral tegmental area (VTA), and interacting stress circuitry play an important role in the development of addiction. VTA GABAergic interneurons are the primary inhibitory regulators of DA neurons and a subset of VTA GABA(A) receptors may be implicated in the switch from heavy drinking to dependence. GABA(A) receptors modulate anxiety and response to stress; important elements of sustained drinking and relapse. The GABA(A) receptor subunit genes clustered on chromosome 4 are highly expressed in the reward pathway. Several recent studies have provided strong evidence that one of these genes, GABRA2, is implicated in alcoholism in humans. The influence of the interaction between ethanol and GABA(A) receptors in the reward pathway on the development of alcoholism together with genetic and epigenetic vulnerabilities will be explored in this review.

Role of GABAA receptors in the physiology and pharmacology of sleep.

PubMed

Winsky-Sommerer, Raphaëlle

2009-05-01

Most sedative-hypnotics used in insomnia treatment target the gamma-aminobutyric acid (GABA)(A) receptors. A vast repertoire of GABA(A) receptor subtypes has been identified and displays specific electrophysiological and functional properties. GABA(A)-mediated inhibition traditionally refers to 'phasic' inhibition, arising from synaptic GABA(A) receptors which transiently inhibit neurons. However, there is growing evidence that peri- or extra-synaptic GABA(A) receptors are continuously activated by low GABA concentrations and mediate a 'tonic' conductance. This slower type of signaling appears to play a key role in controlling cell excitability. This review aims at summarizing recent knowledge on GABA transmission, including the emergence of tonic conductance, and highlighting the importance of GABA(A) receptor heterogeneity. The mechanism of action of sedative-hypnotic drugs and their effects on sleep and the electroencephalogram will be reported. Furthermore, studies using genetically engineered mice will be emphasized, providing insights into the role of GABA(A) receptors in mechanisms underlying physiological and pharmacological sleep. Finally, we will address the potential of GABA(A) receptor pharmacology for the treatment of insomnia.

Fungal hallucinogens psilocin, ibotenic acid, and muscimol: analytical methods and biologic activities.

PubMed

Stebelska, Katarzyna

2013-08-01

Psychoactive drugs of fungal origin, psilocin, ibotenic acid, and muscimol among them have been proposed for recreational use and popularized since the 1960s, XX century. Despite their well-documented neurotoxicity, they reached reputation of being safe and nonaddictive. Scientific efforts to find any medical application for these hallucinogens in psychiatry, psychotherapy, and even for religious rituals support are highly controversial. Even if they show any healing potential, their usage in psychotherapy is in some cases inadequate and may additionally harm seriously suffering patients. Hallucinogens are thought to reduce cognitive functions. However, in case of indolealkylamines, such as psilocin, some recent findings suggest their ability to improve perception and mental skills, what would motivate the consumption of "magic mushrooms." The present article offers an opportunity to find out what are the main symptoms of intoxication with mushrooms containing psilocybin/psilocin, muscimol, and ibotenic acid. The progress in analytical methods for detection of them in fungal material, food, and body fluids is reviewed. Findings on the mechanisms of their biologic activity are summarized. Additionally, therapeutic potential of these fungal psychoactive compounds and health risk associated with their abuse are discussed.

Tobacco smoking interferes with GABAA receptor neuroadaptations during prolonged alcohol withdrawal

PubMed Central

Cosgrove, Kelly P.; McKay, Reese; Esterlis, Irina; Kloczynski, Tracy; Perkins, Evgenia; Bois, Frederic; Pittman, Brian; Lancaster, Jack; Glahn, David C.; O’Malley, Stephanie; Carson, Richard E.; Krystal, John H.

2014-01-01

Understanding the effects of tobacco smoking on neuroadaptations in GABAA receptor levels over alcohol withdrawal will provide critical insights for the treatment of comorbid alcohol and nicotine dependence. We conducted parallel studies in human subjects and nonhuman primates to investigate the differential effects of tobacco smoking and nicotine on changes in GABAA receptor availability during acute and prolonged alcohol withdrawal. We report that alcohol withdrawal with or without concurrent tobacco smoking/nicotine consumption resulted in significant and robust elevations in GABAA receptor levels over the first week of withdrawal. Over prolonged withdrawal, GABAA receptors returned to control levels in alcohol-dependent nonsmokers, but alcohol-dependent smokers had significant and sustained elevations in GABAA receptors that were associated with craving for alcohol and cigarettes. In nonhuman primates, GABAA receptor levels normalized by 1 mo of abstinence in both groups—that is, those that consumed alcohol alone or the combination of alcohol and nicotine. These data suggest that constituents in tobacco smoke other than nicotine block the recovery of GABAA receptor systems during sustained alcohol abstinence, contributing to alcohol relapse and the perpetuation of smoking. PMID:25453062

Increased efficiency of the GABAA and GABAB receptor–mediated neurotransmission in the Ts65Dn mouse model of Down syndrome

PubMed Central

Kleschevnikov, Alexander M.; Belichenko, Pavel V.; Gall, Jessica; George, Lizzy; Nosheny, Rachel; Maloney, Michael T.; Salehi, Ahmad; Mobley, William C.

2011-01-01

Cognitive impairment in Down syndrome (DS) involves the hippocampus. In the Ts65Dn mouse model of DS, deficits in hippocampus-dependent learning and synaptic plasticity were linked to enhanced inhibition. However, the mechanistic basis of changes in inhibitory efficiency remains largely unexplored, and efficiency of the GABAergic synaptic neurotransmission has not yet been investigated in direct electrophysiological experiments. To investigate this important feature of neurobiology of DS, we examined synaptic and molecular properties of the GABAergic system in the dentate gyrus (DG) of adult Ts65Dn mice. Both GABAA and GABAB receptor-mediated components of evoked inhibitory postsynaptic currents (IPSCs) were significantly increased in Ts65Dn vs. control (2N) DG granule cells. These changes were unaccompanied by alterations in hippocampal levels of GABAA (α1, α2, α3, α5 and γ2) or GABAB (Gbr1a and Gbr1b) receptor subunits. Immunoreactivity for GAD65, a marker for GABAergic terminals, was also unchanged. In contrast, there was a marked change in functional parameters of GABAergic synapses. Paired stimulations showed reduced paired-pulse ratios of both GABAA and GABAB receptor-mediated IPSC components (IPSC2/IPSC1), suggesting an increase in presynaptic release of GABA. Consistent with increased gene dose, the level of the Kir3.2 subunit of potassium channels, effectors for postsynaptic GABAB receptors, was increased. This change was associated with enhanced postsynaptic GABAB/Kir3.2 signaling following application of the GABAB receptor agonist baclofen. Thus, both GABAA and GABAB receptor-mediated synaptic efficiency is increased in the Ts65Dn DG, thus likely contributing to deficient synaptic plasticity and poor learning in DS. PMID:22062771

The α5 subunit-containing GABAA receptors contribute to chronic pain

PubMed Central

Bravo-Hernández, Mariana; Corleto, José A.; Barragán-Iglesias, Paulino; González-Ramírez, Ricardo; Pineda-Farias, Jorge B.; Felix, Ricardo; Calcutt, Nigel A.; Delgado-Lezama, Rodolfo; Marsala, Martin; Granados-Soto, Vinicio

2016-01-01

It has been recently proposed that α5-subunit containing GABAA receptors (α5-GABAA receptors) that mediate tonic inhibition might be involved in pain. The purpose of this study was to investigate the contribution of α5-GABAA receptors in the loss of GABAergic inhibition and in formalin-, Complete Freund’s adjuvant (CFA)- and L5/L6 spinal nerve ligation-induced long-lasting hypersensitivity. Formalin or CFA injection and L5/L6 spinal nerve ligation produced long-lasting allodynia and hyperalgesia. Moreover, formalin injection impaired the rate-dependent depression (RDD) of the Hofmann reflex. Peripheral and intrathecal pre-treatment or post-treatment with the α5-GABAA receptor antagonist, L-655,708 (0.15–15 nmol) prevented and reversed, respectively, these long-lasting behaviors. Formalin injection increased α5-GABAA receptors mRNA expression in the spinal cord and dorsal root ganglia (DRG) mainly at 3 days. α5-GABAA receptors were localized in the dorsal spinal cord and DRG co-labeling with NeuN, CGRP and IB4 suggesting their presence in peptidergic and non-peptidergic neurons. These receptors were found mainly in small- and medium-size neurons. Formalin injection enhanced α5-GABAA receptors fluorescence intensity in spinal cord and DRG at 3 and 6 days. Intrathecal administration of L-655,708 (15 nmol) prevented and reversed formalin-induced impairment of RDD. These results suggest that α5-GABAA receptors play a role in the loss of GABAergic inhibition and contribute to long-lasting secondary allodynia and hyperalgesia. PMID:26545088

Interneuron- and GABAA receptor-specific inhibitory synaptic plasticity in cerebellar Purkinje cells

NASA Astrophysics Data System (ADS)

He, Qionger; Duguid, Ian; Clark, Beverley; Panzanelli, Patrizia; Patel, Bijal; Thomas, Philip; Fritschy, Jean-Marc; Smart, Trevor G.

2015-07-01

Inhibitory synaptic plasticity is important for shaping both neuronal excitability and network activity. Here we investigate the input and GABAA receptor subunit specificity of inhibitory synaptic plasticity by studying cerebellar interneuron-Purkinje cell (PC) synapses. Depolarizing PCs initiated a long-lasting increase in GABA-mediated synaptic currents. By stimulating individual interneurons, this plasticity was observed at somatodendritic basket cell synapses, but not at distal dendritic stellate cell synapses. Basket cell synapses predominantly express β2-subunit-containing GABAA receptors; deletion of the β2-subunit ablates this plasticity, demonstrating its reliance on GABAA receptor subunit composition. The increase in synaptic currents is dependent upon an increase in newly synthesized cell surface synaptic GABAA receptors and is abolished by preventing CaMKII phosphorylation of GABAA receptors. Our results reveal a novel GABAA receptor subunit- and input-specific form of inhibitory synaptic plasticity that regulates the temporal firing pattern of the principal output cells of the cerebellum.

Induction of θ-frequency oscillations in the rat medial septal diagonal band slice by metabotropic glutamate receptor agonists.

PubMed

Lu, C B; Ouyang, G; Henderson, Z; Li, X

2011-03-17

The aim of this study was to examine the role of metabotropic glutamate receptors (mGluR) in the generation of oscillatory field activity at theta frequency (4-12 Hz) in the medial septal slice prepared from rat brain. Bath application of mGluR agonists and antagonists showed that activation of mGluR1-type receptors produces persistent theta frequency oscillations in a dose-responsive manner. This activity, induced by the group I mGluR agonist (RS)-3,5-dihydroxyphenylglycine (DHPG), was reduced by ionotropic glutamate receptor antagonists and abolished by further addition of a GABAA receptor antagonist. However, addition of a GABAA receptor antagonist on its own converted the DHPG-induced oscillations to intermittent episodes of accentuated theta frequency activity following a burst. In a proportion of slices, DHPG induced large amplitude field population spiking activity (100-300 μV) which is correlated linearly with the field theta oscillations and is sensitive to glutamate receptor antagonists, suggesting a role of this type of spikes in theta generation induced by DHPG. These data demonstrate that DHPG-sensitive neuronal networks within medial septum generate theta rhythmic activity and are differentially modulated by excitatory and inhibitory ionotropic neurotransmissions. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

Reversible inactivation of rostral nucleus raphe pallidus attenuates acute autonomic responses but not their habituation to repeated audiogenic stress in rats.

PubMed

Nyhuis, Tara J; Masini, Cher V; Taufer, Kirsten L; Day, Heidi E W; Campeau, Serge

2016-01-01

The medullary nucleus raphe pallidus (RPa) mediates several autonomic responses evoked by acute stress exposure, including tachycardia and hyperthermia. The present study assessed whether the RPa contributes to the decline/habituation of these responses observed during repeated audiogenic stress. Adult male rats were implanted with cannulae aimed at the RPa, and abdominal E-mitters that wirelessly acquire heart rate and core body temperature. After surgical recovery, animals were injected with muscimol or vehicle (aCSF) in the RPa region, followed by 30 min of 95-dBA loud noise or no noise control exposures on 3 consecutive days at 24-h intervals. Forty-eight hours after the third exposure, animals were exposed to an additional, but injection-free, loud noise or no noise test to assess habituation of hyperthermia and tachycardia. Three days later, rats were restrained for 30-min to evaluate their ability to display normal acute autonomic responses following the repeated muscimol injection regimen. The results indicated that the inhibition of cellular activity induced by the GABAA-receptor agonist muscimol centered in the RPa region reliably attenuated acute audiogenic stress-evoked tachycardia and hyperthermia, compared with vehicle-injected rats. Animals in the stress groups exhibited similar attenuated tachycardia and hyperthermia during the injection-free fourth audiogenic stress exposure, and displayed similar and robust increases in these responses to the subsequent restraint test. These results suggest that cellular activity in neurons of the RPa region is necessary for the expression of acute audiogenic stress-induced tachycardia and hyperthermia, but may not be necessary for the acquisition of habituated tachycardic responses to repeated stress.

Reversible inactivation of rostral nucleus raphe pallidus attenuates acute autonomic responses but not their habituation to repeated audiogenic stress in rats

PubMed Central

Nyhuis, Tara J.; Masini, Cher V.; Taufer, Kirsten L.; Day, Heidi E.W.; Campeau, Serge

2016-01-01

The medullary nucleus raphe pallidus (RPa) mediates several autonomic responses evoked by acute stress exposure, including tachycardia and hyperthermia. The present study assessed whether the RPa contributes to the decline/habituation of these responses observed during repeated audiogenic stress. Adult male rats were implanted with cannulae aimed at the RPa, and abdominal E-mitters that wirelessly acquire heart rate and core body temperature. After surgical recovery, animals were injected with muscimol or vehicle (aCSF) in the RPa region, followed by 30 minutes of 95-dBA loud noise or no noise control exposures on 3 consecutive days at 24-hr intervals. Forty-eight hours after the third exposure, animals were exposed to an additional, but injection-free, loud noise or no noise test to assess habituation of hyperthermia and tachycardia. Three days later, rats were restrained for 30-minutes to evaluate their ability to display normal acute autonomic responses following the repeated muscimol injection regimen. The results indicated that the inhibition of cellular activity induced by the GABAA-receptor agonist muscimol centered in the RPa region reliably attenuated acute audiogenic stress-evoked tachycardia and hyperthermia, compared with vehicle-injected rats. Animals in the stress groups exhibited similarly attenuated tachycardia and hyperthermia during the injection-free fourth audiogenic stress exposure, and displayed similar and robust increases in these responses to the subsequent restraint test. These results suggest that cellular activity in neurons of the RPa region is necessary for the expression of acute audiogenic stress-induced tachycardia and hyperthermia, but may not be necessary for the acquisition of habituated tachycardic responses to repeated stress. PMID:26998558

Trace and contextual fear conditioning require neural activity and NMDA receptor-dependent transmission in the medial prefrontal cortex

PubMed Central

Gilmartin, Marieke R.; Helmstetter, Fred J.

2010-01-01

The contribution of the medial prefrontal cortex (mPFC) to the formation of memory is a subject of considerable recent interest. Notably, the mechanisms supporting memory acquisition in this structure are poorly understood. The mPFC has been implicated in the acquisition of trace fear conditioning, a task that requires the association of a conditional stimulus (CS) and an aversive unconditional stimulus (UCS) across a temporal gap. In both rat and human subjects, frontal regions show increased activity during the trace interval separating the CS and UCS. We investigated the contribution of prefrontal neural activity in the rat to the acquisition of trace fear conditioning using microinfusions of the γ-aminobutyric acid type A (GABAA) receptor agonist muscimol. We also investigated the role of prefrontal N-methyl-d-aspartate (NMDA) receptor-mediated signaling in trace fear conditioning using the NMDA receptor antagonist 2-amino-5-phosphonovaleric acid (APV). Temporary inactivation of prefrontal activity with muscimol or blockade of NMDA receptor-dependent transmission in mPFC impaired the acquisition of trace, but not delay, conditional fear responses. Simultaneously acquired contextual fear responses were also impaired in drug-treated rats exposed to trace or delay, but not unpaired, training protocols. Our results support the idea that synaptic plasticity within the mPFC is critical for the long-term storage of memory in trace fear conditioning. PMID:20504949

Posterior insular cortex is necessary for conditioned inhibition of fear.

PubMed

Foilb, Allison R; Flyer-Adams, Johanna G; Maier, Steven F; Christianson, John P

2016-10-01

Veridical detection of safety versus danger is critical to survival. Learned signals for safety inhibit fear, and so when presented, reduce fear responses produced by danger signals. This phenomenon is termed conditioned inhibition of fear. Here, we report that CS+/CS- fear discrimination conditioning over 5 days in rats leads the CS- to become a conditioned inhibitor of fear, as measured by the classic tests of conditioned inhibition: summation and retardation of subsequent fear acquisition. We then show that NMDA-receptor antagonist AP5 injected to posterior insular cortex (IC) before training completely prevented the acquisition of a conditioned fear inhibitor, while intra-AP5 to anterior and medial IC had no effect. To determine if the IC contributes to the recall of learned fear inhibition, injections of the GABAA agonist muscimol were made to posterior IC before a summation test. This resulted in fear inhibition per se, which obscured inference to the effect of IC inactivation with recall of the safety cue. Control experiments sought to determine if the role of the IC in conditioned inhibition learning could be reduced to simpler fear discrimination function, but fear discrimination and recall were unaffected by AP5 or muscimol, respectively, in the posterior IC. These data implicate a role of posterior IC in the learning of conditioned fear inhibitors. Copyright © 2016 Elsevier Inc. All rights reserved.

17α-ethinyl estradiol attenuates depressive-like behavior through GABAA receptor activation/nitrergic pathway blockade in ovariectomized mice.

PubMed

Saeedi Saravi, Seyed Soheil; Arefidoust, Alireza; Yaftian, Rahele; Saeedi Saravi, Seyed Sobhan; Dehpour, Ahmad Reza

2016-04-01

This study was performed to investigate the antidepressant-like effect of 17α-ethinyl estradiol (EE2) in ovariectomized (OVX) mice and the possible role of nitrergic and gamma aminobutyric acid (GABA)ergic pathways in this paradigm. Bilateral ovariectomy was performed in female mice, and different doses of EE2 were intraperitoneally injected either alone or combined with GABAA agonist, diazepam, GABAA antagonist, flumazenil, non-specific nitric oxide synthase (NOS) inhibitor, N(ω)-nitro-L-arginine methyl ester (L-NAME), specific nNOS inhibitor, 7-nitroindazole (7-NI), a nitric oxide (NO) precursor, L-arginine, and selective PDE5I, sildenafil. After locomotion assessment, immobility times were recorded in the forced swimming test (FST) and tail suspension test (TST). Moreover, hippocampal nitrite concentrations were measured in the examined groups. Ten days after ovariectomy, a significant prolonged immobility times were observed. EE2 (0.3 and 1μg/kg and 0.03, 0.1, and 1mg/kg) caused antidepressant-like activity in OVX mice in FST and TST. Diazepam (1 and 5mg/kg), L-NAME (30mg/kg), and 7-NI (100mg/kg) significantly reduced the immobility times. Co-administration of minimal and sub-effective doses of EE2 and diazepam (0.3μg/kg and 0.5mg/kg, respectively) exerted a significant antidepressant-like effect. The same effect was observed in combination of minimal and sub-effective doses of EE2 and either L-NAME or 7-NI. Moreover, combination of minimal and sub-effective doses of EE2, diazepam either L-NAME, or 7-NI emphasized the significant robust antidepressant-like activity. The study has demonstrated that lowest dose of EE2 exerts a significant antidepressant-like behavior. It is suggested that suppression of NO system, as well as GABAA activation, may be responsible for antidepressant-like activity of EE2 in OVX mice. Moreover, GABAA activation may inhibit nitrergic pathway.

SH-I-048A, AN IN VITRO NONSELECTIVE SUPER-AGONIST AT THE BENZODIAZEPINE SITE OF GABAA RECEPTORS: THE APPROXIMATED ACTIVATION OF RECEPTOR SUBTYPES MAY EXPLAIN BEHAVIORAL EFFECTS

PubMed Central

Obradović, Aleksandar Lj.; Joksimović, Srđan; Poe, Michael M.; Ramerstorfer, Joachim; Varagic, Zdravko; Namjoshi, Ojas; Batinić, Bojan; Radulović, Tamara; Marković, Bojan; Roth, Brian; Sieghart, Werner; Cook, James M.; Savić, Miroslav M.

2014-01-01

Enormous progress in understanding the role of four populations of benzodiazepine-sensitive GABAA receptors was paralleled by the puzzling findings suggesting that substantial separation of behavioral effects may be accomplished by apparently non-selective modulators. We report on SH-I-048A, a newly-synthesized chiral positive modulator of GABAA receptors characterized by exceptional subnanomolar affinity, high efficacy and non-selectivity. Its influence on behavior was assessed in Wistar rats and contrasted to that obtained with 2 mg/kg diazepam. SH-I-048A reached micromolar concentrations in brain tissue, while the unbound fraction in brain homogenate was around 1.5%. The approximated electrophysiological responses, which estimated free concentrations of SH-I-048A or diazepam are able to elicit, suggested a similarity between the 10 mg/kg dose of the novel ligand and 2 mg/kg diazepam; however, SH-I-048A was relatively more active at α1- and α5-containing GABAA receptors. Behaviorally, SH-I-048A induced sedative, muscle relaxant and ataxic effects, reversed mechanical hyperalgesia 24 hours after injury, while it was devoid of clear anxiolytic actions and did not affect water-maze performance. While lack of clear anxiolytic actions may be connected with an enhanced potentiation at α1-containing GABAA receptors, the observed behavior in the rotarod, water maze and peripheral nerve injury tests was possibly affected by its prominent action at receptors containing the α5 subunit. The current results encourage further innovative approaches aimed at linking in vitro and in vivo data in order to help define fine-tuning mechanisms at four sensitive receptor populations that underlie subtle differences in behavioral profiles of benzodiazepine site ligands. PMID:24472579

Dissociating anxiolytic and sedative effects of GABAAergic drugs using temperature and locomotor responses to acute stress

PubMed Central

Klanker, Marianne; Groenink, Lucianne; Korte, S. Mechiel; Cook, James M.; Van Linn, Michael L.; Hopkins, Seth C.; Olivier, Berend

2009-01-01

Rationale The stress-induced hyperthermia (SIH) model is an anxiety model that uses the transient rise in body temperature in response to acute stress. Benzodiazepines produce anxiolytic as well as sedative side effects through nonselective binding to GABAA receptor subunits. The GABAA receptor α1 subunit is associated with sedation, whereas the GABAA receptor α2 and α3 subunits are involved in anxiolytic effects. Objectives We therefore examined the effects of (non) subunit-selective GABAA receptor agonists on temperature and locomotor responses to novel cage stress. Results Using telemetric monitoring of temperature and locomotor activity, we found that nonsubunit-selective GABAA receptor agonist diazepam as well as the α3 subunit-selective receptor agonist TP003 dose-dependently attenuated SIH and locomotor responses. Administration of GABAA receptor α1-selective agonist zolpidem resulted in profound hypothermia and locomotor sedation. The GABAA receptor α1-selective antagonist βCCt antagonized the hypothermia, but did not reverse the SIH response attenuation caused by diazepam and zolpidem. These results suggest an important regulating role for the α1 subunit in thermoregulation and sedation. Ligands of extrasynaptic GABAA receptors such as alcohol and nonbenzodiazepine THIP attenuated the SIH response only at high doses. Conclusions The present study confirms a putative role for the GABAA receptor α1 subunit in hypothermia and sedation and supports a role for α2/3 subunit GABAA receptor agonists in anxiety processes. In conclusion, we show that home cage temperature and locomotor responses to novel home cage stress provide an excellent tool to assess both anxiolytic and sedative effects of various (subunit-selective) GABAAergic compounds. PMID:19169673

Pharmacologic Treatment with GABAB Receptor Agonist of Methamphetamine-Induced Cognitive Impairment in Mice

PubMed Central

Mizoguchi, Hiroyuki; Yamada, Kiyofumi

2011-01-01

Methamphetamine (METH) is a highly addictive drug, and addiction to METH has increased to epidemic proportions worldwide. Chronic use of METH causes psychiatric symptoms, such as hallucinations and delusions, and long-term cognitive deficits, which are indistinguishable from paranoid schizophrenia. The GABA receptor system is known to play a significant role in modulating the dopaminergic neuronal system, which is related to behavioral changes induced by drug abuse. However, few studies have investigated the effects of GABA receptor agonists on cognitive deficits induced by METH. In the present review, we show that baclofen, a GABA receptor agonist, is effective in treating METH-induced impairment of object recognition memory and prepulse inhibition (PPI) of the startle reflex, a measure of sensorimotor gating in mice. Acute and repeated treatment with METH induced a significant impairment of PPI. Furthermore, repeated but not acute treatment of METH resulted in a long-lasting deficit of object recognition memory. Baclofen, a GABAB receptor agonist, dose-dependently ameliorated the METH-induced PPI deficits and object recognition memory impairment in mice. On the other hand, THIP, a GABAA receptor agonist, had no effect on METH-induced cognitive deficits. These results suggest that GABAB receptors may constitute a putative new target in treating cognitive deficits in chronic METH users. PMID:21886573

Context-Dependent Modulation of GABAAR-Mediated Tonic Currents

PubMed Central

Patel, Bijal; Bright, Damian P.; Mortensen, Martin; Frølund, Bente

2016-01-01

Tonic GABA currents mediated by high-affinity extrasynaptic GABAA receptors, are increasingly recognized as important regulators of cell and neuronal network excitability. Dysfunctional GABAA receptor signaling that results in modified tonic GABA currents is associated with a number of neurological disorders. Consequently, developing compounds to selectively modulate the activity of extrasynaptic GABAA receptors underlying tonic inhibition is likely to prove therapeutically useful. Here, we examine the GABAA receptor subtype selectivity of the weak partial agonist, 5-(4-piperidyl)isoxazol-3-ol (4-PIOL), as a potential mechanism for modulating extrasynaptic GABAA receptor-mediated tonic currents. By using recombinant GABAA receptors expressed in HEK293 cells, and native GABAA receptors of cerebellar granule cells, hippocampal neurons, and thalamic relay neurons, 4-PIOL evidently displayed differential agonist and antagonist-type profiles, depending on the extrasynaptic GABAA receptor isoforms targeted. For neurons, this resulted in differential modulation of GABA tonic currents, depending on the cell type studied, their respective GABAA receptor subunit compositions, and critically, on the ambient GABA levels. Unexpectedly, 4-PIOL revealed a significant population of relatively low-affinity γ2 subunit-containing GABAA receptors in the thalamus, which can contribute to tonic inhibition under specific conditions when GABA levels are raised. Together, these data indicate that partial agonists, such as 4-PIOL, may be useful for modulating GABAA receptor-mediated tonic currents, but the direction and extent of this modulation is strongly dependent on relative expression levels of different extrasynaptic GABAA receptor subtypes, and on the ambient GABA levels. SIGNIFICANCE STATEMENT A background level of inhibition (tonic) is important in the brain for controlling neuronal excitability. Increased levels of tonic inhibition are associated with some neurological disorders

Age- and sex-related characteristics of tonic GABA currents in the rat substantia nigra pars reticulata.

PubMed

Chudomel, O; Hasson, H; Bojar, M; Moshé, S L; Galanopoulou, A S

2015-04-01

Previous studies have shown that the pharmacologic effects of GABAergic drugs and the postsynaptic phasic GABAAergic inhibitory responses in the anterior part of the rat substantia nigra pars reticulata (SNRA) are age- and sex-specific. Here, we investigate whether there are age- and sex-related differences in the expression of the δ GABAA receptor (GABAAR) subunit and GABAAR mediated tonic currents. We have used δ-specific immunochemistry and whole cell patch clamp to study GABAAR mediated tonic currents in the SNRA of male and female postnatal day (PN) PN5-9, PN11-16, and PN25-32 rats. We observed age-related decline, but no sex-specific changes, in bicuculline (BIM) sensitive GABAAR tonic current density, which correlated with the decline in δ subunit in the SNRA between PN15 and 30. Furthermore, we show that the GABAAR tonic currents can be modified by muscimol (GABAAR agonist; partial GABACR agonist), THIP (4,5,6,7-tetrahydroisoxazolo (5,4-c)pyridin-3-ol: α4β3δ GABAARs agonist and GABACR antagonist), and zolpidem (α1-subunit selective GABAAR agonist) in age- and sex-dependent manner specific for each drug. We propose that the emergence of the GABAAR-sensitive anticonvulsant effects of the rat SNRA during development may depend upon the developmental decline in tonic GABAergic inhibition of the activity of rat SNRA neurons, although other sex-specific factors are also involved.

Microinjection of muscimol into the dorsomedial hypothalamus suppresses MDMA-evoked sympathetic and behavioral responses

PubMed Central

Rusyniak, Daniel E.; Zaretskaia, Maria V.; Zaretsky, Dmitry V.; DiMicco, Joseph A.

2008-01-01

When given systemically to rats and humans, the drug of abuse 3–4 methylenedioxymethamphetamine (ecstasy, MDMA) elicits hyperthermia, hyperactivity, tachycardia, and hypertension. Chemically stimulating the dorsomedial hypothalamus (DMH), a brain region known to be involved in thermoregulation and in stress responses, causes similar effects. We therefore tested the hypothesis that neuronal activity in the DMH plays a role in MDMA-evoked sympathetic and behavioral responses by microinjecting artificial CSF or muscimol, a neuronal inhibitor, into the DMH prior to intravenous infusion of saline or MDMA in conscious rats. Core temperature, heart rate, mean arterial pressure and locomotor activity were recorded by telemetry every minute for 120 minutes. In rats previously microinjected with CSF, MDMA elicited significant increases from baseline in core temperature (+1.3 ± 0.3°C), locomotion (+50 ± 6 counts/min), heart rate (+142 ± 16 beats/min), and mean arterial pressure (+26 ±3 mmHg). Microinjecting muscimol into the DMH prior to MDMA prevented increases in core temperature and locomotion and attenuated increases in heart rate and mean arterial pressure. These results indicate that neuronal activity in the DMH is necessary for the sympathetic and behavioral responses evoked by MDMA. PMID:18586013

Persistent spatial information in the FEF during object-based short-term memory does not contribute to task performance.

PubMed

Clark, Kelsey L; Noudoost, Behrad; Moore, Tirin

2014-06-01

We previously reported the existence of a persistent spatial signal in the FEF during object-based STM. This persistent activity reflected the location at which the sample appeared, irrespective of the location of upcoming targets. We hypothesized that such a spatial signal could be used to maintain or enhance object-selective memory activity elsewhere in cortex, analogous to the role of a spatial signal during attention. Here, we inactivated a portion of the FEF with GABAa agonist muscimol to test whether the observed activity contributes to object memory performance. We found that, although RTs were slowed for saccades into the inactivated portion of retinotopic space, performance for samples appearing in that region was unimpaired. This contrasts with the devastating effects of the same FEF inactivation on purely spatial working memory, as assessed with the memory-guided saccade task. Thus, in a task in which a significant fraction of FEF neurons displayed persistent, sample location-based activity, disrupting this activity had no impact on task performance.

Muscarinic Long-Term Enhancement of Tonic and Phasic GABAA Inhibition in Rat CA1 Pyramidal Neurons

PubMed Central

Domínguez, Soledad; Fernández de Sevilla, David; Buño, Washington

2016-01-01

Acetylcholine (ACh) regulates network operation in the hippocampus by controlling excitation and inhibition in rat CA1 pyramidal neurons (PCs), the latter through gamma-aminobutyric acid type-A receptors (GABAARs). Although, the enhancing effects of ACh on GABAARs have been reported (Dominguez et al., 2014, 2015), its role in regulating tonic GABAA inhibition has not been explored in depth. Therefore, we aimed at determining the effects of the activation of ACh receptors on responses mediated by synaptic and extrasynaptic GABAARs. Here, we show that under blockade of ionotropic glutamate receptors ACh, acting through muscarinic type 1 receptors, paired with post-synaptic depolarization induced a long-term enhancement of tonic GABAA currents (tGABAA) and puff-evoked GABAA currents (pGABAA). ACh combined with depolarization also potentiated IPSCs (i.e., phasic inhibition) in the same PCs, without signs of interactions of synaptic responses with pGABAA and tGABAA, suggesting the contribution of two different GABAA receptor pools. The long-term enhancement of GABAA currents and IPSCs reduced the excitability of PCs, possibly regulating plasticity and learning in behaving animals. PMID:27833531

Interactions of L-3,5,3'-Triiodothyronine, Allopregnanolone, and Ivermectin with the GABAA Receptor: Evidence for Overlapping Intersubunit Binding Modes

PubMed Central

Westergard, Thomas; Salari, Reza; Martin, Joseph V.; Brannigan, Grace

2015-01-01

Structural mechanisms of modulation of γ-aminobutyric acid (GABA) type A receptors by neurosteroids and hormones remain unclear. The thyroid hormone L-3,5,3’-triiodothyronine (T3) inhibits GABAA receptors at micromolar concentrations and has common features with neurosteroids such as allopregnanolone (ALLOP). Here we use functional experiments on α2β1γ2 GABAA receptors expressed in Xenopus oocytes to detect competitive interactions between T3 and an agonist (ivermectin, IVM) with a crystallographically determined binding site at subunit interfaces in the transmembrane domain of a homologous receptor (glutamate-gated chloride channel, GluCl). T3 and ALLOP also show competitive effects, supporting the presence of both a T3 and ALLOP binding site at one or more subunit interfaces. Molecular dynamics (MD) simulations over 200 ns are used to investigate the dynamics and energetics of T3 in the identified intersubunit sites. In these simulations, T3 molecules occupying all intersubunit sites (with the exception of the α-β interface) display numerous energetically favorable conformations with multiple hydrogen bonding partners, including previously implicated polar/acidic sidechains and a structurally conserved deformation in the M1 backbone. PMID:26421724

Involvement of ERK phosphorylation in brainstem neurons in modulation of swallowing reflex in rats

PubMed Central

Tsujimura, Takanori; Kondo, Masahiro; Kitagawa, Junichi; Tsuboi, Yoshiyuki; Saito, Kimiko; Tohara, Haruka; Ueda, Koichiro; Sessle, Barry J; Iwata, Koichi

2009-01-01

In order to evaluate the neuronal mechanisms underlying functional abnormalities of swallowing in orofacial pain patients, this study investigated the effects of noxious orofacial stimulation on the swallowing reflex, phosphorylated extracellular signal-regulated kinase (pERK) and γ-aminobutyric acid (GABA) immunohistochemical features in brainstem neurons, and also analysed the effects of brainstem lesioning and of microinjection of GABA receptor agonist or antagonist into the nucleus tractus solitarii (NTS) on the swallowing reflex in anaesthetized rats. The swallowing reflex elicited by topical administration of distilled water to the pharyngolaryngeal region was inhibited after capsaicin injection into the facial (whisker pad) skin or lingual muscle. The capsaicin-induced inhibitory effect on the swallowing reflex was itself depressed after the intrathecal administration of MAPK kinase (MEK) inhibitor. No change in the capsaicin-induced inhibitory effect was observed after trigeminal spinal subnucleus caudalis lesioning, but the inhibitory effect was diminished by paratrigeminal nucleus (Pa5) lesioning. Many pERK-like immunoreactive neurons in the NTS showed GABA immunoreactivity. The local microinjection of the GABAA receptor agonist muscimol into the NTS produced a significant reduction in swallowing reflex, and the capsaicin-induced depression of the swallowing reflex was abolished by microinjection of the GABAA receptor antagonist bicuculline into the NTS. The present findings suggest that facial skin–NTS, lingual muscle–NTS and lingual muscle–Pa5–NTS pathways are involved in the modulation of swallowing reflex by facial and lingual pain, respectively, and that the activation of GABAergic NTS neurons is involved in the inhibition of the swallowing reflex following noxious stimulation of facial and intraoral structures. PMID:19124539

GABAA Receptors, Anesthetics and Anticonvulsants in Brain Development

PubMed Central

Henschel, Oliver; Gipson, Keith E.; Bordey, Angelique

2008-01-01

GABA, acting via GABAA receptors, is well-accepted as the main inhibitory neurotransmitter of the mature brain, where it dampens neuronal excitability. The receptor's properties have been studied extensively, yielding important information about its structure, pharmacology, and regulation that are summarized in this review. Several GABAergic drugs have been commonly used as anesthetics, sedatives, and anticonvulsants for decades. However, findings that GABA has critical functions in brain development, in particular during the late embryonic and neonatal period, raise worthwhile questions regarding the side effects of GABAergic drugs that may lead to long-term cognitive deficits. Here, we will review some of these drugs in parallel with the control of CNS development that GABA exerts via activation of GABAA receptors. This review aims to provide a basic science and clinical perspective on the function of GABA and related pharmaceuticals acting at GABAA receptors. PMID:18537647

Interactions of L-3,5,3'-Triiodothyronine [corrected], Allopregnanolone, and Ivermectin with the GABAA Receptor: Evidence for Overlapping Intersubunit Binding Modes.

PubMed

Westergard, Thomas; Salari, Reza; Martin, Joseph V; Brannigan, Grace

2015-01-01

Structural mechanisms of modulation of γ-aminobutyric acid (GABA) type A receptors by neurosteroids and hormones remain unclear. The thyroid hormone L-3,5,3'-triiodothyronine (T3) inhibits GABAA receptors at micromolar concentrations and has common features with neurosteroids such as allopregnanolone (ALLOP). Here we use functional experiments on α2β1γ2 GABAA receptors expressed in Xenopus oocytes to detect competitive interactions between T3 and an agonist (ivermectin, IVM) with a crystallographically determined binding site at subunit interfaces in the transmembrane domain of a homologous receptor (glutamate-gated chloride channel, GluCl). T3 and ALLOP also show competitive effects, supporting the presence of both a T3 and ALLOP binding site at one or more subunit interfaces. Molecular dynamics (MD) simulations over 200 ns are used to investigate the dynamics and energetics of T3 in the identified intersubunit sites. In these simulations, T3 molecules occupying all intersubunit sites (with the exception of the α-β interface) display numerous energetically favorable conformations with multiple hydrogen bonding partners, including previously implicated polar/acidic sidechains and a structurally conserved deformation in the M1 backbone.

Development of F-18 Labeled Radiotracers for PET Imaging of Brain Alpha-1 Noradrenergic Receptors: Potential PTSD Vulnerability and Diagnostic Biomarkers

DTIC Science & Technology

2012-09-01

Amitriptyline D1 [3H]SCH233930 SKF38393 D2 [3H]N-methylspiperone Haloperidol D3 [3H]N-methylspiperone Chlorpromazine D4 [3H]N...Salvinorin A OPIOIDS Mu Opioid [3H]DAMGO DAMGO Sigma 1 [3H]Pentazocine Haloperidol UNCLEAR Sigma 2 [3H]DTG Haloperidol GABA GABAA [3H]Muscimol GABA

Anxiety and Depression: Mouse Genetics and Pharmacological Approaches to the Role of GABAA Receptor Subtypes

PubMed Central

Smith, Kiersten S.; Rudolph, Uwe

2012-01-01

GABAA receptors mediate fast synaptic inhibitory neurotransmission throughout the central nervous system. Recent work indicates a role for GABAA receptors in physiologically modulating anxiety and depression levels. In this review, we summarize research that led to the identification of the essential role of GABAA receptors in counteracting trait anxiety and depression-related behaviors, and research aimed at identifying individual GABAA receptor subtypes involved in physiological and pharmacological modulation of emotions. PMID:21810433

Context-Dependent Modulation of GABAAR-Mediated Tonic Currents.

PubMed

Patel, Bijal; Bright, Damian P; Mortensen, Martin; Frølund, Bente; Smart, Trevor G

2016-01-13

Tonic GABA currents mediated by high-affinity extrasynaptic GABAA receptors, are increasingly recognized as important regulators of cell and neuronal network excitability. Dysfunctional GABAA receptor signaling that results in modified tonic GABA currents is associated with a number of neurological disorders. Consequently, developing compounds to selectively modulate the activity of extrasynaptic GABAA receptors underlying tonic inhibition is likely to prove therapeutically useful. Here, we examine the GABAA receptor subtype selectivity of the weak partial agonist, 5-(4-piperidyl)isoxazol-3-ol (4-PIOL), as a potential mechanism for modulating extrasynaptic GABAA receptor-mediated tonic currents. By using recombinant GABAA receptors expressed in HEK293 cells, and native GABAA receptors of cerebellar granule cells, hippocampal neurons, and thalamic relay neurons, 4-PIOL evidently displayed differential agonist and antagonist-type profiles, depending on the extrasynaptic GABAA receptor isoforms targeted. For neurons, this resulted in differential modulation of GABA tonic currents, depending on the cell type studied, their respective GABAA receptor subunit compositions, and critically, on the ambient GABA levels. Unexpectedly, 4-PIOL revealed a significant population of relatively low-affinity γ2 subunit-containing GABAA receptors in the thalamus, which can contribute to tonic inhibition under specific conditions when GABA levels are raised. Together, these data indicate that partial agonists, such as 4-PIOL, may be useful for modulating GABAA receptor-mediated tonic currents, but the direction and extent of this modulation is strongly dependent on relative expression levels of different extrasynaptic GABAA receptor subtypes, and on the ambient GABA levels. A background level of inhibition (tonic) is important in the brain for controlling neuronal excitability. Increased levels of tonic inhibition are associated with some neurological disorders but there are no

Plasticity of spatial hearing: behavioural effects of cortical inactivation

PubMed Central

Nodal, Fernando R; Bajo, Victoria M; King, Andrew J

2012-01-01

The contribution of auditory cortex to spatial information processing was explored behaviourally in adult ferrets by reversibly deactivating different cortical areas by subdural placement of a polymer that released the GABAA agonist muscimol over a period of weeks. The spatial extent and time course of cortical inactivation were determined electrophysiologically. Muscimol-Elvax was placed bilaterally over the anterior (AEG), middle (MEG) or posterior ectosylvian gyrus (PEG), so that different regions of the auditory cortex could be deactivated in different cases. Sound localization accuracy in the horizontal plane was assessed by measuring both the initial head orienting and approach-to-target responses made by the animals. Head orienting behaviour was unaffected by silencing any region of the auditory cortex, whereas the accuracy of approach-to-target responses to brief sounds (40 ms noise bursts) was reduced by muscimol-Elvax but not by drug-free implants. Modest but significant localization impairments were observed after deactivating the MEG, AEG or PEG, although the largest deficits were produced in animals in which the MEG, where the primary auditory fields are located, was silenced. We also examined experience-induced spatial plasticity by reversibly plugging one ear. In control animals, localization accuracy for both approach-to-target and head orienting responses was initially impaired by monaural occlusion, but recovered with training over the next few days. Deactivating any part of the auditory cortex resulted in less complete recovery than in controls, with the largest deficits observed after silencing the higher-level cortical areas in the AEG and PEG. Although suggesting that each region of auditory cortex contributes to spatial learning, differences in the localization deficits and degree of adaptation between groups imply a regional specialization in the processing of spatial information across the auditory cortex. PMID:22547635

Differential effects of short- and long-term zolpidem treatment on recombinant α1β2γ2s subtype of GABAA receptors in vitro

PubMed Central

Vlainić, Josipa; Jembrek, Maja Jazvinšćak; Vlainić, Toni; Štrac, Dubravka Švob; Peričić, Danka

2012-01-01

Aim: Zolpidem is a non-benzodiazepine agonist at benzodiazepine binding site in GABAA receptors, which is increasingly prescribed. Recent studies suggest that prolonged zolpidem treatment induces tolerance. The aim of this study was to explore the adaptive changes in GABAA receptors following short and long-term exposure to zolpidem in vitro. Methods: Human embryonic kidney (HEK) 293 cells stably expressing recombinant α1β2γ2s GABAA receptors were exposed to zolpidem (1 and 10 μmol/L) for short-term (2 h daily for 1, 2, or 3 consecutive days) or long-term (continuously for 48 h). Radioligand binding studies were used to determine the parameters of [3H]flunitrazepam binding sites. Results: A single (2 h) or repeated (2 h daily for 2 or 3 d) short-term exposure to zolpidem affected neither the maximum number of [3H]flunitrazepam binding sites nor the affinity. In both control and short-term zolpidem treated groups, addition of GABA (1 nmol/L–1 mmol/L) enhanced [3H]flunitrazepam binding in a concentration-dependent manner. The maximum enhancement of [3H]flunitrazepam binding in short-term zolpidem treated group was not significantly different from that in the control group. In contrast, long-term exposure to zolpidem resulted in significantly increase in the maximum number of [3H]flunitrazepam binding sites without changing the affinity. Furthermore, long-term exposure to zolpidem significantly decreased the ability of GABA to stimulate [3H]flunitrazepam binding. Conclusion: The results suggest that continuous, but not intermittent and short-term, zolpidem-exposure is able to induce adaptive changes in GABAA receptors that could be related to the development of tolerance and dependence. PMID:22922343

Decreased hepatocyte membrane potential differences and GABAA-beta3 expression in human hepatocellular carcinoma.

PubMed

Minuk, Gerald Y; Zhang, Manna; Gong, Yuewen; Minuk, Leonard; Dienes, Hans; Pettigrew, Norman; Kew, Michael; Lipschitz, Jeremy; Sun, Dongfeng

2007-03-01

To determine whether hepatocyte membrane potential differences (PDs) are depolarized in human HCC and whether depolarization is associated with changes in GABAA receptor expression, hepatocyte PDs and gamma-aminobutyric acid (GABA)A receptor messenger RNA (mRNA) and protein expression were documented in HCC tissues via microelectrode impalement, real-time reverse-transcriptase polymerase chain reaction, and Western blot analysis, respectively. HCC tissues were significantly depolarized (-19.8+/-1.3 versus -25.9+/-3.2 mV, respectively [P<0.05]), and GABAA-beta3 expression was down-regulated (GABAA-beta3 mRNA and protein expression in HCC; 5,693+/-1,385 and 0.29+/-0.11 versus 11,046+/-4,979 copies/100 mg RNA and 0.62+/-0.16 optical density in adjacent tumor tissues, respectively [P=0.002 and P<0.0001, respectively]) when compared with adjacent nontumor tissues. To determine the physiological relevance of the down-regulation, human malignant hepatocytes deficient in GABAA-beta3 receptor expression (Huh-7 cells) were transfected with GABAA-beta3 complementary DNA (cDNA) or vector alone and injected into nu/nu nude mice (n=16-17 group). Tumors developed after a mean (+/-SD) of 51+/-6 days (range: 41-60 days) in 7/16 (44%) mice injected with vector-transfected cells and 70+/-12 days (range: 59-86 days) in 4/17 (24%) mice injected with GABAA-beta3 cDNA-transfected cells (P<0.005). The results of this study indicate that (1) human HCC tissues are depolarized compared with adjacent nontumor tissues, (2) hepatic GABAA-beta3 receptor expression is down-regulated in human HCC, and (3) restoration of GABAA-beta3 receptor expression results in attenuated in vivo tumor growth in nude mice.

An Allosteric Coagonist Model for Propofol Effects on α1β2γ2L γ-Aminobutyric Acid Type A Receptors

PubMed Central

Ruesch, Dirk; Neumann, Elena; Wulf, Hinnerk; Forman, Stuart A.

2011-01-01

Background Propofol produces its major actions via γ-aminobutyric acid type A (GABAA) receptors. At low concentrations, propofol enhances agonist-stimulated GABAA receptor activity, and high propofol concentrations directly activate receptors. Etomidate produces similar effects, and there is convincing evidence that a single class of etomidate sites mediate both agonist modulation and direct GABAA receptor activation. It is unknown if the propofol binding site(s) on GABAA receptors that modulate agonist-induced activity also mediate direct activation. Methods GABAA α1β2γ2L receptors were heterologously expressed in Xenopus oocytes and activity was quantified using voltage clamp electrophysiology. We tested whether propofol and etomidate display the same linkage between agonist modulation and direct activation of GABAA receptors by identifying equi-efficacious drug solutions for direct activation. We then determined whether these drug solutions produce equal modulation of GABA-induced receptor activity. We also measured propofol-dependent direct activation and modulation of low GABA responses. Allosteric coagonist models similar to that established for etomidate, but with variable numbers of propofol sites, were fitted to combined data. Results Solutions of 19 μM propofol and 10 μM etomidate were found to equally activate GABAA receptors. These two drug solutions also produced indistinguishable modulation of GABA-induced receptor activity. Combined electrophysiological data behaved in a manner consistent with allosteric co-agonist models with more than one propofol site. The best fit was observed when the model assumed three equivalent propofol sites. Conclusions Our results support the hypothesis that propofol, like etomidate, acts at GABAA receptor sites mediating both GABA modulation and direct activation. PMID:22104494

SB-205384 Is a Positive Allosteric Modulator of Recombinant GABAA Receptors Containing Rat α3, α5, or α6 Subunit Subtypes Coexpressed with β3 and γ2 Subunits

PubMed Central

Heidelberg, Laura S.; Warren, James W.

2013-01-01

Many drugs used to treat anxiety are positive modulators of GABAA receptors, which mediate fast inhibitory neurotransmission. The GABAA receptors can be assembled from a combination of at least 16 different subunits. The receptor’s subunit composition determines its pharmacologic and functional properties, and subunit expression varies throughout the brain. A primary goal for new treatments targeting GABAA receptors is the production of subunit-selective modulators acting upon a discrete population of receptors. The anxiolytic 4-amino-7-hydroxy-2-methyl-5,6,7,8,-tetrahydrobenzo[b]thieno[2,3-b]pyridine-3-carboxylic acid, but-2-ynyl ester (SB-205384) is widely considered to be selective for α3-containing GABAA receptors. However, it has been tested only on α1-, α2-, and α3-containing receptors. We examined the activity of SB-205384 at recombinant receptors containing the six different α subunits and found that receptors containing the α3, α5, and α6 subunits were potentiated by SB-205384, with the α6 subunit conferring the greatest responsiveness. Properties associated with chimeric α1/α6 subunits suggested that multiple structural domains influence sensitivity to SB-205384. Point mutations of residues within the extracellular N-terminal domain identified a leucine residue located in loop E of the agonist binding site as an important determinant of high sensitivity to modulation. In the α6 subunit the identity of this residue is species-dependent, with the leucine found in rat subunits but not in human. Our results indicate that SB-205384 is not an α3-selective modulator, and instead acts at several GABAA receptor isoforms. These findings have implications for the side-effect profile of this anxiolytic as well as for its use in neuronal and animal studies as a marker for contribution from α3-containing receptors. PMID:23902941

Opioid systems in the lateral hypothalamus regulate feeding behavior through orexin and GABA neurons.

PubMed

Ardianto, C; Yonemochi, N; Yamamoto, S; Yang, L; Takenoya, F; Shioda, S; Nagase, H; Ikeda, H; Kamei, J

2016-04-21

The hypothalamus controls feeding behavior. Since central opioid systems may regulate feeding behavior, we examined the role of μ-, δ- and κ-opioid receptors in the lateral hypothalamus (LH), the hunger center, in feeding behavior of mice. Non-selective (naloxone; 3 mg/kg, s.c.) and selective μ- (β-funaltrexamine, β-FNA; 10 mg/kg, s.c.), δ- (naltrindole; 3 mg/kg, s.c.) and κ- (norbinaltorphimine, norBNI; 20 mg/kg, s.c.) opioid receptor antagonists significantly decreased food intake in food-deprived mice. The injection of naloxone (20 μg/side) into the LH significantly decreased food intake whereas the injection of naloxone (20 μg/side) outside of the LH did not affect food intake. The injection of β-FNA (2 μg/side), naltrindole (1 μg/side) or norBNI (2 μg/side) into the LH significantly decreased food intake. Furthermore, all these antagonists significantly decreased the mRNA level of preproorexin, but not those of other hypothalamic neuropeptides. In addition, the injection of the GABAA receptor agonist muscimol (5 μg/side) into the LH significantly decreased food intake, and this effect was abolished by the GABAA receptor antagonist bicuculline (50 μg/side). Muscimol (1mg/kg, i.p.) decreased the mRNA level of preproorexin in the hypothalamus. Naloxone (3mg/kg, s.c.) significantly increased the GABA level in the LH and both bicuculline and the GABA release inhibitor 3-mercaptopropionic acid (3-MP, 5 μg/side) attenuated the inhibitory effect of naloxone on feeding behavior. 3-MP also attenuated the effects of β-FNA and norBNI, but not that of naltrindole. These results show that opioid systems in the LH regulate feeding behavior through orexin neurons. Moreover, μ- and κ-, but not δ-, opioid receptor antagonists inhibit feeding behavior by activating GABA neurons in the LH. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Transient inactivation of basolateral amygdala during selective satiation disrupts reinforcer devaluation in rats

PubMed Central

West, Elizabeth A.; Forcelli, Patrick A.; Murnen, Alice T.; McCue, David L.; Gale, Karen; Malkova, Ludise

2012-01-01

Basolateral amygdala (BLA) function is critical for flexible, goal-directed behavior, including performance on reinforcer devaluation tasks. Here we tested, in rats, the hypothesis that BLA is critical for conditioned reinforcer devaluation during the period when the primary reinforcer (food) is being devalued (by feeding it to satiety), but not thereafter for guiding behavioral choices. We used a spatially-independent task, which employed two visual cues, each predicting one of two foods. An instrumental action (lever press) was required for reinforcer delivery. After training, rats received BLA or sham lesions, or cannulae implanted in BLA. Under control conditions (sham lesions, saline infusions), devaluation of one food significantly decreased responding to the cue associated with that food, when both cues were presented simultaneously during extinction. BLA lesions impaired this devaluation effect. Transient inactivation of BLA by microinfusion of the GABAA agonist muscimol resulted in an impairment, only when BLA was inactivated during satiation. When muscimol was infused after satiation and, therefore, BLA was inactivated only during the choice test, rats showed no impairment. Thus, BLA is necessary for registering or updating cues to reflect updated reinforcer values, but not for guiding choices once the value has been updated. Our results are the first to describe the contribution of rat BLA to specific components of reinforcer devaluation, and are the first to show impairment in reinforcer devaluation following transient inactivation in the rat. PMID:22845705

Potentiation of tonic GABAergic inhibition by activation of postsynaptic kainate receptors.

PubMed

Jiang, L; Kang, D; Kang, J

2015-07-09

Presynaptic kainate-type glutamate ionotropic receptors (KARs) that mediate either the depression or the facilitation of GABA release have been intensively studied. Little attention has been given to the modulation of GABAA receptors (GABAARs) by postsynaptic KARs. Recent studies suggest that two GABAAR populations, synaptic (sGABAAR) and extrasynaptic (eGABAAR) GABAARs, mediate phasic and tonic forms of inhibition, respectively. Tonic inhibition plays an important role in the excitability of neuronal circuits and the occurrence of epileptic seizures. For this study, we are the first to report that the activation of postsynaptic KARs by the KAR agonist, Kainic acid (KA, 5 μM), enhanced tonic inhibition by potentiating eGABAARs. KA enhanced THIP-induced eGABAAR currents and prolonged the rise and decay time of muscimol-induced sGABAAR/eGABAAR currents, but also depressed the amplitude of evoked inhibitory postsynaptic currents (IPSCs), unitary IPSCs (uIPSCs), and muscimol-induced sGABAAR/eGABAAR currents. The PKC inhibitor, staurosporine (1 μM), in the patch pipette solution fully blocked the KA-induced potentiation of tonic inhibition, suggesting the involvement of an intracellular PKC pathway. Our study suggests that the activation of postsynaptic KARs potentiates eGABAARs but depresses sGABAARs. By activating postsynaptic KARs, synaptically released glutamate depresses phasic inhibition to facilitate neuronal plasticity, but potentiates tonic inhibition to protect neurons from over-excitation. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Brain functional network changes following Prelimbic area inactivation in a spatial memory extinction task.

PubMed

Méndez-Couz, Marta; Conejo, Nélida M; Vallejo, Guillermo; Arias, Jorge L

2015-01-01

Several studies suggest a prefrontal cortex involvement during the acquisition and consolidation of spatial memory, suggesting an active modulating role at late stages of acquisition processes. Recently, we have reported that the prelimbic and infralimbic areas of the prefrontal cortex, among other structures, are also specifically involved in the late phases of spatial memory extinction. This study aimed to evaluate whether the inactivation of the prelimbic area of the prefrontal cortex impaired spatial memory extinction. For this purpose, male Wistar rats were implanted bilaterally with cannulae into the prelimbic region of the prefrontal cortex. Animals were trained during 5 consecutive days in a hidden platform task and tested for reference spatial memory immediately after the last training session. One day after completing the training task, bilateral infusion of the GABAA receptor agonist Muscimol was performed before the extinction protocol was carried out. Additionally, cytochrome c oxidase histochemistry was applied to map the metabolic brain activity related to the spatial memory extinction under prelimbic cortex inactivation. Results show that animals acquired the reference memory task in the water maze, and the extinction task was successfully completed without significant impairment. However, analysis of the functional brain networks involved by cytochrome oxidase activity interregional correlations showed changes in brain networks between the group treated with Muscimol as compared to the saline-treated group, supporting the involvement of the mammillary bodies at a the late stage in the memory extinction process. Copyright © 2015 Elsevier B.V. All rights reserved.

Functional inactivation of the orbitofrontal cortex disrupts context-induced reinstatement of alcohol seeking in rats.

PubMed

Bianchi, Paula Cristina; Carneiro de Oliveira, Paulo Eduardo; Palombo, Paola; Leão, Rodrigo Molini; Cogo-Moreira, Hugo; Planeta, Cleopatra da Silva; Cruz, Fábio Cardoso

2018-05-01

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. 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. 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. Copyright © 2018 Elsevier B.V. All rights reserved.

Evidence for the involvement of extinction-associated inhibitory learning in the forced swimming test.

PubMed

Campus, P; Colelli, V; Orsini, C; Sarra, D; Cabib, S

2015-02-01

The forced swimming test (FST) remains one of the most used tools for screening antidepressants in rodent models. Nonetheless, the nature of immobility, its main behavioral measure, is still a matter of debate. The present study took advantage of our recent finding that mice of the inbred DBA/2J strain require a functioning left dorsolateral striatum (DLS) to consolidate long-term memory of FST to test whether immobility is the outcome of stress-related learning. Infusion of the GABA-A agonist muscimol in the left DLS immediately after a single experience of FST prevented and infusion in the left or the right amygdala impaired recall of the acquired levels of immobility in a probe test performed 24h later. Post-training left DLS infusion of muscimol, at a dose capable of preventing retention of FST-induced immobility, did not influence 24h retention of inhibitory avoidance training or of the escape response acquired in a water T-maze. However, this same treatment prevented 24h retention of the extinction training of the consolidated escape response. These results indicate that a left DLS-centered memory system selectively mediates memory consolidation of FST and of escape extinction and support the hypothesis that immobility is the result of extinction-like inhibitory learning involving all available escape responses due to the inescapable/unavoidable nature of FST experience. Copyright © 2014 Elsevier B.V. All rights reserved.

Modulatory Effects of Eschscholzia californica Alkaloids on Recombinant GABAA Receptors

PubMed Central

Fedurco, Milan; Gregorová, Jana; Šebrlová, Kristýna; Kantorová, Jana; Peš, Ondřej; Baur, Roland; Sigel, Erwin; Táborská, Eva

2015-01-01

The California poppy (Eschscholzia californica Cham.) contains a variety of natural compounds including several alkaloids found exclusively in this plant. Because of the sedative, anxiolytic, and analgesic effects, this herb is currently sold in pharmacies in many countries. However, our understanding of these biological effects at the molecular level is still lacking. Alkaloids detected in E. californica could be hypothesized to act at GABAA receptors, which are widely expressed in the brain mainly at the inhibitory interneurons. Electrophysiological studies on a recombinant α 1 β 2 γ 2 GABAA receptor showed no effect of N-methyllaurotetanine at concentrations lower than 30 μM. However, (S)-reticuline behaved as positive allosteric modulator at the α 3, α 5, and α 6 isoforms of GABAA receptors. The depressant properties of aerial parts of E. californica are assigned to chloride-current modulation by (S)-reticuline at the α 3 β 2 γ 2 and α 5 β 2 γ 2 GABAA receptors. Interestingly, α 1, α 3, and α 5 were not significantly affected by (R)-reticuline, 1,2-tetrahydroreticuline, codeine, and morphine—suspected (S)-reticuline metabolites in the rodent brain. PMID:26509084

Prefrontal Cortical GABA Modulation of Spatial Reference and Working Memory

PubMed Central

Auger, Meagan L.

2015-01-01

Background: Dysfunction in prefrontal cortex (PFC) GABA transmission has been proposed to contribute to cognitive dysfunction in schizophrenia, yet how this system regulates different cognitive and mnemonic functions remains unclear. Methods: We assessed the effects of pharmacological reduction of GABAA signaling in the medial PFC of rats on spatial reference/working memory using different versions of the radial-arm maze task. We used a massed-trials procedure to probe how PFC GABA regulates susceptibility to proactive interference. Male rats were well-trained to retrieve food from the same 4 arms of an 8-arm maze, receiving 5 trials/day (1–2min intervals). Results: Infusions of the GABAA receptor antagonist bicuculline (12.5–50ng) markedly increased working and reference memory errors and response latencies. Similar treatments also impaired short-term memory on an 8-baited arm task. These effects did not appear to be due to increased susceptibility to proactive interference. In contrast, PFC inactivation via infusion of GABA agonists baclofen/muscimol did not affect reference/working memory. In comparison to the pronounced effects on the 8-arm maze tasks, PFC GABAA antagonism only causes a slight and transient decrease in accuracy on a 2-arm spatial discrimination. Conclusions: These findings demonstrate that prefrontal GABA hypofunction severely disrupts spatial reference and short-term memory and that disinhibition of the PFC can, in some instances, perturb memory processes not normally dependent on the frontal lobes. Moreover, these impairments closely resemble those observed in schizophrenic patients, suggesting that perturbation in PFC GABA signaling may contribute to these types of cognitive deficits associated with the disorder. PMID:25552433

Prefrontal cortical GABA modulation of spatial reference and working memory.

PubMed

Auger, Meagan L; Floresco, Stan B

2014-10-31

Dysfunction in prefrontal cortex (PFC) GABA transmission has been proposed to contribute to cognitive dysfunction in schizophrenia, yet how this system regulates different cognitive and mnemonic functions remains unclear. We assessed the effects of pharmacological reduction of GABAA signaling in the medial PFC of rats on spatial reference/working memory using different versions of the radial-arm maze task. We used a massed-trials procedure to probe how PFC GABA regulates susceptibility to proactive interference. Male rats were well-trained to retrieve food from the same 4 arms of an 8-arm maze, receiving 5 trials/day (1-2 min intervals). Infusions of the GABAA receptor antagonist bicuculline (12.5-50 ng) markedly increased working and reference memory errors and response latencies. Similar treatments also impaired short-term memory on an 8-baited arm task. These effects did not appear to be due to increased susceptibility to proactive interference. In contrast, PFC inactivation via infusion of GABA agonists baclofen/muscimol did not affect reference/working memory. In comparison to the pronounced effects on the 8-arm maze tasks, PFC GABAA antagonism only causes a slight and transient decrease in accuracy on a 2-arm spatial discrimination. These findings demonstrate that prefrontal GABA hypofunction severely disrupts spatial reference and short-term memory and that disinhibition of the PFC can, in some instances, perturb memory processes not normally dependent on the frontal lobes. Moreover, these impairments closely resemble those observed in schizophrenic patients, suggesting that perturbation in PFC GABA signaling may contribute to these types of cognitive deficits associated with the disorder. © The Author 2014. Published by Oxford University Press on behalf of CINP.

Anxiolytic-like actions of the hexane extract from leaves of Annona cherimolia in two anxiety paradigms: possible involvement of the GABA/benzodiazepine receptor complex.

PubMed

López-Rubalcava, C; Piña-Medina, B; Estrada-Reyes, R; Heinze, G; Martínez-Vázquez, M

2006-01-11

A hexane extract of leaves of Annona cherimolia produced anxiolytic-like actions when administered to mice and tested in two animal models of anxiety: the mouse avoidance exploratory behavior and the burying behavior tests. In order to discard unspecific drug-actions on general activity, all treatments studied in the anxiety paradigms were also analyzed in the open field test. Results showed that A. cherimolia induced anxiolytic-like actions at the doses of 6.25, 12.5, 25.0 and 50.0 mg/kg. Picrotoxin (0.25 mg/kg), a GABA-gated chloride ion channel blocker, antagonized the anxiolytic-like actions of A. cherimolia, while a sub-effective dose of muscimol (0.5 mg/kg), a selective GABA(A) receptor agonist, facilitated the effects of a sub-optimal dose of A. cherimolia (3.12 mg/kg). Thus, the involvement of the GABA(A) receptor complex in the anxiolytic-like actions of A. cherimolia hexane extract is suggested. In addition the extract was also able to enhance the duration of sodium pentobarbital induced sleeping time. Taken together, results indicate that the hexane extract of A. cherimolia has depressant activity on the Central Nervous System and could interact with the GABA(A) receptor complex. On the other hand, the chromatographic separation of this extract led to the isolation of palmitone, and beta-sitosterol as major constituents. In addition a GC-MS study of some fractions revealed the presence of several compounds such beta-cariophyllene, beta-selinene, alpha-cubebene, and linalool that have been reported to show effects on behavior that could explain some of the extract effects.

A novel GABA(A) alpha 5 receptor inhibitor with therapeutic potential.

PubMed

Ling, István; Mihalik, Balázs; Etherington, Lori-An; Kapus, Gábor; Pálvölgyi, Adrienn; Gigler, Gábor; Kertész, Szabolcs; Gaál, Attila; Pallagi, Katalin; Kiricsi, Péter; Szabó, Éva; Szénási, Gábor; Papp, Lilla; Hársing, László G; Lévay, György; Spedding, Michael; Lambert, Jeremy J; Belelli, Delia; Barkóczy, József; Volk, Balázs; Simig, Gyula; Gacsályi, István; Antoni, Ferenc A

2015-10-05

Novel 2,3-benzodiazepine and related isoquinoline derivatives, substituted at position 1 with a 2-benzothiophenyl moiety, were synthesized to produce compounds that potently inhibited the action of GABA on heterologously expressed GABAA receptors containing the alpha 5 subunit (GABAA α5), with no apparent affinity for the benzodiazepine site. Substitutions of the benzothiophene moiety at position 4 led to compounds with drug-like properties that were putative inhibitors of extra-synaptic GABAA α5 receptors and had substantial blood-brain barrier permeability. Initial characterization in vivo showed that 8-methyl-5-[4-(trifluoromethyl)-1-benzothiophen-2-yl]-1,9-dihydro-2H-[1,3]oxazolo[4,5-h][2,3]benzodiazepin-2-one was devoid of sedative, pro-convulsive or motor side-effects, and enhanced the performance of rats in the object recognition test. In summary, we have discovered a first-in-class GABA-site inhibitor of extra-synaptic GABAA α5 receptors that has promising drug-like properties and warrants further development. Copyright © 2015 Elsevier B.V. All rights reserved.

Increased GABA(A) inhibition of the RVLM after hindlimb unloading in rats

NASA Technical Reports Server (NTRS)

Moffitt, Julia A.; Heesch, Cheryl M.; Hasser, Eileen M.

2002-01-01

Attenuated baroreflex-mediated increases in renal sympathetic nerve activity (RSNA) in hindlimb unloaded (HU) rats apparently are due to changes within the central nervous system. We hypothesized that GABA(A) receptor-mediated inhibition of the rostral ventrolateral medulla (RVLM) is increased after hindlimb unloading. Responses to bilateral microinjection of the GABA(A) antagonist (-)-bicuculline methiodide (BIC) into the RVLM were examined before and during caudal ventrolateral medulla (CVLM) inhibition in Inactin-anesthetized control and HU rats. Increases in mean arterial pressure (MAP), heart rate (HR), and RSNA in response to BIC in the RVLM were significantly enhanced in HU rats. Responses to bilateral CVLM blockade were not different. When remaining GABA(A) inhibition in the RVLM was blocked by BIC during CVLM inhibition, the additional increases in MAP and RSNA were significantly greater in HU rats. These data indicate that GABA(A) receptor-mediated inhibition of RVLM neurons is augmented after hindlimb unloading. Effects of input from the CVLM were unaltered. Thus, after cardiovascular deconditioning in rodents, the attenuated increase in sympathetic nerve activity in response to hypotension is associated with greater GABA(A) receptor-mediated inhibition of RVLM neurons originating at least in part from sources other than the CVLM.

Gi-Coupled γ-Aminobutyric Acid–B Receptors Cross-Regulate Phospholipase C and Calcium in Airway Smooth Muscle

PubMed Central

Mizuta, Kentaro; Mizuta, Fumiko; Xu, Dingbang; Masaki, Eiji; Panettieri, Reynold A.

2011-01-01

γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and exerts its actions via both ionotropic (GABAA) and metabotropic (GABAB) receptors. Although the functional expression of GABAB receptors coupled to the Gi protein was reported for airway smooth muscle, the role of GABAB receptors in airway responsiveness remains unclear. We investigated whether Gi-coupled GABAB receptors cross-regulate phospholipase C (PLC), an enzyme classically regulated by Gq-coupled receptors in human airway smooth muscle cells. Both the GABAB-selective agonist baclofen and the endogenous ligand GABA significantly increased the synthesis of inositol phosphate, whereas GABAA receptor agonists, muscimol, and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol exerted no effect. The baclofen-induced synthesis of inositol phosphate and transient increases in [Ca2+]i were blocked by CGP35348 and CGP55845 (selective GABAB antagonists), pertussis toxin (PTX, which inactivates the Gi protein), gallein (a Gβγ signaling inhibitor), U73122 (an inhibitor of PLC-β), and xestospongin C, an inositol 1,4,5-triphosphate receptor blocker. Baclofen also potentiated the bradykinin-induced synthesis of inositol phosphate and transient increases in [Ca2+]i, which were blocked by CGP35348 or PTX. Moreover, baclofen potentiated the substance P–induced contraction of airway smooth muscle in isolated guinea pig tracheal rings. In conclusion, the stimulation of GABAB receptors in human airway smooth muscle cells rapidly mobilizes intracellular Ca2+ stores by the synthesis of inositol phosphate via the activation of PLC-β, which is stimulated by Gβγ protein liberated from Gi proteins coupled to GABAB receptors. Furthermore, crosstalk between GABAB receptors and Gq-coupled receptors potentiates the synthesis of inositol phosphate, transient increases in [Ca2+]i, and smooth muscle contraction through Gi proteins. PMID:21719794

Analysis of Neural Systems Involved in Modulation of Memory Storage

DTIC Science & Technology

1993-02-01

doses of the muscarinic cholinergic agonist oxotremorine (Castellano and McGaugh, 1991). In experiments (unpublished) using intra- amygdala injections...and McGaugh, J.L. Oxotremorine attenuates retrograde amnesia induced by posttraining administration of the, GABAergic agonists muscimol and baclofen

A molecular characterization of the agonist binding site of a nematode cys-loop GABA receptor

PubMed Central

Kaji, Mark D; Kwaka, Ariel; Callanan, Micah K; Nusrat, Humza; Desaulniers, Jean-Paul; Forrester, Sean G

2015-01-01

Background and Purpose Cys-loop GABA receptors represent important targets for human chemotherapeutics and insecticides and are potential targets for novel anthelmintics (nematicides). However, compared with insect and mammalian receptors, little is known regarding the pharmacological characteristics of nematode Cys-loop GABA receptors. Here we have investigated the agonist binding site of the Cys-loop GABA receptor UNC-49 (Hco-UNC-49) from the parasitic nematode Haemonchus contortus. Experimental Approach We used two-electrode voltage-clamp electrophysiology to measure channel activation by classical GABA receptor agonists on Hco-UNC-49 expressed in Xenopus laevis oocytes, along with site-directed mutagenesis and in silico homology modelling. Key Results The sulphonated molecules P4S and taurine had no effect on Hco-UNC-49. Other classical Cys-loop GABAA receptor agonists tested on the Hco-UNC-49B/C heteromeric channel had a rank order efficacy of GABA > trans-4-aminocrotonic acid > isoguvacine > imidazole-4-acetic acid (IMA) > (R)-(−)-4-amino-3-hydroxybutyric acid [R(−)-GABOB] > (S)-(+)-4-amino-3-hydroxybutyric acid [S(+)-GABOB] > guanidinoacetic acid > isonipecotic acid > 5-aminovaleric acid (DAVA) (partial agonist) > β-alanine (partial agonist). In silico ligand docking revealed some variation in binding between agonists. Mutagenesis of a key serine residue in binding loop C to threonine had minimal effects on GABA and IMA but significantly increased the maximal response to DAVA and decreased twofold the EC50 for R(−)- and S(+)-GABOB. Conclusions and Implications The pharmacological profile of Hco-UNC-49 differed from that of vertebrate Cys-loop GABA receptors and insect resistance to dieldrin receptors, suggesting differences in the agonist binding pocket. These findings could be exploited to develop new drugs that specifically target GABA receptors of parasitic nematodes. PMID:25850584

Pharmacology of Ramelteon, a Selective MT1/MT2 Receptor Agonist: A Novel Therapeutic Drug for Sleep Disorders

PubMed Central

Miyamoto, Masaomi

2009-01-01

An estimated one-third of the general population is affected by insomnia, and this number is increasing due to more stressful working conditions and the progressive aging of society. However, current treatment of insomnia with hypnotics, gamma-aminobutyric acid A (GABAA) receptor modulators, induces various side effects, including cognitive impairment, motor disturbance, dependence, tolerance, hangover, and rebound insomnia. Ramelteon (Rozerem; Takeda Pharmaceutical Company Limited, Osaka, Japan) is an orally active, highly selective melatonin MT1/MT2 receptor agonist. Unlike the sedative hypnotics that target GABAA receptor complexes, ramelteon is a chronohypnotic that acts on the melatonin MT1 and MT2 receptors, which are primarily located in the suprachiasmatic nucleus, the body's “master clock.” As such, ramelteon possesses the first new therapeutic mechanism of action for a prescription insomnia medication in over three decades. Ramelteon has demonstrated sleep-promoting effects in clinical trials, and coupled with its favorable safety profile and lack of abuse potential or dependence, this chronohypnotic provides an important treatment option for insomnia. PMID:19228178

Benzodiazepine-site pharmacology on GABAA receptors in histaminergic neurons.

PubMed

May, A C; Fleischer, W; Kletke, O; Haas, H L; Sergeeva, O A

2013-09-01

The histaminergic tuberomamillary nucleus (TMN) of the posterior hypothalamus controls the cognitive aspects of vigilance which is reduced by common sedatives and anxiolytics. The receptors targeted by these drugs in histaminergic neurons are unknown. TMN neurons express nine different subunits of the GABAA receptor (GABAA R) with three α- (α1, α2 and α5) and two γ- (γ1, γ 2) subunits, which confer different pharmacologies of the benzodiazepine-binding site. We investigated the actions of zolpidem, midazolam, diazepam, chlordiazepoxide, flumazenil (Ro15-1788) and methyl-6,7-dimethoxy-4-ethyl-β-carboline-3-carboxylate (DMCM) in TMN neurons using mouse genetics, electrophysiological and molecular biological methods. We find the sensitivity of GABAA R to zolpidem, midazolam and DMCM significantly reduced in TMN neurons from γ2F77I mice, but modulatory activities of diazepam, chlordiazepoxide and flumazenil not affected. Potencies and efficacies of these compounds are in line with the dominance of α2- and α1-subunit containing receptors associated with γ2- or γ1-subunits. Functional expression of the γ1-subunit is supported by siRNA-based knock-down experiments in γ2F77I mice. GABAA R of TMN neurons respond to a variety of common sedatives with a high affinity binding site (γ2F77I) involved. The γ1-subunit likely contributes to the action of common sedatives in TMN neurons. This study is relevant for understanding the role of neuronal histamine and benzodiazepines in disorders of sleep and metabolism. © 2013 The British Pharmacological Society.

GABAergic regulation of REM sleep in reticularis pontis oralis and caudalis in rats.

PubMed

Sanford, Larry D; Tang, Xiangdong; Xiao, Jihua; Ross, Richard J; Morrison, Adrian R

2003-08-01

The nucleus reticularis pontis oralis (RPO) and nucleus reticularis pontis caudalis (RPC) are implicated in the generation of rapid eye movement sleep (REM). Work in cats has indicated that GABA in RPO plays a role in the regulation of REM. We assessed REM after local microinjections into RPO and RPC of the gamma-aminobutyric acid-A (GABA(A)) agonist, muscimol (MUS), and the GABA(A) antagonist, bicuculline (BIC). Rats (90-day-old male Sprague-Dawley) were implanted with electrodes for recording electroencephalographs (EEG) and electromyographs (EMG). Guide cannulae were aimed into RPO (n = 9) and RPC (n = 8) for microinjecting MUS (200, 1,000.0 microM) and BIC (0.056, 0.333, 1.0, 1,000.0, and 10,000.0 microM). Animals received bilateral microinjections of saline, MUS, and BIC (0.2 microl microinjected at 0.1 microl/min) into each region followed by 6-h sleep recordings. In RPO, MUS (1,000.0 microM) suppressed REM and BIC (1,000.0 microM) enhanced REM. In RPC, MUS (200, 1,000.0 microM) suppressed REM, but BIC (1,000.0 microM and less) did not significantly affect REM. Higher concentrations of BIC (10,000.0 microM) injected into RPO (n = 9) and RPC (n = 4) produced wakefulness and escape behavior. The results indicate that GABA in RPO/RPC is involved in the regulation of REM and suggest site-specific differences in this regulation.

Targeting brain-derived neurotrophic factor in the medial thalamus for the treatment of central poststroke pain in a rodent model.

PubMed

Shih, Hsi-Chien; Kuan, Yung-Hui; Shyu, Bai-Chung

2017-07-01

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.

Using drug combinations to assess potential contributions of non-GABAA receptors in the discriminative stimulus effects of the neuroactive steroid pregnanolone in rats.

PubMed

Eppolito, Amy K; Kodeih, Hanna R; Gerak, Lisa R

2014-10-01

Neuroactive steroids are increasingly implicated in the development of depression and anxiety and have been suggested as possible treatments for these disorders. While neuroactive steroids, such as pregnanolone, act primarily at γ-aminobutyric acidA (GABAA) receptors, other mechanisms might contribute to their behavioral effects and could increase their clinical effectiveness, as compared with drugs acting exclusively at GABAA receptors (e.g., benzodiazepines). The current study examined the role of non-GABAA receptors, including N-methyl-d-aspartate (NMDA) and serotonin3 (5-HT3) receptors, in the discriminative stimulus effects of pregnanolone. Separate groups of rats discriminated either 3.2mg/kg pregnanolone from vehicle or 0.32mg/kg of the benzodiazepine midazolam from vehicle while responding under a fixed-ratio 10 schedule for food pellets. When administered alone in both groups, pregnanolone and midazolam produced ≥80% drug-lever responding, the NMDA receptor antagonists dizocilpine and phencyclidine produced ≥60 and ≥30% drug-lever responding, respectively, and the 5-HT3 receptor agonist 1-(m-chlorophenyl)-biguanide (CPBG) and morphine produced <20% drug-lever responding up to doses that markedly decreased response rates. When studied together, neither dizocilpine, phencyclidine, CPBG nor morphine significantly altered the midazolam dose-effect curve in either group. Given that CPBG is without effect, it is unlikely that 5-HT3 receptors contribute substantially to the discriminative stimulus effects of pregnanolone. Similarities across groups in effects of dizocilpine and phencyclidine suggest that NMDA receptors do not differentially contribute to the effects of pregnanolone. Thus, NMDA and 5-HT3 receptors are not involved in the discriminative stimulus effects of pregnanolone. Copyright © 2014 Elsevier Inc. All rights reserved.

Cell type specificity of GABA(A) receptor mediated signaling in the hippocampus.

PubMed

Semyanov, A

2003-08-01

Inhibitory signaling mediated by ionotropic GABA(1) receptors generally acts as a major brake against excessive excitability in the brain. This is especially relevant in epilepsy-prone structures such as the hippocampus, in which GABA(A) receptor mediated inhibition is critical in suppressing epileptiform activity. Indeed, potentiating GABA(A) receptor mediated signaling is an important target for antiepileptic drug therapy. GABA(A) receptor mediated inhibition has different roles in the network dependent on the target neuron. Inhibiting principal cells will thus reduce network excitability, whilst inhibiting interneurons will increase network excitability; GABAergic therapeutic agents do not distinguish between these two alternatives, which may explain why, on occasion, GABAergic antiepileptic drugs can be proconvulsant. The importance of the target-cell for the effect of neuroactive drugs has emerged from a number of recent studies. Immunocytochemical data have suggested non-uniform distribution of GABA(A) receptor subunits among hippocampal interneurons and pyramidal cells. This has been confirmed by subsequent electropharmacological data. These have demonstrated that compounds which act on GABA(A) receptors or the extracellular GABA concentration can have distinct effects in different neuronal populations. Recently, it has also been discovered that presynaptic glutamate heteroreceptors can modulate GABA release in the hippocampus in a postsynaptic cell-specific manner. Since systemically administrated drugs may act on different neuronal subtypes, they can exhibit paradoxical effects. Distinguishing compounds that have target specific effects on GABAergic signaling may lead to novel and more effective treatments against epilepsy.

Zolpidem modulation of phasic and tonic GABA currents in the rat dorsal motor nucleus of the vagus

PubMed Central

Gao, Hong; Smith, Bret N.

2010-01-01

Zolpidem is a widely prescribed sleep aid with relative selectivity for GABAA receptors containing α1–3 subunits. We examined the effects of zolpidem on the inhibitory currents mediated by GABAA receptors using whole-cell patch-clamp recordings from DMV neurons in transverse brainstem slices from rat. Zolpidem prolonged the decay time of mIPSCs and of muscimol-evoked whole-cell GABAergic currents, and it occasionally enhanced the amplitude of mIPSCs. The effects were blocked by flumazenil, a benzodiazepine antagonist. Zolpidem also hyperpolarized the resting membrane potential, with a concomitant decrease in input resistance and action potential firing activity in a subset of cells. Zolpidem did not clearly alter the GABAA receptor-mediated tonic current (Itonic) under baseline conditions, but after elevating extracellular GABA concentration with nipecotic acid, a non-selective GABA transporter blocker, zolpidem consistently and significantly increased the tonic GABA current. This increase was suppressed by flumazenil and gabazine. These results suggest that α1–3 subunits are expressed in synaptic GABAA receptors on DMV neurons. The baseline tonic GABA current is likely not mediated by these same low affinity, zolpidem-sensitive GABAA receptors. However, when the extracellular GABA concentration is increased, zolpidem-sensitive extrasynaptic GABAA receptors containing α1–3 subunits contribute to the Itonic. PMID:20226798

Topographic and functional neuroanatomical study of GABAergic disinhibitory striatum-nigral inputs and inhibitory nigrocollicular pathways: neural hodology recruiting the substantia nigra, pars reticulata, for the modulation of the neural activity in the inferior colliculus involved with panic-like emotions.

PubMed

Castellan-Baldan, Lissandra; da Costa Kawasaki, Mateus; Ribeiro, Sandro José; Calvo, Fabrício; Corrêa, Vani Maria Alves; Coimbra, Norberto Cysne

2006-08-01

Considering the influence of the substantia nigra on mesencephalic neurons involved with fear-induced reactions organized in rostral aspects of the dorsal midbrain, the present work investigated the topographical and functional neuroanatomy of similar influence on caudal division of the corpora quadrigemina, addressing: (a) the neural hodology connecting the neostriatum, the substantia nigra, periaqueductal gray matter and inferior colliculus (IC) neural networks; (b) the influence of the inhibitory neostriatonigral-nigrocollicular GABAergic links on the control of the defensive behavior organized in the IC. The effects of the increase or decrease of activity of nigrocollicular inputs on defensive responses elicited by either electrical or chemical stimulation of the IC were also determined. Electrolytic or chemical lesions of the substantia nigra, pars reticulata (SNpr), decreased the freezing and escape behaviors thresholds elicited by electrical stimulation of the IC, and increased the behavioral responses evoked by the GABAA blockade in the same sites of the mesencephalic tectum (MT) electrically stimulated. These findings were corroborated by similar effects caused by microinjections of the GABAA-receptor agonist muscimol in the SNpr, followed by electrical and chemical stimulations of the IC. The GABAA blockade in the SNpr caused a significant increase in the defensive behavior thresholds elicited by electrical stimulation of the IC and a decrease in the mean incidence of panic-like responses induced by microinjections of bicuculline in the mesencephalic tectum (inferior colliculus). These findings suggest that the substantia nigra receives GABAergic inputs that modulate local and also inhibitory GABAergic outputs toward the IC. In fact, neurotracing experiments with fast blue and iontophoretic microinjections of biotinylated dextran amine either into the inferior colliculus or in the reticular division of the substantia nigra demonstrated a neural link

Metabolic products of linalool and modulation of GABAA receptors

NASA Astrophysics Data System (ADS)

Milanos, Sinem; Elsharif, Shaimaa A.; Janzen, Dieter; Buettner, Andrea; Villmann, Carmen

2017-06-01

Terpenoids are major subcomponents in aroma substances which harbor sedative physiological potential. We have demonstrated that various monoterpenoids such as the acyclic linalool enhance GABAergic currents in an allosteric manner in vitro upon overexpression of inhibitory a1b2 GABAA receptors in various expression systems. However, in plants or humans, i.e. following intake via inhalation or ingestion, linalool undergoes metabolic modifications including oxygenation and acetylation, which may affect the modulatory efficacy of the generated linalool derivatives. Here, we analyzed the modulatory potential of linalool derivatives at a1b2g2 GABAA receptors upon transient overexpression. Following receptor expression control, electrophysiological recordings in a whole cell configuration were used to determine the chloride influx upon co-application of GABA EC5-10 together with the modulatory substance. Our results show that only oxygenated linalool metabolites at carbon 8 positively affect GABAergic currents whereas derivatives hydroxylated or carboxylated at carbon 8 were rather ineffective. Acetylated linalool derivatives resulted in non-significant changes of GABAergic currents. We can conclude that metabolism of linalool reduces its positive allosteric potential at GABAA receptors compared to the significant potentiation effects of the parent molecule linalool itself.

Role played by periaqueductal gray neurons in parasympathetically mediated fear bradycardia in conscious rats.

PubMed

Koba, Satoshi; Inoue, Ryo; Watanabe, Tatsuo

2016-06-01

Freezing, a characteristic pattern of defensive behavior elicited by fear, is associated with a decrease in the heart rate. Central mechanisms underlying fear bradycardia are poorly understood. The periaqueductal gray (PAG) in the midbrain is known to contribute to autonomic cardiovascular adjustments associated with various emotional behaviors observed during active or passive defense reactions. The purpose of this study was to elucidate the role played by PAG neurons in eliciting fear bradycardia. White noise sound (WNS) exposure at 90 dB induced freezing behavior and elicited bradycardia in conscious rats. The WNS exposure-elicited bradycardia was mediated parasympathetically because intravenous administration of atropine abolished the bradycardia (P < 0.05). Moreover, WNS exposure-elicited bradycardia was mediated by neuronal activation of the lateral/ventrolateral PAG (l/vlPAG) because bilateral microinjection of muscimol, a GABAA agonist, into the l/vlPAG significantly suppressed the bradycardia. It is noted that muscimol microinjected bilaterally into the dorsolateral PAG had no effect on WNS exposure-elicited bradycardia. Furthermore, retrograde neuronal tracing experiments combined with immunohistochemistry demonstrated that a number of l/vlPAG neurons that send direct projections to the nucleus ambiguus (NA) in the medulla, a major origin of parasympathetic preganglionic neurons to the heart, were activated by WNS exposure. Based on these findings, we propose that the l/vlPAG-NA monosynaptic pathway transmits fear-driven central signals, which elicit bradycardia through parasympathetic outflow. © 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Nucleus accumbens GABAergic inhibition generates intense eating and fear that resists environmental retuning and needs no dopamine

PubMed Central

Richard, Jocelyn M.; Plawecki, Andrea M.; Berridge, Kent C.

2013-01-01

Intense fearful behavior and/or intense appetitive eating behavior can be generated by localized amino acid inhibitions along a rostrocaudal anatomical gradient within medial shell of nucleus accumbens of the rat. This can be produced by microinjections in medial shell of either the GABAA agonist muscimol (mimicking intrinsic GABAergic inputs) or the AMPA antagonist DNQX (disrupting corticolimbic glutamate inputs). At rostral sites in medial shell, each drug robustly stimulates appetitive eating and food intake, whereas at more caudal sites the same drugs instead produce increasingly fearful behaviors such as escape, distress vocalizations, and defensive treading (an antipredator behavior rodents emit to snakes and scorpions). Previously we showed that intense motivated behaviors generated by glutamate blockade require local endogenous dopamine and can be modulated in valence by environmental ambience. Here we investigated whether GABAergic generation of intense appetitive and fearful motivations similarly depends on local dopamine signals, and whether the valence of motivations generated by GABAergic inhibition can also be retuned by changes in environmental ambience. We report that the answer to both questions is ‘no’. Eating and fear generated by GABAergic inhibition of accumbens shell does not need endogenous dopamine. Also, the appetitive/fearful valence generated by GABAergic muscimol microinjections resists environmental retuning and is determined almost purely by rostrocaudal anatomical placement. These results suggest that NAc GABAergic release of fear and eating are relatively independent of modulatory dopamine signals, and more anatomically pre-determined in valence balance than release of the same intense behaviors by glutamate disruptions. PMID:23551138

Calcium/calmodulin-dependent kinase II phosphorylation of the GABAA receptor alpha1 subunit modulates benzodiazepine binding.

PubMed

Churn, Severn B; Rana, Aniruddha; Lee, Kangmin; Parsons, J Travis; De Blas, Angel; Delorenzo, Robert J

2002-09-01

gamma-Aminobutyric acid (GABA) is the primary neurotransmitter that is responsible for the fast inhibitory synaptic transmission in the central nervous system. A major post-translational mechanism that can rapidly regulate GABAAR function is receptor phosphorylation. This study was designed to test the effect of endogenous calcium and calmodulin-dependent kinase II (CaM kinase II) activation on both allosteric modulator binding and GABAA receptor subunit phosphorylation. Endogenous CaM kinase II activity was stimulated, and GABAA receptors were subsequently analyzed for bothallosteric modulator binding properties and immunoprecipitated and analyzed for subunit phosphorylation levels. A significant increase in allosteric-modulator binding of the GABAAR was observed under conditions maximal for CaM kinase II activation. In addition, CaM kinase II activation resulted in a direct increase in phosphorylation of the GABAA receptor alpha1 subunit. The data suggest that the CaM kinase II-dependent phosphorylation of the GABAA receptor alpha1 subunit modulated allosteric modulator binding to the GABAA receptor.

Receptor Subtype Alterations: Bases of Neuronal Plasticity and Learning

DTIC Science & Technology

1990-12-18

oxotremorine -M binding in rabbit anterior thalamus and cingulate cortex increased during the course of discriminative avoidance conditioning (DAC). Since there...anterior thalamus between training-induced neuronal plasticities and changes in oxotremorine -M binding. 3) The concentrations of noradrenaline, serotonin...binding protocols included the following: M 1, 3H-pirenzepine; M 2, 3H- oxotremorine -M in the p resence of unlabeled pirenzepine; GABAA, 3H-muscimol; M, and

Association between alcoholism and the genetic polymorphisms of the GABAA receptor genes on chromosome 5q33-34 in Korean population.

PubMed

Park, Chul-Soo; Park, So-Young; Lee, Chul-Soon; Sohn, Jin-Wook; Hahn, Gyu-Hee; Kim, Bong-Jo

2006-06-01

Family, twin, and adoption studies have demonstrated that genes play an important role in the development of alcoholism. We investigated the association between alcoholism and the genetic polymorphisms of the GABAA receptor genes on chromosome 5q33-34 in Korean population. The genotype of the GABAA receptor gene polymorphisms were determined by performing polymerase chain reaction genotyping for 172 normal controls and 162 male alcoholics who are hospitalized in alcoholism treatment institute. We found a significant association between the genetic polymorphisms of the GABAA alpha1 and GABAA alpha6 receptor gene and alcoholism. The GG genotype of the GABAA alpha1 receptor gene was associated with the onset age of alcoholism and alcohol withdrawal symptoms, and a high score on the Korean version of the ADS. However, there was no association between the genetic polymorphisms of the GABAA beta2 and gamma2 receptor gene and alcoholisms. Our finding suggest that genetic polymorphisms of the GABAA alpha1 and GABAA alpha6 receptor gene may be associated with the development of alcoholism and that the GG genotype of the GABAA alpha1 receptor gene play an important role in the development of the early onset and the severe type of alcoholism.

Modulation of GABAA receptors by valerian extracts is related to the content of valerenic acid.

PubMed

Trauner, Gabriele; Khom, Sophia; Baburin, Igor; Benedek, Birgit; Hering, Steffen; Kopp, Brigitte

2008-01-01

Valeriana Officinalis L . is a traditionally used sleep remedy, however, the mechanism of action and the substances responsible for its sedative and sleep-enhancing properties are not fully understood. As we previously identified valerenic acid as a subunit-specific allosteric modulator of GABAA receptors, we now investigated the relation between modulation of GABAA receptors by Valerian extracts of different polarity and the content of sesquiterpenic acids (valerenic acid, acetoxyvalerenic acid). All extracts were analysed by HPLC concerning the content of sesquiterpenic acids. GABAA receptors composed of alpha 1, beta 2 and gamma 2S subunits were expressed in Xenopus laevis oocytes and the modulation of chloride currents through GABAA receptors (IGABA) by Valerian extracts was investigated using the two-microelectrode voltage clamp technique. Apolar extracts induced a significant enhancement of IGABA, whereas polar extracts showed no effect. These results were confirmed by fractionating a highly active ethyl acetate extract: again fractions with high contents of valerenic acid exhibited strong receptor activation. In addition, removal of sesquiterpenic acids from the ethyl acetate extract led to a loss of I (GABA) enhancement. In conclusion, our data show that the extent of GABAA receptor modulation by Valerian extracts is related to the content of valerenic acid.

Effects of Insecticidal Ketones Present in Mint Plants on GABAA Receptor from Mammalian Neurons

PubMed Central

Sánchez-Borzone, Mariela Eugenia; Marin, Leticia Delgado; García, Daniel Asmed

2017-01-01

Background: The genus Mentha, an important member of the Lamiaceae family, is represented by many species commonly known as mint. The insecticidal activity of Mentha oil and its main components has been tested and established against various insects/pests. Among these, the ketone monoterpenes that are most common in different Mentha species demonstrated insect toxicity, with pulegone being the most active, followed by carvone and menthone. Considering that the GABAA receptor (GABAA-R) is one of the main insecticide targets on neurons, and that pulegone would modulate the insect GABA system, it may be expected that the insecticidal properties of Mentha ketones are mediated by their interaction with this receptor. Objective: In order to discern the pharmacological actions of these products when used as insecticides on mammalian organisms, we evaluated the pharmacologic activity of ketones, commonly present in Mentha plants, on native GABAA-R from rats. Materials and Methods: Determination of ketones effects on allosterically enhanced benzodiazepine binding, using primary cultures of cortical neurons, which express functional receptors and MTT assay to evaluate their cell toxicity. Results: Our results seem to indicate that ketone components of Mentha, with proven repellent or insecticide activity, were able to behave as GABAA-R negative allosteric modulators in murine cells and consequently could exhibit convulsant activity in mammalians. Only pulegone at the highest assayed concentration (2 mM) showed a significant reduction in cell viability after exposure for 24 hr. Conclusion: The present results strongly suggest that the ketone components of Mentha are able to exhibit convulsant activity in mammalian organisms, but functional assays and in vivo experiments would be necessary to corroborate this proposed action. SUMMARY The pharmacological activity of insecticide ketones, commonly present in Mentha plants, was evaluated on native GABAA receptor from mammalian

Rapid compensatory changes in GABA receptor efficacy in rat vestibular neurones after unilateral labyrinthectomy

PubMed Central

Yamanaka, Toshiaki; Him, Aydin; Cameron, Susan A; Dutia, Mayank B

2000-01-01

The inhibitory effects of the GABAA agonist muscimol and the GABAB agonist baclofen on tonically active medial vestibular nucleus (MVN) neurones were recorded in slices of the rat dorsal brainstem in vitro, to determine whether any changes occurred in the functional efficacy of GABAergic inhibition in these cells during the initial rapid stage of ‘vestibular compensation’, the behavioural recovery that takes place after unilateral labyrinthectomy (UL). These experiments were carried out in preparations where the midline was cut, severing all commissural connections between the two vestibular nuclei. Slices of the MVN were prepared from normal animals and animals that had been unilaterally labyrinthectomised 4 h earlier. The mean in vitro discharge rate of MVN neurones in the rostral region of the ipsi-lesional nucleus after UL was significantly higher than that in control slices, confirming our earlier reports of an increase in intrinsic excitability of these cells in the early stage of vestibular compensation. The in vitro discharge rates of caudal ipsi-lesional MVN cells, and rostral and caudal contra-lesional MVN cells, were not different from controls. Muscimol and baclofen caused reversible, dose-related inhibition of the tonic discharge rate of MVN cells in control slices. In slices prepared from UL animals, MVN cells in the rostral region of the ipsi-lesional nucleus showed a marked downregulation of their response to both muscimol and baclofen, seen as a rightward shift and a decrease in slope of the dose-response relationships for the two agonists. In the contra-lesional nucleus, there was a small but significant upregulation of the responsiveness of both rostral and caudal MVN cells to baclofen, and a marked upregulation of the responsiveness of caudal MVN cells to muscimol. In slices from animals that had undergone bilateral labyrinthectomy 4 h earlier, the downregulation of the functional efficacy of GABA receptors in the rostral MVN cells did not

Activity of Protein Kinase C is Important for 3α,5α-THP’s Actions at Dopamine Type 1-like and/or GABAA receptors in the Ventral Tegmental Area for Lordosis of Rats

PubMed Central

Frye, Cheryl A.; Walf, Alicia A.

2008-01-01

In the ventral tegmental area, progestogens facilitate sexual receptivity of rodents via actions at dopamine type 1-like and/or γ-aminobutyric type A receptors and activation of downstream signal transduction molecules. In the present study, we investigated whether effects of progesterone’s metabolite, 3α,5α-THP, to enhance lordosis via actions at these receptors in the ventral tegmental area requires phospholipase C-dependent protein kinase C. The objective of this study was to test the hypothesis that: if progestogens’ actions through dopamine type 1-like and/or γ-aminobutyric type A receptors in the ventral tegmental area for lordosis require protein kinase C, then inhibiting protein kinase C in the ventral tegmental area should reduce 3α,5α-THP-facilitated lordosis and its enhancement by dopamine type 1-like or γ-aminobutyric type A receptor agonists. Ovariectomized, E2 (10 μg s.c. at hr 0)-primed rats were tested for their baseline lordosis responses and then received a series of three infusions to the ventral tegmental area: first, bisindolylmaleimide (75 nM/side) or vehicle; second, SKF38393 (100 ng/side), muscimol (100 ng/side), or vehicle; third, 3α,5α-THP (100, 200 ng) or vehicle. Rats were pre-tested for lordosis and motor behavior and then tested for lordosis after each infusion and 10 and 60 mins after the last infusion. Rats were tested for motor behavior following their last lordosis test. As has been previously demonstrated, 3α,5α-THP infusions to the ventral tegmental area increased lordosis and effects were further enhanced by infusions of SKF38393 and muscimol. Infusions of bisindolylmaleimide to the ventral tegmental area attenuated 3α,5α-THP-, SKF38393-, and/or muscimol-facilitated lordosis. Effects on lordosis were not solely due to changes in general motor behavior. Thus, 3α,5α-THP’s actions in the ventral tegmental area through membrane receptors may require activity of protein kinase C. PMID:18675324

Bud extracts from Tilia tomentosa Moench inhibit hippocampal neuronal firing through GABAA and benzodiazepine receptors activation.

PubMed

Allio, Arianna; Calorio, Chiara; Franchino, Claudio; Gavello, Daniela; Carbone, Emilio; Marcantoni, Andrea

2015-08-22

Tilia tomentosa Moench bud extracts (TTBEs) is used in traditional medicine for centuries as sedative compound. Different plants belonging to the Tilia genus have shown their efficacy in the treatment of anxiety but still little is known about the mechanism of action of their bud extracts. To evaluate the action of TTBEs as anxiolytic and sedative compound on in vitro hippocampal neurons. The anxiolytic effect of TTBEs was assayed by testing the effects of these compounds on GABAA receptor-activated chloride current of hippocampal neurons by means of the patch-clamp technique and microelectrode-arrays (MEAs). TTBEs acutely administered on mouse hippocampal neurons, activated a chloride current comparable to that measured in the presence of GABA (100 µM). Bicuculline (100 µM) and picrotoxin (100 µM) blocked about 90% of this current, while the remaining 10% was blocked by adding the benzodiazepine (BDZ) antagonist flumazenil (30 µM). Flumazenil alone blocked nearly 60% of the TTBEs activated current, suggesting that TTBEs binds to both GABAA and BDZ receptor sites. Application of high-doses of TTBEs on spontaneous active hippocampal neurons grown for 3 weeks on MEAs blocked the synchronous activity of these neurons. The effects were mimicked by GABA and prevented by picrotoxin (100µM) and flumazenil (30 µM). At minimal doses, TTBEs reduced the frequency of synchronized bursts and increased the cross-correlation index of synchronized neuronal firing. Our data suggest that TTBEs mimics GABA and BDZ agonists by targeting hippocampal GABAergic synapses and inhibiting network excitability by increasing the strength of inhibitory synaptic outputs. Our results contribute toward the validation of TTBEs as effective sedative and anxiolytic compound. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

GABAA receptor-expressing neurons promote consumption in Drosophila melanogaster.

PubMed

Cheung, Samantha K; Scott, Kristin

2017-01-01

Feeding decisions are highly plastic and bidirectionally regulated by neurons that either promote or inhibit feeding. In Drosophila melanogaster, recent studies have identified four GABAergic interneurons that act as critical brakes to prevent incessant feeding. These GABAergic neurons may inhibit target neurons that drive consumption. Here, we tested this hypothesis by examining GABA receptors and neurons that promote consumption. We find that Resistance to dieldrin (RDL), a GABAA type receptor, is required for proper control of ingestion. Knockdown of Rdl in a subset of neurons causes overconsumption of tastants. Acute activation of these neurons is sufficient to drive consumption of appetitive substances and non-appetitive substances and acute silencing of these neurons decreases consumption. Taken together, these studies identify GABAA receptor-expressing neurons that promote Drosophila ingestive behavior and provide insight into feeding regulation.

Role of brain allopregnanolone in the plasticity of γ-aminobutyric acid type A receptor in rat brain during pregnancy and after delivery

PubMed Central

Concas, A.; Mostallino, M. C.; Porcu, P.; Follesa, P.; Barbaccia, M. L.; Trabucchi, M.; Purdy, R. H.; Grisenti, P.; Biggio, G.

1998-01-01

The relation between changes in brain and plasma concentrations of neurosteroids and the function and structure of γ-aminobutyric acid type A (GABAA) receptors in the brain during pregnancy and after delivery was investigated in rats. In contrast with plasma, where all steroids increased in parallel, the kinetics of changes in the cerebrocortical concentrations of progesterone, allopregnanolone (AP), and allotetrahydrodeoxycorticosterone (THDOC) diverged during pregnancy. Progesterone was already maximally increased between days 10 and 15, whereas AP and allotetrahydrodeoxycorticosterone peaked around day 19. The stimulatory effect of muscimol on 36Cl− uptake by cerebrocortical membrane vesicles was decreased on days 15 and 19 of pregnancy and increased 2 days after delivery. Moreover, the expression in cerebral cortex and hippocampus of the mRNA encoding for γ2L GABAA receptor subunit decreased during pregnancy and had returned to control values 2 days after delivery. Also α1,α2, α3, α4, β1, β2, β3, and γ2S mRNAs were measured and failed to change during pregnancy. Subchronic administration of finasteride, a 5α-reductase inhibitor, to pregnant rats reduced the concentrations of AP more in brain than in plasma as well as prevented the decreases in both the stimulatory effect of muscimol on 36Cl− uptake and the decrease of γ2L mRNA observed during pregnancy. These results indicate that the plasticity of GABAA receptors during pregnancy and after delivery is functionally related to fluctuations in endogenous brain concentrations of AP whose rate of synthesis/metabolism appears to differ in the brain, compared with plasma, in pregnant rats. PMID:9789080

Cyclohexanol analogues are positive modulators of GABAA receptor currents and act as general anaesthetics in vivo

USDA-ARS?s Scientific Manuscript database

GABAA receptors meet all the pharmacological criteria required to be considered important general anaesthetic targets. In the following study, the modulatory effects of various commercially available and novel cyclohexanol were investigated on recombinant human '-aminobutyric acid (GABAA, a1ß2'2s) r...

Alcohol-induced tolerance and physical dependence in mice with ethanol insensitive α1 GABAA receptors

PubMed Central

Werner, David F.; Swihart, Andrew R.; Ferguson, Carolyn; Lariviere, William R.; Harrison, Neil L.; Homanics, Gregg E.

2009-01-01

Background Although many people consume alcohol (ethanol), it remains unknown why some become addicted. Elucidating the molecular mechanisms of tolerance and physical dependence (withdrawal) may provide insight into alcohol addiction. While the exact molecular mechanisms of ethanol action are unclear, γ-aminobutyric acid type A receptors (GABAA-Rs) have been extensively implicated in ethanol action. The α1 GABAA-R subunit is associated with tolerance and physical dependence, but its exact role remains unknown. In this report, we tested the hypothesis that α1-GABAA-Rs mediate in part these effects of ethanol. Methods Ethanol-induced behavioral responses related to tolerance and physical dependence were investigated in knockin mice that have ethanol-insensitive α1 GABAA-Rs and wildtype controls. Acute functional tolerance (AFT) was assessed using the stationary dowel and loss of righting reflex assays. Chronic tolerance was assessed on the loss of righting reflex, fixed speed rotarod, hypothermia, and radiant tail flick assays following ten consecutive days of ethanol exposure. Withdrawal-related hyperexcitability was assessed by handling-induced convulsions following 3 cycles of ethanol vapor exposure/withdrawal. Immunoblots were used to assess α1 protein levels. Results Compared to controls, knockin mice displayed decreased AFT and chronic tolerance to ethanol-induced motor ataxia, and also displayed heightened ethanol-withdrawal hyperexcitability. No differences between wildtype and knockin mice were seen in other ethanol-induced behavioral measures. Following chronic exposure to ethanol, control mice displayed reductions in α1 protein levels, but knockins did not. Conclusions We conclude that α1-GABAA-Rs play a role in tolerance to ethanol-induced motor ataxia and withdrawal-related hyperexcitability. However, other aspects of behavioral tolerance and physical dependence do not rely on α1-containing GABAA-Rs. PMID:19032579

α2-containing GABAA receptors expressed in hippocampal region CA3 control fast network oscillations

PubMed Central

Heistek, Tim S; Ruiperez-Alonso, Marta; Timmerman, A Jaap; Brussaard, Arjen B; Mansvelder, Huibert D

2013-01-01

GABAA receptors are critically involved in hippocampal oscillations. GABAA receptor α1 and α2 subunits are differentially expressed throughout the hippocampal circuitry and thereby may have distinct contributions to oscillations. It is unknown which GABAA receptor α subunit controls hippocampal oscillations and where these receptors are expressed. To address these questions we used transgenic mice expressing GABAA receptor α1 and/or α2 subunits with point mutations (H101R) that render these receptors insensitive to allosteric modulation at the benzodiazepine binding site, and tested how increased or decreased function of α subunits affects hippocampal oscillations. Positive allosteric modulation by zolpidem prolonged decay kinetics of hippocampal GABAergic synaptic transmission and reduced the frequency of cholinergically induced oscillations. Allosteric modulation of GABAergic receptors in CA3 altered oscillation frequency in CA1, while modulation of GABA receptors in CA1 did not affect oscillations. In mice having a point mutation (H101R) at the GABAA receptor α2 subunit, zolpidem effects on cholinergically induced oscillations were strongly reduced compared to wild-type animals, while zolpidem modulation was still present in mice with the H101R mutation at the α1 subunit. Furthermore, genetic knockout of α2 subunits strongly reduced oscillations, whereas knockout of α1 subunits had no effect. Allosteric modulation of GABAergic receptors was strongly reduced in unitary connections between fast spiking interneurons and pyramidal neurons in CA3 of α2H101R mice, but not of α1H101R mice, suggesting that fast spiking interneuron to pyramidal neuron synapses in CA3 contain α2 subunits. These findings suggest that α2-containing GABAA receptors expressed in the CA3 region provide the inhibition that controls hippocampal rhythm during cholinergically induced oscillations. PMID:23109109

Benzodiazepine-induced anxiolysis and reduction of conditioned fear are mediated by distinct GABAA receptor subtypes in mice

PubMed Central

Smith, Kiersten S.; Engin, Elif; Meloni, Edward G.; Rudolph, Uwe

2012-01-01

GABAA receptor modulating drugs such as benzodiazepines (BZs) have been used to treat anxiety disorders for over five decades. In order to determine whether the same or different GABAA receptor subtypes are necessary for the anxiolytic-like action of BZs in unconditioned anxiety and conditioned fear models, we investigated the role of different GABAA receptor subtypes by challenging wild type, α1(H101R), α2(H101R) and α3(H126R) mice bred on the C57BL/6J background with diazepam or chlordiazepoxide in the elevated plus maze and the fear-potentiated startle paradigms. Both drugs significantly increased open arm exploration in the elevated plus maze in wild type, α1(H101R) and α3(H126R), but this effect was abolished in α2(H101R) mice; these were expected results based on previous published results. In contrast, while administration of diazepam and chlordiazepoxide significantly attenuated fear-potentiated startle (FPS) in wild type mice and α3(H126R) mice, the fear-reducing effects of these drugs were absent in both α1(H101R) and α2(H101R) point mutants, indicating that both α1- and α2-containing GABAA receptors are necessary for BZs to exert their effects on conditioned fear responses.. Our findings illustrate both an overlap and a divergence between the GABAA receptor subtype requirements for the impact of BZs, specifically that both α1- and α2-containing GABAA receptors are necessary for BZs to reduce conditioned fear whereas only α2-containing GABAA receptors are needed for BZ-induced anxiolysis in unconditioned tests of anxiety. This raises the possibility that GABAergic pharmacological interventions for specific anxiety disorders can be differentially tailored. PMID:22465203

Modulation of neurosteroid potentiation by protein kinases at synaptic- and extrasynaptic-type GABAA receptors

PubMed Central

Adams, Joanna M.; Thomas, Philip; Smart, Trevor G.

2015-01-01

GABAA receptors are important for inhibition in the CNS where neurosteroids and protein kinases are potent endogenous modulators. Acting individually, these can either enhance or depress receptor function, dependent upon the type of neurosteroid or kinase and the receptor subunit combination. However, in vivo, these modulators probably act in concert to fine-tune GABAA receptor activity and thus inhibition, although how this is achieved remains unclear. Therefore, we investigated the relationship between these modulators at synaptic-type α1β3γ2L and extrasynaptic-type α4β3δ GABAA receptors using electrophysiology. For α1β3γ2L, potentiation of GABA responses by tetrahydro-deoxycorticosterone was reduced after inhibiting protein kinase C, and enhanced following its activation, suggesting this kinase regulates neurosteroid modulation. In comparison, neurosteroid potentiation was reduced at α1β3S408A,S409Aγ2L receptors, and unaltered by PKC inhibitors or activators, indicating that phosphorylation of β3 subunits is important for regulating neurosteroid activity. To determine whether extrasynaptic-type GABAA receptors were similarly modulated, α4β3δ and α4β3S408A,S409Aδ receptors were investigated. Neurosteroid potentiation was reduced at both receptors by the kinase inhibitor staurosporine. By contrast, neurosteroid-mediated potentiation at α4S443Aβ3S408A,S409Aδ receptors was unaffected by protein kinase inhibition, strongly suggesting that phosphorylation of α4 and β3 subunits is required for regulating neurosteroid activity at extrasynaptic receptors. Western blot analyses revealed that neurosteroids increased phosphorylation of β3S408,S409 implying that a reciprocal pathway exists for neurosteroids to modulate phosphorylation of GABAA receptors. Overall, these findings provide important insight into the regulation of GABAA receptors in vivo, and into the mechanisms by which GABAergic inhibitory transmission may be simultaneously tuned by

Inhibitory input from slowly adapting lung stretch receptors to retrotrapezoid nucleus chemoreceptors

PubMed Central

Moreira, Thiago S; Takakura, Ana C; Colombari, Eduardo; West, Gavin H; Guyenet, Patrice G

2007-01-01

The retrotrapezoid nucleus (RTN) contains CO2-activated interneurons with properties consistent with central respiratory chemoreceptors. These neurons are glutamatergic and express the transcription factor Phox2b. Here we tested whether RTN neurons receive an input from slowly adapting pulmonary stretch receptors (SARs) in halothane-anaesthetized ventilated rats. In vagotomized rats, RTN neurons were inhibited to a variable extent by stimulating myelinated vagal afferents using the lowest intensity needed to inhibit the phrenic nerve discharge (PND). In rats with intact vagus nerves, RTN neurons were inhibited, also to a variable extent, by increasing positive end-expiratory pressure (PEEP; 2–6 cmH2O). The cells most sensitive to PEEP were inhibited during each lung inflation at rest and were instantly activated by stopping ventilation. Muscimol (GABA-A agonist) injection in or next to the solitary tract at area postrema level desynchronized PND from ventilation, eliminated the lung inflation-synchronous inhibition of RTN neurons and their steady inhibition by PEEP but did not change their CO2 sensitivity. Muscimol injection into the rostral ventral respiratory group eliminated PND but did not change RTN neuron response to either lung inflation, PEEP increases, vagal stimulation or CO2. Generalized glutamate receptor blockade with intracerebroventricular (i.c.v.) kynurenate eliminated PND and the response of RTN neurons to lung inflation but did not change their CO2 sensitivity. PEEP-sensitive RTN neurons expressed Phox2b. In conclusion, RTN chemoreceptors receive an inhibitory input from myelinated lung stretch receptors, presumably SARs. The lung input to RTN may be di-synaptic with inhibitory pump cells as sole interneurons. PMID:17255166

The role of dorsomedial hypotalamus ionotropic glutamate receptors in the hypertensive and tachycardic responses evoked by Tityustoxin intracerebroventricular injection.

PubMed

Silva, F C; Guidine, Patrícia Alves Maia; Machado, Natalia Lima; Xavier, Carlos Henrique; de Menezes, R C; Moraes-Santos, Tasso; Moraes, Márcio Flávio; Chianca, Deoclécio Alves

2015-03-01

The scorpion envenoming syndrome is an important worldwide public health problem due to its high incidence and potential severity of symptoms. Some studies address the high sensitivity of the central nervous system to this toxin action. It is known that cardiorespiratory manifestations involve the activation of the autonomic nervous system. However, the origin of this modulation remains unclear. Considering the important participation of the dorsomedial hypotalamus (DMH) in the cardiovascular responses during emergencial situations, the aim of this work is to investigate the involvement of the DMH on cardiovascular responses induced by intracerebroventricular (icv) injection of Tityustoxin (TsTX, a α-type toxin extracted from the Tityus serrulatus scorpion venom). Urethane-anaesthetized male Wistar rats (n=30) were treated with PBS, muscimol or ionotropic glutamate receptor antagonists, bilaterally in DMH and later, with an icv injection of TsTX, or treated only with PBS in both regions. TsTX evoked a marked increase in mean arterial pressure and heart rate in all control rats. Interestingly, injection of muscimol, a GABAA receptor agonist, did not change the pressor and tachycardic responses evoked by TsTX. Remarkably, the injection ionotropic glutamate receptors antagonists in DMH abolished the pressor and the tachycardic response evoked by TsTX. Our data suggest that the central circuit recruited by TsTX, whose activation results in an array of physiological and behavioral alterations, depend on the activation of DMH ionotropic glutamate receptors. Moreover, our data provide new insights on the central mechanisms involved in the development of symptoms in the severe scorpion envenomation syndrome. Copyright © 2014 Elsevier Inc. All rights reserved.

A Critical Role of Lateral Hypothalamus in Context-Induced Relapse to Alcohol Seeking after Punishment-Imposed Abstinence

PubMed Central

Rabei, Rana; Kaganovsky, Konstantin; Caprioli, Daniele; Bossert, Jennifer M.; Bonci, Antonello

2014-01-01

In human alcoholics, abstinence is often self-imposed, despite alcohol availability, because of the negative consequences of excessive use. During abstinence, relapse is often triggered by exposure to contexts associated with alcohol use. We recently developed a rat model that captures some features of this human condition: exposure to the alcohol self-administration environment (context A), after punishment-imposed suppression of alcohol self-administration in a different environment (context B), provoked renewal of alcohol seeking in alcohol-preferring P rats. The mechanisms underlying context-induced renewal of alcohol seeking after punishment-imposed abstinence are unknown. Here, we studied the role of the lateral hypothalamus (LH) and its forebrain projections in this effect. We first determined the effect of context-induced renewal of alcohol seeking on Fos (a neuronal activity marker) expression in LH. We next determined the effect of LH reversible inactivation by GABAA + GABAB receptor agonists (muscimol + baclofen) on this effect. Finally, we determined neuronal activation in brain areas projecting to LH during context-induced renewal tests by measuring double labeling of the retrograde tracer cholera toxin subunit B (CTb; injected in LH) with Fos. Context-induced renewal of alcohol seeking after punishment-imposed abstinence was associated with increased Fos expression in LH. Additionally, renewal was blocked by muscimol + baclofen injections into LH. Finally, double-labeling analysis of CTb + Fos showed that context-induced renewal of alcohol seeking after punishment-imposed abstinence was associated with selective activation of accumbens shell neurons projecting to LH. The results demonstrate an important role of LH in renewal of alcohol seeking after punishment-imposed abstinence and suggest a role of accumbens shell projections to LH in this form of relapse. PMID:24872550

Comparison of Steroid Modulation of Spontaneous Inhibitory Postsynaptic Currents in Cultured Hippocampal Neurons and Steady-State Single-Channel Currents from Heterologously Expressed α1β2γ2L GABAA Receptors

PubMed Central

Chakrabarti, Sampurna; Qian, Mingxing; Krishnan, Kathiresan; Covey, Douglas F.; Mennerick, Steven

2016-01-01

Neuroactive steroids are efficacious modulators of γ-aminobutyric acid type A receptor (GABAA) receptor function. The effects of steroids on the GABAA receptor are typically determined by comparing steady-state single-channel open probability or macroscopic peak responses elicited by GABA in the absence and presence of a steroid. Due to differences in activation conditions (exposure duration, concentration of agonist), it is not obvious whether modulation measured using typical experimental protocols can be used to accurately predict the effect of a modulator on native receptors under physiologic conditions. In the present study, we examined the effects of 14 neuroactive steroids and analogs on the properties of spontaneous inhibitory postsynaptic currents (sIPSCs) in cultured rat hippocampal neurons. The goal was to determine whether the magnitude of modulation of the decay time course of sIPSCs correlates with the extent of modulation and kinetic properties of potentiation as determined in previous single-channel studies. The steroids were selected to cover a wide range of efficacy on heterologously expressed rat α1β2γ2L GABAA receptors, ranging from essentially inert to highly efficacious (strong potentiators of single-channel and macroscopic peak responses). The data indicate a strong correlation between prolongation of the decay time course of sIPSCs and potentiation of single-channel open probability. Furthermore, changes in intracluster closed time distributions were the single best predictor of prolongation of sIPSCs. We infer that the information obtained in steady-state single-channel recordings can be used to forecast modulation of synaptic currents. PMID:26769414

Allyl m-Trifluoromethyldiazirine Mephobarbital: An Unusually Potent Enantioselective and Photoreactive Barbiturate General Anesthetic

DOE Office of Scientific and Technical Information (OSTI.GOV)

Savechenkov, Pavel Y.; Zhang, Xi; Chiara, David C.

2012-12-10

We synthesized 5-allyl-1-methyl-5-(m-trifluoromethyl-diazirynylphenyl)barbituric acid (14), a trifluoromethyldiazirine-containing derivative of general anesthetic mephobarbital, separated the racemic mixture into enantiomers by chiral chromatography, and determined the configuration of the (+)-enantiomer as S by X-ray crystallography. Additionally, we obtained the {sup 3}H-labeled ligand with high specific radioactivity. R-(-)-14 is an order of magnitude more potent than the most potent clinically used barbiturate, thiopental, and its general anesthetic EC{sub 50} approaches those for propofol and etomidate, whereas S-(+)-14 is 10-fold less potent. Furthermore, at concentrations close to its anesthetic potency, R-(-)-14 both potentiated GABA-induced currents and increased the affinity for the agonist muscimol inmore » human {alpha}1{beta}2/3{gamma}2L GABA{sub A} receptors. Finally, R-(-)-14 was found to be an exceptionally efficient photolabeling reagent, incorporating into both {alpha}1 and {beta}3 subunits of human {alpha}1{beta}3 GABAA receptors. These results indicate R-(-)-14 is a functional general anesthetic that is well-suited for identifying barbiturate binding sites on Cys-loop receptors.« less

Temporary inactivation of the anterior part of the bed nucleus of the stria terminalis blocks alarm pheromone-induced defensive behavior in rats

PubMed Central

Breitfeld, Tino; Bruning, Johann E. A.; Inagaki, Hideaki; Takeuchi, Yukari; Kiyokawa, Yasushi; Fendt, Markus

2015-01-01

Rats emit an alarm pheromone in threatening situations. Exposure of rats to this alarm pheromone induces defensive behaviors, such as head out behavior, and increases c-Fos expression in brain areas involved in the mediation of defensive behaviors. One of these brain areas is the anterior bed nucleus of the stria terminalis (aBNST). The goal of the present study was to investigate if pharmacological inactivation of the aBNST by local microinjections of the GABAA receptor-agonist muscimol modulates alarm pheromone-induced defensive behaviors. We first established the behavioral paradigm of alarm pheromone-induced defensive behaviors in Sprague-Dawley rats in our laboratory. In a second experiment, we inactivated the aBNST, then exposed rats to one of four different odors (neck odor, female urine, alarm pheromone, fox urine) and tested the effects of the aBNST inactivation on the behavior in response to these odors. Our data show that temporary inactivation of the aBNST blocked head out behavior in response to the alarm pheromone. This indicates that the aBNST plays an important role in the mediation of the alarm pheromone-induced defensive behavior in rats. PMID:26441496

The depolarizing action of GABA in cultured hippocampal neurons is not due to the absence of ketone bodies.

PubMed

Waddell, Jaylyn; Kim, Jimok; Alger, Bradley E; McCarthy, Margaret M

2011-01-01

Two recent reports propose that the depolarizing action of GABA in the immature brain is an artifact of in vitro preparations in which glucose is the only energy source. The authors argue that this does not mimic the physiological environment because the suckling rats use ketone bodies and pyruvate as major sources of metabolic energy. Here, we show that availability of physiologically relevant levels of ketone bodies has no impact on the excitatory action of GABA in immature cultured hippocampal neurons. Addition of β-hydroxybutyrate (BHB), the primary ketone body in the neonate rat, affected neither intracellular calcium elevation nor membrane depolarizations induced by the GABA-A receptor agonist muscimol, when assessed with calcium imaging or perforated patch-clamp recording, respectively. These results confirm that the addition of ketone bodies to the extracellular environment to mimic conditions in the neonatal brain does not reverse the chloride gradient and therefore render GABA hyperpolarizing. Our data are consistent with the existence of a genuine "developmental switch" mechanism in which GABA goes from having a predominantly excitatory role in immature cells to a predominantly inhibitory one in adults.

Dual Modulators of GABA-A and Alpha 7 Nicotinic Receptors for Treating Autism

DTIC Science & Technology

2015-10-01

AWARD NUMBER: W81XWH-13-1-0144 TITLE: Dual Modulators of GABA-A and Alpha 7 Nicotinic Receptors for Treating Autism PRINCIPAL INVESTIGATOR...SUBTITLE 5a. CONTRACT NUMBER Dual Modulators of GABA-A and Alpha 7 Nicotinic Receptors for Treating Autism 5b. GRANT NUMBER W81XWH-13-1-0144 5c...ABSTRACT Autism spectrum disorder (ASD) is a polygenic signaling disorder that may result, in part, from an imbalance in excitatory and inhibitory

A network of autism linked genes stabilizes two pools of synaptic GABAA receptors

PubMed Central

Tong, Xia-Jing; Hu, Zhitao; Liu, Yu; Anderson, Dorian; Kaplan, Joshua M

2015-01-01

Changing receptor abundance at synapses is an important mechanism for regulating synaptic strength. Synapses contain two pools of receptors, immobilized and diffusing receptors, both of which are confined to post-synaptic elements. Here we show that immobile and diffusing GABAA receptors are stabilized by distinct synaptic scaffolds at C. elegans neuromuscular junctions. Immobilized GABAA receptors are stabilized by binding to FRM-3/EPB4.1 and LIN-2A/CASK. Diffusing GABAA receptors are stabilized by the synaptic adhesion molecules Neurexin and Neuroligin. Inhibitory post-synaptic currents are eliminated in double mutants lacking both scaffolds. Neurexin, Neuroligin, and CASK mutations are all linked to Autism Spectrum Disorders (ASD). Our results suggest that these mutations may directly alter inhibitory transmission, which could contribute to the developmental and cognitive deficits observed in ASD. DOI: http://dx.doi.org/10.7554/eLife.09648.001 PMID:26575289

Gi-coupled γ-aminobutyric acid-B receptors cross-regulate phospholipase C and calcium in airway smooth muscle.

PubMed

Mizuta, Kentaro; Mizuta, Fumiko; Xu, Dingbang; Masaki, Eiji; Panettieri, Reynold A; Emala, Charles W

2011-12-01

γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and exerts its actions via both ionotropic (GABA(A)) and metabotropic (GABA(B)) receptors. Although the functional expression of GABA(B) receptors coupled to the G(i) protein was reported for airway smooth muscle, the role of GABA(B) receptors in airway responsiveness remains unclear. We investigated whether G(i)-coupled GABA(B) receptors cross-regulate phospholipase C (PLC), an enzyme classically regulated by G(q)-coupled receptors in human airway smooth muscle cells. Both the GABA(B)-selective agonist baclofen and the endogenous ligand GABA significantly increased the synthesis of inositol phosphate, whereas GABA(A) receptor agonists, muscimol, and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol exerted no effect. The baclofen-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i) were blocked by CGP35348 and CGP55845 (selective GABA(B) antagonists), pertussis toxin (PTX, which inactivates the G(i) protein), gallein (a G(βγ) signaling inhibitor), U73122 (an inhibitor of PLC-β), and xestospongin C, an inositol 1,4,5-triphosphate receptor blocker. Baclofen also potentiated the bradykinin-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i), which were blocked by CGP35348 or PTX. Moreover, baclofen potentiated the substance P-induced contraction of airway smooth muscle in isolated guinea pig tracheal rings. In conclusion, the stimulation of GABA(B) receptors in human airway smooth muscle cells rapidly mobilizes intracellular Ca(2+) stores by the synthesis of inositol phosphate via the activation of PLC-β, which is stimulated by G(βγ) protein liberated from G(i) proteins coupled to GABA(B) receptors. Furthermore, crosstalk between GABA(B) receptors and G(q)-coupled receptors potentiates the synthesis of inositol phosphate, transient increases in [Ca(2+)](i), and smooth muscle contraction through G

cAMP and forskolin decrease gamma-aminobutyric acid-gated chloride flux in rat brain synaptoneurosomes.

PubMed Central

Heuschneider, G; Schwartz, R D

1989-01-01

The effects of the cyclic nucleotide cAMP on gamma-aminobutyric acid-gated chloride channel function were investigated. The membrane-permeant cAMP analog N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate inhibited muscimol-induced 36Cl- uptake into rat cerebral cortical synaptoneurosomes in a concentration-dependent manner (IC50 = 1.3 mM). The inhibition was due to a decrease in the maximal effect of muscimol, with no change in potency. Similar effects were observed with 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate, 8-bromoadenosine 3',5'-cyclic monophosphate, and the phosphodiesterase inhibitor isobutylmethylxanthine. The effect of endogenous cAMP accumulation on the gamma-aminobutyric acid-gated Cl- channel was studied with forskolin, an activator of adenylate cyclase. Under identical conditions, in the intact synaptoneurosomes, forskolin inhibited muscimol-induced 36Cl- uptake and generated cAMP with similar potencies (IC50 = 14.3 microM; EC50 = 6.2 microM, respectively). Surprisingly, 1,9-dideoxyforskolin, which does not activate adenylate cyclase, also inhibited the muscimol response, suggesting that forskolin and its lipophilic derivatives may interact with the Cl- channel directly. Indeed, forskolin inhibition of muscimol-induced 36Cl- uptake was extremely rapid (within 5 sec), preceding the accumulation of sufficient levels of cAMP. After 5 min, a slower phase of inhibition was seen, similar to the time course for cAMP accumulation. The data suggest that gamma-aminobutyric acid (GABAA) receptor function in brain can be regulated by cAMP-dependent phosphorylation. PMID:2468163

Reversal of pathological pain through specific spinal GABAA receptor subtypes.

PubMed

Knabl, Julia; Witschi, Robert; Hösl, Katharina; Reinold, Heiko; Zeilhofer, Ulrike B; Ahmadi, Seifollah; Brockhaus, Johannes; Sergejeva, Marina; Hess, Andreas; Brune, Kay; Fritschy, Jean-Marc; Rudolph, Uwe; Möhler, Hanns; Zeilhofer, Hanns Ulrich

2008-01-17

Inflammatory diseases and neuropathic insults are frequently accompanied by severe and debilitating pain, which can become chronic and often unresponsive to conventional analgesic treatment. A loss of synaptic inhibition in the spinal dorsal horn is considered to contribute significantly to this pain pathology. Facilitation of spinal gamma-aminobutyric acid (GABA)ergic neurotransmission through modulation of GABA(A) receptors should be able to compensate for this loss. With the use of GABA(A)-receptor point-mutated knock-in mice in which specific GABA(A) receptor subtypes have been selectively rendered insensitive to benzodiazepine-site ligands, we show here that pronounced analgesia can be achieved by specifically targeting spinal GABA(A) receptors containing the alpha2 and/or alpha3 subunits. We show that their selective activation by the non-sedative ('alpha1-sparing') benzodiazepine-site ligand L-838,417 (ref. 13) is highly effective against inflammatory and neuropathic pain yet devoid of unwanted sedation, motor impairment and tolerance development. L-838,417 not only diminished the nociceptive input to the brain but also reduced the activity of brain areas related to the associative-emotional components of pain, as shown by functional magnetic resonance imaging in rats. These results provide a rational basis for the development of subtype-selective GABAergic drugs for the treatment of chronic pain, which is often refractory to classical analgesics.

Postsynaptic activity reverses the sign of the acetylcholine-induced long-term plasticity of GABAA inhibition

PubMed Central

Domínguez, Soledad; Fernández de Sevilla, David; Buño, Washington

2014-01-01

Acetylcholine (ACh) regulates forms of plasticity that control cognitive functions but the underlying mechanisms remain largely unknown. ACh controls the intrinsic excitability, as well as the synaptic excitation and inhibition of CA1 hippocampal pyramidal cells (PCs), cells known to participate in circuits involved in cognition and spatial navigation. However, how ACh regulates inhibition in function of postsynaptic activity has not been well studied. Here we show that in rat PCs, a brief pulse of ACh or a brief stimulation of cholinergic septal fibers combined with repeated depolarization induces strong long-term enhancement of GABAA inhibition (GABAA-LTP). Indeed, this enhanced inhibition is due to the increased activation of α5βγ2 subunit-containing GABAA receptors by the GABA released. GABAA-LTP requires the activation of M1-muscarinic receptors and an increase in cytosolic Ca2+. In the absence of PC depolarization ACh triggered a presynaptic depolarization-induced suppression of inhibition (DSI), revealing that postsynaptic activity gates the effects of ACh from presynaptic DSI to postsynaptic LTP. These results provide key insights into mechanisms potentially linked with cognitive functions, spatial navigation, and the homeostatic control of abnormal hyperexcitable states. PMID:24938789

GABAa and GABAc receptor-mediated modulation of responses to color stimuli: electroretinographic study in the turtle Emys orbicularis.

PubMed

Kupenova, Petia; Vitanova, Lily; Popova, Elka

2010-04-01

GABAergic transmission is involved in color coding in the retina. The specific contribution of different GABA receptors to spectral sensitivity of the retinal responses is not well characterized. We studied GABAa and GABAc receptor-mediated effects on the intensity-response functions of the electroretinographic ON (b-wave) and OFF (d-wave) responses to color stimuli. For this purpose, we compared the effects of GABAa receptor blockade by bicuculline with the effects of GABAa + GABAc receptor blockade by picrotoxin. The blockade of both GABAa and GABAc receptors caused an amplitude increase of the electroretinographic responses, but the effects of the two blockades depended in a specific manner on stimulus intensity and wavelength. The effects of GABAa receptor blockade showed distinct color ON/OFF asymmetry. The absolute and relative sensitivities of the ON responses to blue stimuli and OFF responses to red stimuli were increased to the greatest degree while the sensitivity of the ON responses to red stimuli and OFF responses to blue stimuli was least increased. In contrast, color ON/OFF asymmetry was not typical of the effects of GABAc receptor blockade. The most prominent GABAc effect was the sensitivity increase of the ON and OFF responses to blue stimuli and, to some lesser extent, to green stimuli. The results of this study indicate a specific role of GABAa and GABAc receptor-mediated influences in processing of chromatic information in the distal retina.

Frequency-dependent actions of benzodiazepines on GABAA receptors in cultured murine cerebellar granule cells.

PubMed Central

Mellor, J R; Randall, A D

1997-01-01

1. Miniature IPSCs recorded from cultured murine cerebellar granule cells increased in half-width and amplitude following application of the benzodiazepine (BDZ) Flunitrazepam (Flu, 1 microM). The increase in the half-width was much greater than that in the amplitude. 2. Five-millisecond applications of 1 mM GABA to nucleated outside-out patches elicited rapidly rising biexponentially decaying responses that resembled IPSCs. Flu had no effect on the amplitude of such responses, but consistently slowed their deactivation by approximately 50%. This effect was reversed by Flu washout or application of the BDZ antagonist Ro15-1788. The partial inverse agonist. Ro15-4513 speeded deactivation and depressed peak current amplitude by 23 +/- 12%. 3. The EC50 for GABA was between 45 and 50 microM. At submaximally effective agonist concentrations, Flu increased response amplitude and slowed response deactivation. Both effects were present in all cells taken from young cultures (4-7 days in vitro) but the latter was absent in 55% of the neurones obtained from older cultures (14-27 days in vitro). 4. With 120 ms applications of 20 microM GABA, responses activated monoexponentially (time constant, 39.8 +/- 2.8 ms) and deactivated biexponentially (time constants, 40.4 +/- 2.1 and 251 +/- 15 ms). Application of Flu slowed both activation and deactivation. The latter effect arose from an increased contribution of the slower component of decay. 5. Desensitization of responses to 1 mM GABA was biexponential, with time constants of 47 +/- 11 and 479 +/- 49 ms. Flu speeded desensitization by decreasing both fast and slow time constants. GABAA receptor desensitization consistently slowed subsequent deactivation. No significant relationship between the level of desensitization and the amount of slowing of deactivation produced by Flu was found. 6. Responses to paired 5 ms applications of 1 mM GABA indicated that the slowing of deactivation and the speeding of desensitization produced by

Lipid raft integrity affects GABAA receptor, but not NMDA receptor modulation by psychopharmacological compounds.

PubMed

Nothdurfter, Caroline; Tanasic, Sascha; Di Benedetto, Barbara; Uhr, Manfred; Wagner, Eva-Maria; Gilling, Kate E; Parsons, Chris G; Rein, Theo; Holsboer, Florian; Rupprecht, Rainer; Rammes, Gerhard

2013-07-01

Lipid rafts have been shown to play an important role for G-protein mediated signal transduction and the function of ligand-gated ion channels including their modulation by psychopharmacological compounds. In this study, we investigated the functional significance of the membrane distribution of NMDA and GABAA receptor subunits in relation to the accumulation of the tricyclic antidepressant desipramine (DMI) and the benzodiazepine diazepam (Diaz). In the presence of Triton X-100, which allowed proper separation of the lipid raft marker proteins caveolin-1 and flotillin-1 from the transferrin receptor, all receptor subunits were shifted to the non-raft fractions. In contrast, under detergent-free conditions, NMDA and GABAA receptor subunits were detected both in raft and non-raft fractions. Diaz was enriched in non-raft fractions without Triton X-100 in contrast to DMI, which preferentially accumulated in lipid rafts. Impairment of lipid raft integrity by methyl-β-cyclodextrine (MβCD)-induced cholesterol depletion did not change the inhibitory effect of DMI at the NMDA receptor, whereas it enhanced the potentiating effect of Diaz at the GABAA receptor at non-saturating concentrations of GABA. These results support the hypothesis that the interaction of benzodiazepines with the GABAA receptor likely occurs outside of lipid rafts while the antidepressant DMI acts on ionotropic receptors both within and outside these membrane microdomains.

Antiseizure Activity of Midazolam in Mice Lacking δ-Subunit Extrasynaptic GABAA Receptors

PubMed Central

Reddy, Sandesh D.; Younus, Iyan; Clossen, Bryan L.

2015-01-01

Midazolam is a benzodiazepine anticonvulsant with rapid onset and short duration of action. Midazolam is the current drug of choice for acute seizures and status epilepticus, including those caused by organophosphate nerve agents. The antiseizure activity of midazolam is thought to result from its allosteric potentiation of synaptic GABAA receptors in the brain. However, there are indications that benzodiazepines promote neurosteroid synthesis via the 18-kDa cholesterol transporter protein (TSPO). Therefore, we investigated the role of neurosteroids and their extrasynaptic GABAA receptor targets in the antiseizure activity of midazolam. Here, we used δ-subunit knockout (DKO) mice bearing a targeted deletion of the extrasynaptic receptors to investigate the contribution of the extrasynaptic receptors to the antiseizure activity of midazolam using the 6-Hz and hippocampus kindling seizure models. In both models, midazolam produced rapid and dose-dependent protection against seizures (ED50, 0.4 mg/kg). Moreover, the antiseizure potency of midazolam was undiminished in DKO mice compared with control mice. Pretreatment with PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide], a TSPO blocker, or finasteride, a 5α-reductase neurosteroid inhibitor, did not affect the antiseizure effect of midazolam. The antiseizure activity of midazolam was significantly reversed by pretreatment with flumazenil, a benzodiazepine antagonist. Plasma and brain levels of the neurosteroid allopregnanolone were not significantly greater in midazolam-treated animals. These studies therefore provide strong evidence that neurosteroids and extrasynaptic GABAA receptors are not involved in the antiseizure activity of midazolam, which mainly occurs through synaptic GABAA receptors via direct binding to benzodiazepine sites. This study reaffirms midazolam’s use for controlling acute seizures and status epilepticus. PMID:25784648

Towards functional selectivity for α6β3γ2 GABAA receptors: a series of novel pyrazoloquinolinones

PubMed Central

Treven, Marco; Siebert, David C B; Holzinger, Raphael; Bampali, Konstantina; Fabjan, Jure; Varagic, Zdravko; Wimmer, Laurin; Steudle, Friederike; Scholze, Petra; Schnürch, Michael; Mihovilovic, Marko D

2017-01-01

Background and Purpose The GABAA receptors are ligand‐gated ion channels, which play an important role in neurotransmission. Their variety of binding sites serves as an appealing target for many clinically relevant drugs. Here, we explored the functional selectivity of modulatory effects at specific extracellular α+/β− interfaces, using a systematically varied series of pyrazoloquinolinones. Experimental Approach Recombinant GABAA receptors were expressed in Xenopus laevis oocytes and modulatory effects on GABA‐elicited currents by the newly synthesized and reference compounds were investigated by the two‐electrode voltage clamp method. Key Results We identified a new compound which, to the best of our knowledge, shows the highest functional selectivity for positive modulation at α6β3γ2 GABAA receptors with nearly no residual activity at the other αxβ3γ2 (x = 1–5) subtypes. This modulation was independent of affinity for α+/γ− interfaces. Furthermore, we demonstrated for the first time a compound that elicits a negative modulation at specific extracellular α+/β− interfaces. Conclusion and Implications These results constitute a major step towards a potential selective positive modulation of certain α6‐containing GABAA receptors, which might be useful to elicit their physiological role. Furthermore, these studies pave the way towards insights into molecular principles that drive positive versus negative allosteric modulation of specific GABAA receptor isoforms. PMID:29127702

Effects of Antecedent GABAA Activation With Alprazolam on Counterregulatory Responses to Hypoglycemia in Healthy Humans

PubMed Central

Hedrington, Maka S.; Farmerie, Stephnie; Ertl, Andrew C.; Wang, Zhihui; Tate, Donna B.; Davis, Stephen N.

2010-01-01

OBJECTIVE To date, there are no data investigating the effects of GABAA activation on counterregulatory responses during repeated hypoglycemia in humans. The aim of this study was to determine the effects of prior GABAA activation using the benzodiazepine alprazolam on the neuroendocrine and autonomic nervous system (ANS) and metabolic counterregulatory responses during next-day hypoglycemia in healthy humans. RESEARCH DESIGN AND METHODS Twenty-eight healthy individuals (14 male and 14 female, age 27 ± 6 years, BMI 24 ± 3 kg/m2, and A1C 5.2 ± 0.1%) participated in four randomized, double-blind, 2-day studies. Day 1 consisted of either morning and afternoon 2-h hyperinsulinemic euglycemia or 2-h hyperinsulinemic hypoglycemia (2.9 mmol/l) with either 1 mg alprazolam or placebo administered 30 min before the start of each clamp. Day 2 consisted of a single-step hyperinsulinemic-hypoglycemic clamp of 2.9 mmol/l. RESULTS Despite similar hypoglycemia (2.9 ± 1 mmol/l) and insulinemia (672 ± 108 pmol/l) during day 2 studies, GABAA activation with alprazolam during day 1 euglycemia resulted in significant blunting (P < 0.05) of ANS (epinephrine, norepinephrine, muscle sympathetic nerve activity, and pancreatic polypeptide), neuroendocrine (glucagon and growth hormone), and metabolic (glucose kinetics, lipolysis, and glycogenolysis) counterregulatory responses. GABAA activation with alprazolam during prior hypoglycemia caused further significant (P < 0.05) decrements in subsequent glucagon, growth hormone, pancreatic polypeptide, and muscle sympathetic nerve activity counterregulatory responses. CONCLUSIONS Alprazolam activation of GABAA pathways during day 1 hypoglycemia can play an important role in regulating a spectrum of key physiologic responses during subsequent (day 2) hypoglycemia in healthy man. PMID:20086227

Effects of antecedent GABAA activation with alprazolam on counterregulatory responses to hypoglycemia in healthy humans.

PubMed

Hedrington, Maka S; Farmerie, Stephnie; Ertl, Andrew C; Wang, Zhihui; Tate, Donna B; Davis, Stephen N

2010-04-01

To date, there are no data investigating the effects of GABA(A) activation on counterregulatory responses during repeated hypoglycemia in humans. The aim of this study was to determine the effects of prior GABA(A) activation using the benzodiazepine alprazolam on the neuroendocrine and autonomic nervous system (ANS) and metabolic counterregulatory responses during next-day hypoglycemia in healthy humans. Twenty-eight healthy individuals (14 male and 14 female, age 27 +/- 6 years, BMI 24 +/- 3 kg/m(2), and A1C 5.2 +/- 0.1%) participated in four randomized, double-blind, 2-day studies. Day 1 consisted of either morning and afternoon 2-h hyperinsulinemic euglycemia or 2-h hyperinsulinemic hypoglycemia (2.9 mmol/l) with either 1 mg alprazolam or placebo administered 30 min before the start of each clamp. Day 2 consisted of a single-step hyperinsulinemic-hypoglycemic clamp of 2.9 mmol/l. Despite similar hypoglycemia (2.9 +/- 1 mmol/l) and insulinemia (672 +/- 108 pmol/l) during day 2 studies, GABA(A) activation with alprazolam during day 1 euglycemia resulted in significant blunting (P < 0.05) of ANS (epinephrine, norepinephrine, muscle sympathetic nerve activity, and pancreatic polypeptide), neuroendocrine (glucagon and growth hormone), and metabolic (glucose kinetics, lipolysis, and glycogenolysis) counterregulatory responses. GABA(A) activation with alprazolam during prior hypoglycemia caused further significant (P < 0.05) decrements in subsequent glucagon, growth hormone, pancreatic polypeptide, and muscle sympathetic nerve activity counterregulatory responses. Alprazolam activation of GABA(A) pathways during day 1 hypoglycemia can play an important role in regulating a spectrum of key physiologic responses during subsequent (day 2) hypoglycemia in healthy man.

Extrasynaptic GABAA receptors in the crosshairs of hormones and ethanol

PubMed Central

Mody, Istvan

2008-01-01

Gamma-aminobutyric acid (GABA) is the main chemical inhibitory neurotransmitter in the brain. In the central nervous system (CNS) it acts on two distinct types of receptor: an ion channel, i.e., an “ionotropic” receptor permeable to Cl− and HCO3− (GABAA receptors) and a G-protein coupled “metabotropic” receptor that is linked to various effector mechanisms (GABAB receptors). This review will summarize novel developments in the physiology and pharmacology of GABAA receptors (GABAARs), specifically those found outside synapses. The focus will be on a particular combination of GABAAR subunits sensitive to ovarian and adrenal cortical steroid hormone metabolites that are synthesized in the brain (neurosteroids) and to sobriety impairing concentrations of ethanol. These receptors may be the final common pathway for interactions between ethanol and ovarian and stress-related neurosteroids. PMID:17714830

Methods for recording and measuring tonic GABAA receptor-mediated inhibition

PubMed Central

Bright, Damian P.; Smart, Trevor G.

2013-01-01

Tonic inhibitory conductances mediated by GABAA receptors have now been identified and characterized in many different brain regions. Most experimental studies of tonic GABAergic inhibition have been carried out using acute brain slice preparations but tonic currents have been recorded under a variety of different conditions. This diversity of recording conditions is likely to impact upon many of the factors responsible for controlling tonic inhibition and can make comparison between different studies difficult. In this review, we will firstly consider how various experimental conditions, including age of animal, recording temperature and solution composition, are likely to influence tonic GABAA conductances. We will then consider some technical considerations related to how the tonic conductance is measured and subsequently analyzed, including how the use of current noise may provide a complementary and reliable method for quantifying changes in tonic current. PMID:24367296

Dynamic differentiation of GABAA-sensitive influences on orientation selectivity of complex cells in the cat striate cortex.

PubMed

Pfleger, B; Bonds, A B

1995-01-01

The influence of GABAA receptors on orientation selectivity of cat complex cells was tested by iontophoresis of the GABAA receptor blockers bicuculline and N-methyl-bicuculline while stimulating with drifting sinusoidal gratings. Reduction of orientation tuning was markedly less than reported in previous studies that used drifting bars as visual stimuli. Only 3/31 cells lost orientation selectivity, with an average increase in bandwidth of 33%, as opposed to half the cells losing selectivity and a bandwidth increase for the remainder of 47% as reported previously. Infusion of GABAA blockers revealed a prominent stimulus onset transient response, lasting about 120 ms, that showed a broadening of orientation selectivity comparable to that found using drifting bars under similar circumstances. We believe that drifting gratings emphasize a steady-state response component that retains, in the presence of GABAA blockers, significant orientation selectivity. Because the onset transient is initially unselective for orientation, we suggest that the steady-state, orientation-selective response component develops from an alternate inhibitory mechanism, possibly mediated by GABAB receptors.

Characterization of GABA/sub A/ receptor-mediated /sup 36/chloride uptake in rat brain synaptoneurosomes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luu, M.D.; Morrow, A.L.; Paul, S.M.

1987-09-07

..gamma..-Aminobutyric acid (GABA) receptor-mediated /sup 36/chloride (/sup 36/Cl/sup -/) uptake was measured in synaptoneurosomes from rat brain. GABA and GABA agonists stimulated /sup 36/Cl/sup -/ uptake in a concentration-dependent manner with the following order of potency: Muscimol>GABA>piperidine-4-sulfonic acid (P4S)>4,5,6,7-tetrahydroisoxazolo-(5,4-c)pyridin-3-ol (THIP)=3-aminopropanesulfonic acid (3APS)>>taurine. Both P4S and 3APS behaved as partial agonists, while the GABA/sub B/ agonist, baclofen, was ineffective. The response to muscimol was inhibited by bicuculline and picrotoxin in a mixed competitive/non-competitive manner. Other inhibitors of GABA receptor-opened channels or non-neuronal anion channels such as penicillin, picrate, furosemide and disulfonic acid stilbenes also inhibited the response to muscimol. A regionalmore » variation in muscimol-stimulated /sup 36/Cl/sup -/ uptake was observed; the largest responses were observed in the cerebral cortex, cerebellum and hippocampus, moderate responses were obtained in the striatum and hypothalamus and the smallest response was observed in the pons-medulla. GABA receptor-mediated /sup 36/Cl/sup -/ uptake was also dependent on the anion present in the media. The muscinol response varied in media containing the following anions: Br/sup -/>Cl/sup -/greater than or equal toNO/sub 3//sup -/>I/sup -/greater than or equal toSCN/sup -/>>C/sub 3/H/sub 5/OO/sup -/greater than or equal toClO/sub 4//sup -/>F/sup -/, consistent with the relative anion permeability through GABA receptor-gated anion channels and the enhancement of convulsant binding to the GABA receptor-gated Cl/sup -/ channel. 43 references, 4 figures, 3 tables.« less

RDX Binds to the GABAA Receptor–Convulsant Site and Blocks GABAA Receptor–Mediated Currents in the Amygdala: A Mechanism for RDX-Induced Seizures

PubMed Central

Williams, Larry R.; Aroniadou-Anderjaska, Vassiliki; Qashu, Felicia; Finne, Huckelberry; Pidoplichko, Volodymyr; Bannon, Desmond I.; Braga, Maria F. M.

2011-01-01

Background Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) is a high-energy, trinitrated cyclic compound that has been used worldwide since World War II as an explosive in both military and civilian applications. RDX can be released in the environment by way of waste streams generated during the manufacture, use, and disposal of RDX-containing munitions and can leach into groundwater from unexploded munitions found on training ranges. For > 60 years, it has been known that exposure to high doses of RDX causes generalized seizures, but the mechanism has remained unknown. Objective We investigated the mechanism by which RDX induces seizures. Methods and results By screening the affinity of RDX for a number of neurotransmitter receptors, we found that RDX binds exclusively to the picrotoxin convulsant site of the γ-aminobutyric acid type A (GABAA) ionophore. Whole-cell in vitro recordings in the rat basolateral amygdala (BLA) showed that RDX reduces the frequency and amplitude of spontaneous GABAA receptor–mediated inhibitory postsynaptic currents and the amplitude of GABA-evoked postsynaptic currents. In extracellular field recordings from the BLA, RDX induced prolonged, seizure-like neuronal discharges. Conclusions These results suggest that binding to the GABAA receptor convulsant site is the primary mechanism of seizure induction by RDX and that reduction of GABAergic inhibitory transmission in the amygdala is involved in the generation of RDX-induced seizures. Knowledge of the molecular site and the mechanism of RDX action with respect to seizure induction can guide therapeutic strategies, allow more accurate development of safe thresholds for exposures, and help prevent the development of new explosives or other munitions that could pose similar health risks. PMID:21362589

α4βδ GABAA Receptors Reduce Dendritic Spine Density In CA1 Hippocampus And Impair Relearning Ability Of Adolescent Female Mice: Effects Of A GABA Agonist And A Stress Steroid

PubMed Central

Afroz, Sonia; Shen, Hui; Smith, Sheryl S.

2017-01-01

Synaptic pruning underlies the transition from an immature to an adult CNS through refinements of neuronal circuits. Our recent study indicates that pubertal synaptic pruning is triggered by the inhibition generated by extrasynaptic α4βδ GABAA receptors (GABARs) which are increased for 10 d on dendritic spines of CA1 pyramidal cells at the onset of puberty (PND 35–44) in the female mouse, suggesting α4βδ GABARs as a novel target for the regulation of adolescent synaptic pruning. In the present study we used a pharmacological approach to further examine the role of these receptors in altering spine density during puberty of female mice and the impact of these changes on spatial learning, assessed in adulthood. Two drugs were chronically administered during the pubertal period (PND 35–44): the GABA agonist gaboxadol (GBX, 0.1 mg/kg, i.p.), to enhance current gated by α4βδ GABARs and the neurosteroid/stress steroid THP (3α-OH-5β-pregnan-20-one, 10 mg/kg, i.p.) to decrease expression of α4βδ. Spine density was determined on PND 56 with Golgi staining. Spatial learning and relearning were assessed using the multiple object relocation task (MPORT) and an active place avoidance task (APA) on PND 56. Pubertal GBX decreased spine density post-pubertally by 70% (P<0.05), while decreasing α4βδ expression with THP increased spine density by two-fold (P<0.05), in both cases, with greatest effects on the mushroom spines. Adult relearning ability was compromised in both hippocampus-dependent tasks after pubertal administration of either drug. These findings suggest that an optimal spine density produced by α4βδ GABARs is necessary for optimal cognition in adults. PMID:28189613

Extrasynaptic αβ subunit GABAA receptors on rat hippocampal pyramidal neurons

PubMed Central

Mortensen, Martin; Smart, Trevor G

2006-01-01

Extrasynaptic GABAA receptors that are tonically activated by ambient GABA are important for controlling neuronal excitability. In hippocampal pyramidal neurons, the subunit composition of these extrasynaptic receptors may include α5βγ and/or α4βδ subunits. Our present studies reveal that a component of the tonic current in the hippocampus is highly sensitive to inhibition by Zn2+. This component is probably not mediated by either α5βγ or α4βδ receptors, but might be explained by the presence of αβ isoforms. Using patch-clamp recording from pyramidal neurons, a small tonic current measured in the absence of exogenous GABA exhibited both high and low sensitivity to Zn2+ inhibition (IC50 values, 1.89 and 223 μm, respectively). Using low nanomolar and micromolar GABA concentrations to replicate tonic currents, we identified two components that are mediated by benzodiazepine-sensitive and -insensitive receptors. The latter indicated that extrasynaptic GABAA receptors exist that are devoid of γ2 subunits. To distinguish whether the benzodiazepine-insensitive receptors were αβ or αβδ isoforms, we used single-channel recording. Expressing recombinant α1β3γ2, α5β3γ2, α4β3δ and α1β3 receptors in human embryonic kidney (HEK) or mouse fibroblast (Ltk) cells, revealed similar openings with high main conductances (∼25–28 pS) for γ2 or δ subunit-containing receptors whereas αβ receptors were characterized by a lower main conductance state (∼11 pS). Recording from pyramidal cell somata revealed a similar range of channel conductances, indicative of a mixture of GABAA receptors in the extrasynaptic membrane. The lowest conductance state (∼11 pS) was the most sensitive to Zn2+ inhibition in accord with the presence of αβ receptors. This receptor type is estimated to account for up to 10% of all extrasynaptic GABAA receptors on hippocampal pyramidal neurons. PMID:17023503

Activation of the medial prefrontal cortex by escapable stress is necessary for protection against subsequent inescapable stress-induced potentiation of morphine conditioned place preference.

PubMed

Rozeske, Robert R; Der-Avakian, Andre; Watkins, Linda R; Maier, Steven F

2012-01-01

Stress can be a predisposing factor in the development of psychiatric disorders. However, not all individuals develop psychiatric disorders following a traumatic event. An attempt to understand these individual differences has led to a focus on factors that produce resistance. Interestingly, in rats, an experience with escapable tailshock (ES) before inescapable tailshock (IS) prevents the typical anxiety-like behavioral outcomes of IS. This type of resistance has been termed 'behavioral immunization', and it depends on activation of the medial prefrontal cortex (mPFC) during ES. However, one outcome of IS that is not anxiety-related is potentiation of morphine conditioned place preference (CPP). The present experiments investigated whether prior ES would block IS-induced potentiation of morphine CPP. Rats received either ES, IS or homecage control treatment on day 1 and then either IS or homecage control treatment on day 2. Twenty-four hours following day 2, rats underwent morphine conditioning, and CPP was subsequently assessed. In a second experiment, rats received ES 3, 14 or 56 days prior to IS to determine the duration of behavioral immunization. In a final experiment, rats were microinjected with the GABA(A) agonist muscimol (50 ng/0.5 μL) or saline in the mPFC before day 1 of stress. Prior ES blocked IS-induced potentiation of morphine CPP. This immunizing effect of ES lasted for at least 56 days. Additionally, intra-mPFC muscimol during ES prevented behavioral immunization. These results suggest that prior experience with ES activates the mPFC and produces long-lasting neural alterations that block subsequent IS-induced potentiation of morphine CPP. © 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

A critical role of lateral hypothalamus in context-induced relapse to alcohol seeking after punishment-imposed abstinence.

PubMed

Marchant, Nathan J; Rabei, Rana; Kaganovsky, Konstantin; Caprioli, Daniele; Bossert, Jennifer M; Bonci, Antonello; Shaham, Yavin

2014-05-28

In human alcoholics, abstinence is often self-imposed, despite alcohol availability, because of the negative consequences of excessive use. During abstinence, relapse is often triggered by exposure to contexts associated with alcohol use. We recently developed a rat model that captures some features of this human condition: exposure to the alcohol self-administration environment (context A), after punishment-imposed suppression of alcohol self-administration in a different environment (context B), provoked renewal of alcohol seeking in alcohol-preferring P rats. The mechanisms underlying context-induced renewal of alcohol seeking after punishment-imposed abstinence are unknown. Here, we studied the role of the lateral hypothalamus (LH) and its forebrain projections in this effect. We first determined the effect of context-induced renewal of alcohol seeking on Fos (a neuronal activity marker) expression in LH. We next determined the effect of LH reversible inactivation by GABAA + GABAB receptor agonists (muscimol + baclofen) on this effect. Finally, we determined neuronal activation in brain areas projecting to LH during context-induced renewal tests by measuring double labeling of the retrograde tracer cholera toxin subunit B (CTb; injected in LH) with Fos. Context-induced renewal of alcohol seeking after punishment-imposed abstinence was associated with increased Fos expression in LH. Additionally, renewal was blocked by muscimol + baclofen injections into LH. Finally, double-labeling analysis of CTb + Fos showed that context-induced renewal of alcohol seeking after punishment-imposed abstinence was associated with selective activation of accumbens shell neurons projecting to LH. The results demonstrate an important role of LH in renewal of alcohol seeking after punishment-imposed abstinence and suggest a role of accumbens shell projections to LH in this form of relapse. Copyright © 2014 the authors 0270-6474/14/347447-11$15.00/0.

( sup 3 H)RO15-4513 binding to cerebellar diazepam-sensitive and insensitive GABAA receptors is unchanged by one week of ethanol intake

DOE Office of Scientific and Technical Information (OSTI.GOV)

Martin, M.W.; Chen, J.P.; Wallis, C.

1992-02-26

({sup 3}H)RO15-4513, a partial inverse agonist at GABAA receptors, binds to two sites in cerebellar membranes, one sensitive (DZ-S) and one insensitive (DZ-IS) to inhibition by diazepam. These binding sites may represent different isoforms of the GABAA receptor and may play a role in ethanol (EtOH) dependence. The authors tested the hypothesis that chronic intake of EtOH induces changes in the binding properties of one or both of these putative GABBA receptors. Rats were fed a liquid diet of 4.5% EtOH for 7 d, gavaged with a 3g/kg dose of EtOH, and then sacrificed after 2 h, 12 h, ormore » 4.5 d. Binding of ({sup 3}H)RO15-4513 to cerebellar membranes was performed in the absence or presence of 10{mu}M diazepam (DZ-IS binding); DZ-S binding was calculated as the difference between total and DZ-IS. Nonlinear regression analysis showed that each class of binding site fit a model of mass action binding to a single, noninteractive population of sites. No significant difference was observed between any of the treatment groups in the apparent affinity (Kd) for ({sup 3}H)RO15-4513 at total, DZ-S, or DZ-IS sites following chronic EtOH intake or withdrawal. In addition, no significant difference was observed in the apparent number of DZ-S or DZ-IS binding sites or the ratio of DZ-S to DZ-IS.« less

The nucleus of the optic tract. Its function in gaze stabilization and control of visual-vestibular interaction

NASA Technical Reports Server (NTRS)

Cohen, B.; Reisine, H.; Yokota, J. I.; Raphan, T.

1992-01-01

1. Electrical stimulation of the nucleus of the optic tract (NOT) induced nystagmus and after-nystagmus with ipsilateral slow phases. The velocity characteristics of the nystagmus were similar to those of the slow component of optokinetic nystagmus (OKN) and to optokinetic after-nystagmus (OKAN), both of which are produced by velocity storage in the vestibular system. When NOT was destroyed, these components disappeared. This indicates that velocity storage is activated from the visual system through NOT. 2. Velocity storage produces compensatory eye-in-head and head-on-body movements through the vestibular system. The association of NOT with velocity storage implies that NOT helps stabilize gaze in space during both passive motion and active locomotion in light with an angular component. It has been suggested that "vestibular-only" neurons in the vestibular nuclei play an important role in generation of velocity storage. Similarities between the rise and fall times of eye velocity during OKN and OKAN to firing rates of vestibular-only neurons suggest that these cells may receive their visual input through NOT. 3. One NOT was injected with muscimol, a GABAA agonist. Ipsilateral OKN and OKAN were lost, suggesting that GABA, which is an inhibitory transmitter in NOT, acts on projection pathways to the brain stem. A striking finding was that visual suppression and habituation of contralateral slow phases of vestibular nystagmus were also abolished after muscimol injection. The latter implies that NOT plays an important role in producing visual suppression of the VOR and habituating its time constant. 4. Habituation is lost after nodulus and uvula lesions and visual suppression after lesions of the flocculus and paraflocculus. We postulate that the disappearance of vestibular habituation and of visual suppression of vestibular responses after muscimol injections was due to dysfacilitation of the prominent NOT-inferior olive pathway, inactivating climbing fibers from

Metabotropic Glutamate Receptors in the Trafficking of Ionotropic Glutamate and GABAA Receptors at Central Synapses

PubMed Central

Xiao, Min-Yi; Gustafsson, Bengt; Niu, Yin-Ping

2006-01-01

The trafficking of ionotropic glutamate (AMPA, NMDA and kainate) and GABAA receptors in and out of, or laterally along, the postsynaptic membrane has recently emerged as an important mechanism in the regulation of synaptic function, both under physiological and pathological conditions, such as information processing, learning and memory formation, neuronal development, and neurodegenerative diseases. Non-ionotropic glutamate receptors, primarily group I metabotropic glutamate receptors (mGluRs), co-exist with the postsynaptic ionotropic glutamate and GABAA receptors. The ability of mGluRs to regulate postsynaptic phosphorylation and Ca2+ concentration, as well as their interactions with postsynaptic scaffolding/signaling proteins, makes them well suited to influence the trafficking of ionotropic glutamate and GABAA receptors. Recent studies have provided insights into how mGluRs may impose such an influence at central synapses, and thus how they may affect synaptic signaling and the maintenance of long-term synaptic plasticity. In this review we will discuss some of the recent progress in this area: i) long-term synaptic plasticity and the involvement of mGluRs; ii) ionotropic glutamate receptor trafficking and long-term synaptic plasticity; iii) the involvement of postsynaptic group I mGluRs in regulating ionotropic glutamate receptor trafficking; iv) involvement of postsynaptic group I mGluRs in regulating GABAA receptor trafficking; v) and the trafficking of postsynaptic group I mGluRs themselves. PMID:18615134

GABA(A) receptors in visual and auditory cortex and neural activity changes during basic visual stimulation.

PubMed

Qin, Pengmin; Duncan, Niall W; Wiebking, Christine; Gravel, Paul; Lyttelton, Oliver; Hayes, Dave J; Verhaeghe, Jeroen; Kostikov, Alexey; Schirrmacher, Ralf; Reader, Andrew J; Northoff, Georg

2012-01-01

Recent imaging studies have demonstrated that levels of resting γ-aminobutyric acid (GABA) in the visual cortex predict the degree of stimulus-induced activity in the same region. These studies have used the presentation of discrete visual stimulus; the change from closed eyes to open also represents a simple visual stimulus, however, and has been shown to induce changes in local brain activity and in functional connectivity between regions. We thus aimed to investigate the role of the GABA system, specifically GABA(A) receptors, in the changes in brain activity between the eyes closed (EC) and eyes open (EO) state in order to provide detail at the receptor level to complement previous studies of GABA concentrations. We conducted an fMRI study involving two different modes of the change from EC to EO: an EO and EC block design, allowing the modeling of the haemodynamic response, followed by longer periods of EC and EO to allow the measuring of functional connectivity. The same subjects also underwent [(18)F]Flumazenil PET to measure GABA(A) receptor binding potentials. It was demonstrated that the local-to-global ratio of GABA(A) receptor binding potential in the visual cortex predicted the degree of changes in neural activity from EC to EO. This same relationship was also shown in the auditory cortex. Furthermore, the local-to-global ratio of GABA(A) receptor binding potential in the visual cortex also predicted the change in functional connectivity between the visual and auditory cortex from EC to EO. These findings contribute to our understanding of the role of GABA(A) receptors in stimulus-induced neural activity in local regions and in inter-regional functional connectivity.

Fragrances in oolong tea that enhance the response of GABAA receptors.

PubMed

Hossain, Sheikh Julfikar; Aoshima, Hitoshi; Koda, Hirofumi; Kiso, Yoshinobu

2004-09-01

We electrophysiologically investigated the effect of some fragrant compounds in oolong tea on the response of ionotropic gamma-aminobutyric acid (GABA) receptors (GABAA receptors) which were expressed in Xenopus oocytes. Of the tested fragrances in oolong tea, cis-jasmone, jasmine lactone, linalool oxide and methyl jasmonate significantly potentiated the response. Among these, cis-jasmone and methyl jasmonate potently potentiated the response, having a respective dissociation constant of the compound (Kp) and maximum potentiation (Vm) of 0.49 mM and 322% for cis-jasmone, and 0.84 mM and 450% for methyl jasmonate. Inhalation of 0.1% cis-jasmone or methyl jasmonate significantly increased the sleeping time of mice induced by pentobarbital, suggesting that these fragrant compounds were absorbed by the brain and thereby potentiated the GABAA receptor response. Both of these compounds may therefore have a tranquillizing effect on the brain.

Nucleus Accumbens Adenosine A2A Receptors Regulate Exertion of Effort by Acting on the Ventral Striatopallidal Pathway

PubMed Central

Mingote, Susana; Font, Laura; Farrar, Andrew M.; Vontell, Regina; Worden, Lila T.; Stopper, Colin M.; Port, Russell G.; Sink, Kelly S.; Bunce, Jamie G.; Chrobak, James J.; Salamone, John D.

2009-01-01

Goal-directed actions are sensitive to work-related response costs, and dopamine in nucleus accumbens is thought to modulate the exertion of effort in motivated behavior. Dopamine-rich striatal areas such as nucleus accumbens also contain high numbers of adenosine A2A receptors, and, for that reason, the behavioral and neurochemical effects of the adenosine A2A receptor agonist CGS 21680 [2-p-(2-carboxyethyl) phenethylamino-5′-N-ethylcarboxamidoadenosine] were investigated. Stimulation of accumbens adenosine A2A receptors disrupted performance of an instrumental task with high work demands (i.e., an interval lever-pressing schedule with a ratio requirement attached) but had little effect on a task with a lower work requirement. Immunohistochemical studies revealed that accumbens neurons that project to the ventral pallidum showed adenosine A2A receptors immunoreactivity. Moreover, activation of accumbens A2A receptors by local injections of CGS 21680 increased extracellular GABA levels in the ventral pallidum. Combined contralateral injections of CGS 21680 into the accumbens and the GABAA agonist muscimol into ventral pallidum (i.e., “disconnection” methods) also impaired response output, indicating that these structures are part of a common neural circuitry regulating the exertion of effort. Thus, accumbens adenosine A2A receptors appear to regulate behavioral activation and effort-related processes by modulating the activity of the ventral striatopallidal pathway. Research on the effort-related functions of these forebrain systems may lead to a greater understanding of pathological features of motivation, such as psychomotor slowing, anergia, and fatigue in depression. PMID:18768698

HPLC-Based Activity Profiling: Discovery of Piperine as a Positive GABAA Receptor Modulator Targeting a Benzodiazepine-Independent Binding Site

PubMed Central

Zaugg, Janine; Baburin, Igor; Strommer, Barbara; Kim, Hyun-Jung; Hering, Steffen; Hamburger, Matthias

2011-01-01

A plant extract library was screened for GABAA receptor activity making use of a two-microelectrode voltage clamp assay on Xenopus laevis oocytes. An ethyl acetate extract of black pepper fruits [Piper nigrum L. (Piperaceae) 100 μg/mL] potentiated GABA-induced chloride currents through GABAA receptors (composed of α1, β2, and γ2S subunits) by 169.1 ± 2.4%. With the aid of an HPLC-based activity profiling approach, piperine (5) was identified as the main active compound, together with 12 structurally related less active or inactive piperamides (1–4, 6–13). Identification was achieved by on-line high-resolution mass spectrometry and off-line microprobe 1D and 2D NMR spectroscopy, using only milligram amounts of extract. Compound 5 induced a maximum potentiation of the chloride currents by 301.9 ± 26.5% with an EC50 of 52.4 ± 9.4 μM. A comparison of the modulatory activity of 5 and other naturally occurring piperamides enabled insights into structural features critical for GABAA receptor modulation. The stimulation of chloride currents through GABAA receptors by compound 5 was not antagonized by flumazenil (10 μM). These data show that piperine (5) represents a new scaffold of positive allosteric GABAA receptor modulators targeting a benzodiazepine-independent binding site. PMID:20085307

Involvement of glutamatergic N-methyl-d-aspartate receptors in the expression of increased head-dipping behaviors in the hole-board tests of olfactory bulbectomized mice.

PubMed

Hirose, Noritaka; Saitoh, Akiyoshi; Kamei, Junzo

2016-10-01

Olfactory bulbectomized (OB) mice produce agitated anxiety-like behaviors in the hole-board test, which was expressed by an increase in head-dipping counts and a decrease in head-dipping latencies. However, the associated mechanisms remain unclear. In the present study, MK-801 (10, 100μg/kg), a selective N-methyl-d-aspartate (NMDA) receptor antagonist, significantly and dose-dependently suppressed the increased head-dipping behaviors in OB mice, without affecting sham mice. Similar results were obtained with another selective NMDA receptor antagonist D-AP5 treatment in OB mice. On the other hand, muscimol, a selective aminobutyric acid type A (GABAA) receptor agonist produced no effects on these hyperemotional behaviors in OB mice at a dose (100μg/kg) that produced anxiolytic-like effects in sham mice. Interestingly, glutamine contents and glutamine/glutamate ratios were significantly increased in the amygdala and frontal cortex of OB mice compared to sham mice. Based on these results, we concluded that the glutamatergic NMDA receptors are involved in the expression of increased head-dipping behaviors in the hole-board tests of OB mice. Accordingly, the changes in glutamatergic transmission in frontal cortex and amygdala may play important roles in the expression of these abnormal behaviors in OB mice. Copyright © 2016. Published by Elsevier B.V.

Antiseizure Activity of Midazolam in Mice Lacking δ-Subunit Extrasynaptic GABA(A) Receptors.

PubMed

Reddy, Sandesh D; Younus, Iyan; Clossen, Bryan L; Reddy, Doodipala Samba

2015-06-01

Midazolam is a benzodiazepine anticonvulsant with rapid onset and short duration of action. Midazolam is the current drug of choice for acute seizures and status epilepticus, including those caused by organophosphate nerve agents. The antiseizure activity of midazolam is thought to result from its allosteric potentiation of synaptic GABA(A) receptors in the brain. However, there are indications that benzodiazepines promote neurosteroid synthesis via the 18-kDa cholesterol transporter protein (TSPO). Therefore, we investigated the role of neurosteroids and their extrasynaptic GABA(A) receptor targets in the antiseizure activity of midazolam. Here, we used δ-subunit knockout (DKO) mice bearing a targeted deletion of the extrasynaptic receptors to investigate the contribution of the extrasynaptic receptors to the antiseizure activity of midazolam using the 6-Hz and hippocampus kindling seizure models. In both models, midazolam produced rapid and dose-dependent protection against seizures (ED50, 0.4 mg/kg). Moreover, the antiseizure potency of midazolam was undiminished in DKO mice compared with control mice. Pretreatment with PK11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide], a TSPO blocker, or finasteride, a 5α-reductase neurosteroid inhibitor, did not affect the antiseizure effect of midazolam. The antiseizure activity of midazolam was significantly reversed by pretreatment with flumazenil, a benzodiazepine antagonist. Plasma and brain levels of the neurosteroid allopregnanolone were not significantly greater in midazolam-treated animals. These studies therefore provide strong evidence that neurosteroids and extrasynaptic GABA(A) receptors are not involved in the antiseizure activity of midazolam, which mainly occurs through synaptic GABA(A) receptors via direct binding to benzodiazepine sites. This study reaffirms midazolam's use for controlling acute seizures and status epilepticus. Copyright © 2015 by The American Society for

Metabotropic glutamate receptors in the trafficking of ionotropic glutamate and GABA(A) receptors at central synapses.

PubMed

Xiao, Min-Yi; Gustafsson, Bengt; Niu, Yin-Ping

2006-01-01

The trafficking of ionotropic glutamate (AMPA, NMDA and kainate) and GABA(A) receptors in and out of, or laterally along, the postsynaptic membrane has recently emerged as an important mechanism in the regulation of synaptic function, both under physiological and pathological conditions, such as information processing, learning and memory formation, neuronal development, and neurodegenerative diseases. Non-ionotropic glutamate receptors, primarily group I metabotropic glutamate receptors (mGluRs), co-exist with the postsynaptic ionotropic glutamate and GABA(A) receptors. The ability of mGluRs to regulate postsynaptic phosphorylation and Ca(2+) concentration, as well as their interactions with postsynaptic scaffolding/signaling proteins, makes them well suited to influence the trafficking of ionotropic glutamate and GABA(A) receptors. Recent studies have provided insights into how mGluRs may impose such an influence at central synapses, and thus how they may affect synaptic signaling and the maintenance of long-term synaptic plasticity. In this review we will discuss some of the recent progress in this area: i) long-term synaptic plasticity and the involvement of mGluRs; ii) ionotropic glutamate receptor trafficking and long-term synaptic plasticity; iii) the involvement of postsynaptic group I mGluRs in regulating ionotropic glutamate receptor trafficking; iv) involvement of postsynaptic group I mGluRs in regulating GABA(A) receptor trafficking; v) and the trafficking of postsynaptic group I mGluRs themselves.

Novel analogues of chlormethiazole are neuroprotective in four cellular models of neurodegeneration by a mechanism with variable dependence on GABAA receptor potentiation

PubMed Central

VandeVrede, Lawren; Tavassoli, Ehsan; Luo, Jia; Qin, Zhihui; Yue, Lan; Pepperberg, David R; Thatcher, Gregory R

2014-01-01

Background and Purpose: Chlormethiazole (CMZ), a clinical sedative/anxiolytic agent, did not reach clinical efficacy in stroke trials despite neuroprotection demonstrated in numerous animal models. Using CMZ as a lead compound, neuroprotective methiazole (MZ) analogues were developed, and neuroprotection and GABAA receptor dependence were studied. Experimental Approach: Eight MZs were selected from a novel library, of which two were studied in detail. Neuroprotection, glutamate release, intracellular calcium and response to GABA blockade by picrotoxin were measured in rat primary cortical cultures using four cellular models of neurodegeneration. GABA potentiation was assayed in oocytes expressing the α1β2γ2 GABAA receptor. Key Results: Neuroprotection against a range of insults was retained even with substantial chemical modification. Dependence on GABAA receptor activity was variable: at the extremes, neuroprotection by GN-28 was universally sensitive to picrotoxin, while GN-38 was largely insensitive. In parallel, effects on extracellular glutamate and intracellular calcium were associated with GABAA dependence. Consistent with these findings, GN-28 potentiated α1β2γ2 GABAA function, whereas GN-38 had a weak inhibitory effect. Neuroprotection against moderate dose oligomeric Aβ1–42 was also tolerant to structural changes. Conclusions and Implications: The results support the concept that CMZ does not contain a single pharmacophore, rather that broad-spectrum neuroprotection results from a GABAA-dependent mechanism represented by GN-28, combined with a mechanism represented in GN-38 that shows the least dependence on GABAA receptors. These findings allow further refinement of the neuroprotective pharmacophore and investigation into secondary mechanisms that will assist in identifying MZ-based compounds of use in treating neurodegeneration. PMID:24116891

Nicotine and Nicotine Abstinence Do Not Interfere with GABAA Receptor Neuroadaptations During Alcohol Abstinence.

PubMed

Hillmer, Ansel T; Kloczynski, Tracy; Sandiego, Christine M; Pittman, Brian; Anderson, Jon M; Labaree, David; Gao, Hong; Huang, Yiyun; Deluliis, Giuseppe; O'Malley, Stephanie S; Carson, Richard E; Cosgrove, Kelly P

2016-04-01

Alcohol dependence and tobacco smoking are highly comorbid, and treating both conditions simultaneously is controversial. Previously, we showed that tobacco smoking interferes with GABAA receptor neuroadaptations during alcohol withdrawal in humans, while this effect did not occur with continued nicotine use during alcohol abstinence in nonhuman primates. Here, we extend our previous work by measuring GABAA receptor availability with positron emission tomography (PET) during drug abstinence in nonhuman primates exposed to alcohol alone, nicotine and alcohol together, and alcohol abstinence with continued nicotine exposure. Twenty-four adolescent male rhesus macaques orally self-administered alcohol and nicotine, available separately in water and saccharin, over 20 weeks. The groups included alcohol alone (n = 8); nicotine and alcohol with simultaneous abstinence (n = 8); nicotine and alcohol with alcohol abstinence while nicotine was still available (n = 8); and a pilot group of animals consuming nicotine alone (n = 6). Animals were imaged with [(11)C]flumazenil PET to measure binding potential (BPND), an index of GABAA receptor availability. Imaging occurred at baseline (drug-naíve), and following alcohol and/or nicotine cessation at 1 day, 8 days, and 12 weeks of abstinence. Generalized linear mixed models were used to examine the time course of [(11)C]flumazenil BPND during alcohol abstinence across groups. Animals consumed 3.95 ± 1.22 g/kg/d alcohol and 55.4 ± 35.1 mg/kg/d nicotine. No significant group effects were observed in [(11)C]flumazenil BPND during alcohol abstinence; however, a main effect of time was detected. Post hoc analyses indicated that all groups abstaining from alcohol exhibited significantly increased GABAA receptor availability at 1 day and 8 days (but not 12 weeks) of abstinence relative to baseline, while no changes in [(11)C]flumazenil BPND during nicotine abstinence alone were observed. These data indicate that neither nicotine nor

Neonatal blockade of GABA-A receptors alters behavioral and physiological phenotypes in adult mice.

PubMed

Salari, Ali-Akbar; Amani, Mohammad

2017-04-01

Gamma-aminobutyric acid (GABA) plays an inhibitory role in the mature brain, and has a complex and bidirectional effect in different parts of the immature brain which affects proliferation, migration and differentiation of neurons during development. There is also increasing evidence suggesting that activation or blockade of the GABA-A receptors during early life can induce brain and behavioral abnormalities in adulthood. We investigated whether neonatal blockade of the GABA-A receptors by bicuculline can alter anxiety- and depression-like behaviors, body weight, food intake, corticosterone and testosterone levels in adult mice (postnatal days 80-95). To this end, neonatal mice were treated with either DMSO or bicuculline (70, 150 and 300μg/kg) during postnatal days 7, 9 and 11. When grown to adulthood, mice were exposed to behavioral tests to measure anxiety- (elevated plus-maze and light-dark box) and depression-like behaviors (tail suspension test and forced swim test). Stress-induced serum corticosterone and testosterone levels, body weight and food intake were also evaluated. Neonatal bicuculline exposure at dose of 300μg/kg decreased anxiety-like behavior, stress-induced corticosterone levels and increased testosterone levels, body weight and food intake, without significantly influencing depression-like behavior in adult male mice. However, no significant changes in these parameters were observed in adult females. These findings suggest that neonatal blockade of GABA-A receptors affects anxiety-like behavior, physiological and hormonal parameters in a sex-dependent manner in mice. Taken together, these data corroborate the concept that GABA-A receptors during early life have an important role in programming neurobehavioral phenotypes in adulthood. Copyright © 2017 ISDN. Published by Elsevier Ltd. All rights reserved.

Garcinol Upregulates GABAA and GAD65 Expression, Modulates BDNF-TrkB Pathway to Reduce Seizures in Pentylenetetrazole (PTZ)-Induced Epilepsy

PubMed Central

Hao, Fang; Jia, Li-Hua; Li, Xiao-Wan; Zhang, Ying-Rui; Liu, Xue-Wu

2016-01-01

Background Epilepsy is the most predominant neurological disorder characterized by recurrent seizures. Despite treatment with antiepileptic drugs, epilepsy still is a challenge to treat, due to the associated adverse effects of the drugs. Previous investigations have shown critical roles of BDNF-TrkB signalling and expression of glutamic acid decarboxylase 65 (GAD65) and GABAA in the brain during epilepsy. Thus, drugs that could modulate BDNF-TrkB signal and expression of GAD65 and GABAA could aid in therapy. Recent experimental data have focussed on plant-derived compounds in treatments. Garcinol (camboginol), is a polyisoprenylated benzophenone derived from the fruit of Garcinia indica. We investigated the effects of garcinol in pentylenetetrazole (PTZ)-induced epileptic models. Material/Methods Seizure scores were measured in epilepsy kindled mice. Neuronal degeneration and apoptosis were assessed by Nissl staining, TUNEL assay, and Fluoro-Jade B staining. Immunohistochemistry was performed to evaluate cleaved caspase-3 expressions. Expression of BDNF, TrkB, GABAA, GAD65, Bad, Bcl-2, Bcl-xL, and Bax were determined by western blots. Results Significantly reduced seizure scores and mortality rates were observed with pretreatment with garcinol. Elevated expression of apoptotic proteins and caspase-3 in kindled mice were effectively downregulated by garcinol. Epileptogenic mice presented increased BDNF and TrkB with considerably decreased GABAA and GAD65 expression. Garcinol significantly enhanced GABAA and GAD65 while it suppressed BDNF and TrkB. Garcinol enhanced the performance of mice in Morris water maze tests. Conclusions Garcinol exerts neuroprotective effects via supressing apoptosis and modulating BDNF-TrkB signalling and GAD65/GABAA expressions and also enhanced cognition and memory of the mice. PMID:27855137

Grp94 Protein Delivers γ-Aminobutyric Acid Type A (GABAA) Receptors to Hrd1 Protein-mediated Endoplasmic Reticulum-associated Degradation.

PubMed

Di, Xiao-Jing; Wang, Ya-Juan; Han, Dong-Yun; Fu, Yan-Lin; Duerfeldt, Adam S; Blagg, Brian S J; Mu, Ting-Wei

2016-04-29

Proteostasis maintenance of γ-aminobutyric acid type A (GABAA) receptors dictates their function in controlling neuronal inhibition in mammalian central nervous systems. However, as a multisubunit, multispan, integral membrane protein, even wild type subunits of GABAA receptors fold and assemble inefficiently in the endoplasmic reticulum (ER). Unassembled and misfolded subunits undergo ER-associated degradation (ERAD), but this degradation process remains poorly understood for GABAA receptors. Here, using the α1 subunits of GABAA receptors as a model substrate, we demonstrated that Grp94, a metazoan-specific Hsp90 in the ER lumen, uses its middle domain to interact with the α1 subunits and positively regulates their ERAD. OS-9, an ER-resident lectin, acts downstream of Grp94 to further recognize misfolded α1 subunits in a glycan-dependent manner. This delivers misfolded α1 subunits to the Hrd1-mediated ubiquitination and the valosin-containing protein-mediated extraction pathway. Repressing the initial ERAD recognition step by inhibiting Grp94 enhances the functional surface expression of misfolding-prone α1(A322D) subunits, which causes autosomal dominant juvenile myoclonic epilepsy. This study clarifies a Grp94-mediated ERAD pathway for GABAA receptors, which provides a novel way to finely tune their function in physiological and pathophysiological conditions. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Evaluation of GABAergic neuroactive steroid 3alpha-hydroxy-5alpha-pregnane-20-one as a neurobiological substrate for the anti-anxiety effect of ethanol in rats.

PubMed

Hirani, Khemraj; Sharma, Ajay N; Jain, Nishant S; Ugale, Rajesh R; Chopde, Chandrabhan T

2005-07-01

Acute systemic ethanol administration is known to elevate plasma and cerebral levels of neuroactive steroid 3alpha-hydroxy-5alpha-pregnane-20-one (3alpha, 5alpha-THP; allopregnanolone) to a concentration sufficient to potentiate GABA(A) receptors. We have earlier demonstrated that 3alpha, 5alpha-THP mediates the antidepressant-like effect of ethanol in Porsolt forced swim test. The aim of the present study is to explain the relationship between endogenous GABAergic neurosteroids and anxiolytic effect of ethanol in Sprague-Dawley rats. The mediation of 3alpha, 5alpha-THP in the anti-anxiety effect of ethanol was assessed by pharmacological interactions of ethanol with various endogenous neurosteroidal modulators and using simulated physiological conditions of altered neurosteroid content in elevated plus maze (EPM) test. Pretreatment of 3alpha, 5alpha-THP (0.5-2.5 mug/rat, i.c.v.) or neurosteroidogenic agents such as 3alpha, 5alpha-THP precursor progesterone (5 or 10 mg/kg, i.p.), 11-beta hydroxylase inhibitor metyrapone (50 or 100 mg/kg, i.p.) or the GABA(A) receptor agonist muscimol (25 ng/rat, i.c.v.) significantly potentiated the anti-anxiety effect of ethanol (1 g/kg, i.p.). On the other hand, the GABAergic antagonistic neurosteroid dehydroepiandrosterone sulphate (DHEAS) (1 mg/kg, i.p.), the GABA(A) receptor blocker bicuculline (1 mg/kg, i.p.), the 5alpha-reductase inhibitor finasteride (50 x 2 mg/kg, s.c.) or the mitochondrial diazepam binding inhibitory receptor antagonist PK11195 (1 mg/kg, i.p.) reduced ethanol-induced preference of time spent and number of entries into open arms. Anti-anxiety effect of ethanol was abolished in adrenalectomized (ADX) rats as compared to sham-operated control. This ADX-induced blockade was restored by prior systemic injection of progesterone, signifying the contribution of peripheral steroidogenesis in ethanol anxiolysis. Socially isolated animals known to exhibit decreased brain 3alpha, 5alpha-THP and GABA(A) receptor

Agonist-induced modulation of inverse agonist efficacy at the beta 2-adrenergic receptor.

PubMed

Chidiac, P; Nouet, S; Bouvier, M

1996-09-01

Sustained stimulation of several G protein-coupled receptors is known to lead to a reduction in the signaling efficacy. This phenomenon, named agonist-induced desensitization, has been best studied for the beta 2-adrenergic receptor (AR) and is characterized by a decreased efficacy of beta-adrenergic agonists to stimulate the adenylyl cyclase activity. Recently, several beta-adrenergic ligands were found to inhibit the spontaneous agonist-independent activity of the beta 2AR. These compounds, termed inverse agonists, have different inhibitory efficacies, ranging from almost neutral antagonists to full inverse agonists. The current study was undertaken to determine whether, as is the case for agonists, desensitization can affect the efficacies of inverse agonists. Agonist-promoted desensitization of the human beta 2AR expressed in Sf9 cells potentiated the inhibitory actions of the inverse agonists, with the extent of the potentiation being inversely proportional to their intrinsic activity. For example, desensitization increased the inhibitory action of the weak inverse agonist labetalol by 29%, whereas inhibition of the spontaneous activity by the strong inverse agonist timolol was not enhanced by the desensitizing stimuli. Interestingly, dichloroisoproterenol acted stochastically as either a weak partial agonist or a weak inverse agonist in control conditions but always behaved as an inverse agonist after desensitization. These data demonstrate that like for agonists, the efficacies of inverse agonists can be modulated by a desensitizing treatment. Also, the data show that the initial state of the receptor can determine whether a ligand behaves as a partial agonist or an inverse agonist.

GABRA2 Alcohol Dependence Risk Allele is Associated with Reduced Expression of Chromosome 4p12 GABAA Subunit Genes in Human Neural Cultures.

PubMed

Lieberman, Richard; Kranzler, Henry R; Joshi, Pujan; Shin, Dong-Guk; Covault, Jonathan

2015-09-01

Genetic variation in a region of chromosome 4p12 that includes the GABAA subunit gene GABRA2 has been reproducibly associated with alcohol dependence (AD). However, the molecular mechanisms underlying the association are unknown. This study examined correlates of in vitro gene expression of the AD-associated GABRA2 rs279858*C-allele in human neural cells using an induced pluripotent stem cell (iPSC) model system. We examined mRNA expression of chromosome 4p12 GABAA subunit genes (GABRG1, GABRA2, GABRA4, and GABRB1) in 36 human neural cell lines differentiated from iPSCs using quantitative polymerase chain reaction and next-generation RNA sequencing. mRNA expression in adult human brain was examined using the BrainCloud and BRAINEAC data sets. We found significantly lower levels of GABRA2 mRNA in neural cell cultures derived from rs279858*C-allele carriers. Levels of GABRA2 RNA were correlated with those of the other 3 chromosome 4p12 GABAA genes, but not other neural genes. Cluster analysis based on the relative RNA levels of the 4 chromosome 4p12 GABAA genes identified 2 distinct clusters of cell lines, a low-expression cluster associated with rs279858*C-allele carriers and a high-expression cluster enriched for the rs279858*T/T genotype. In contrast, there was no association of genotype with chromosome 4p12 GABAA gene expression in postmortem adult cortex in either the BrainCloud or BRAINEAC data sets. AD-associated variation in GABRA2 is associated with differential expression of the entire cluster of GABAA subunit genes on chromosome 4p12 in human iPSC-derived neural cell cultures. The absence of a parallel effect in postmortem human adult brain samples suggests that AD-associated genotype effects on GABAA expression, although not present in mature cortex, could have effects on regulation of the chromosome 4p12 GABAA cluster during neural development. Copyright © 2015 by the Research Society on Alcoholism.

Aripiprazole Increases the PKA Signalling and Expression of the GABAA Receptor and CREB1 in the Nucleus Accumbens of Rats.

PubMed

Pan, Bo; Lian, Jiamei; Huang, Xu-Feng; Deng, Chao

2016-05-01

The GABAA receptor is implicated in the pathophysiology of schizophrenia and regulated by PKA signalling. Current antipsychotics bind with D2-like receptors, but not the GABAA receptor. The cAMP-responsive element-binding protein 1 (CREB1) is also associated with PKA signalling and may be related to the positive symptoms of schizophrenia. This study investigated the effects of antipsychotics in modulating D2-mediated PKA signalling and its downstream GABAA receptors and CREB1. Rats were treated orally with aripiprazole (0.75 mg/kg, ter in die (t.i.d.)), bifeprunox (0.8 mg/kg, t.i.d.), haloperidol (0.1 mg/kg, t.i.d.) or vehicle for 1 week. The levels of PKA-Cα and p-PKA in the prefrontal cortex (PFC), nucleus accumbens (NAc) and caudate putamen (CPu) were detected by Western blots. The mRNA levels of Gabrb1, Gabrb2, Gabrb3 and Creb1, and their protein expression were measured by qRT-PCR and Western blots, respectively. Aripiprazole elevated the levels of p-PKA and the ratio of p-PKA/PKA in the NAc, but not the PFC and CPu. Correlated with this elevated PKA signalling, aripiprazole elevated the mRNA and protein expression of the GABAA (β-1) receptor and CREB1 in the NAc. While haloperidol elevated the levels of p-PKA and the ratio of p-PKA/PKA in both NAc and CPu, it only tended to increase the expression of the GABAA (β-1) receptor and CREB1 in the NAc, but not the CPu. Bifeprunox had no effects on PKA signalling in these brain regions. These results suggest that aripiprazole has selective effects on upregulating the GABAA (β-1) receptor and CREB1 in the NAc, probably via activating PKA signalling.

Alteration of GABAergic synapses and gephyrin clusters in the thalamic reticular nucleus of GABAA receptor alpha3 subunit-null mice.

PubMed

Studer, Remo; von Boehmer, Lotta; Haenggi, Tatjana; Schweizer, Claude; Benke, Dietmar; Rudolph, Uwe; Fritschy, Jean-Marc

2006-09-01

Multiple GABAA-receptor subtypes are assembled from alpha, beta and gamma subunit variants. GABAA receptors containing the alpha3 subunit represent a minor population with a restricted distribution in the CNS. In addition, they predominate in monoaminergic neurons and in the nucleus reticularis thalami (nRT), suggesting a role in the regulation of cortical function and sleep. Mice with a targeted deletion of the alpha3 subunit gene (alpha3(0/0)) are viable and exhibit a subtle behavioural phenotype possibly related to dopaminergic hyperfunction. Here, we investigated immunohistochemically the consequences of the loss of alpha3 subunit for maturation of GABAA receptors and formation of GABAergic synapses in the nRT. Throughout postnatal development, the regional distribution of the alpha1, alpha2, or alpha5 subunit was unaltered in alpha3(0/0) mice and the prominent alpha3 subunit staining of nRT neurons in wildtype mice was not replaced. Subcellularly, as seen by double immunofluorescence, the alpha3 and gamma2 subunit were clustered at postsynaptic sites in the nRT of adult wildtype mice along with the scaffolding protein gephyrin. In alpha3(0/0) mice, gamma2 subunit clustering was disrupted and gephyrin formed large aggregates localized at the cell surface, but unrelated to postsynaptic sites, indicating that nRT neurons lack postsynaptic GABAA receptors in mutant mice. Furthermore, GABAergic terminals were enlarged and reduced in number, suggesting a partial deficit of GABAergic synapses. Therefore, GABAA receptors are required for gephyrin clustering and long-term synapse maintenance. The absence of GABAA-mediated transmission in the nRT may have a significant impact on the function of the thalamo-cortical loop of alpha3(0/0) mice.

Reduced GABAA Receptor α6 Expression in The Trigeminal Ganglion Enhanced Myofascial Nociceptive Response

PubMed Central

Kramer, P. R.; Bellinger, L. L.

2013-01-01

Activation of the GABAA receptor results in inhibition of neuronal activity. One subunit of this multi-subunit receptor termed alpha 6 (Gabrα6) contributed to inflammatory temporomandibular joint (TMJ) nociception but TMJ disorders often include myofascial pain. To address Gabrα6 role in myofascial pain we hypothesized that Gabrα6 has an inhibitory role in myofascial nociceptive responses similar to inflammatory TMJ arthritis. To test this hypothesis a, myofascial nociceptive response was induced by placing a ligature bilaterally on the tendon attachment of the anterior superficial part of a male rat's masseter muscle. Four days after ligature placement Gabrα6 expression was reduced by infusing the trigeminal ganglia (TG) with small interfering RNA (siRNA) having homology to either the Gabrα6 gene (Gabra6 siRNA) or no known gene (control siRNA). After siRNA infusion nociceptive behavioral responses were measured, i.e., feeding behavior and head withdrawal after pressing upon the region above the ligature with von Frey filaments. Neuronal activity in the TG and trigeminal nucleus caudalis and upper cervical region (Vc–C1) was measured by quantitating the amount of phosphorylated extracellular signalregulated kinase (p-ERK). Total Gabrα6 and GABAA receptor contents in the TG and Vc–C1 were determined. Gabrα6 siRNA infusion reduced Gabrα6 and GABAA receptor expression and significantly increased the nociceptive response in both nociceptive assays. Gabra6 siRNA infusion also significantly increased TG p-ERK expression of the ligated rats. From these results we conclude GABAA receptors consisting of the Gabrα6 subunit inhibit TG nociceptive sensory afferents in the trigeminal pathway and have an important role in the regulation of myofascial nociception. PMID:23602886

Differential Potency of 2,6-Dimethylcyclohexanol Isomers for Positive Modulation of GABAA Receptor Currents.

PubMed

Chowdhury, Luvana; Croft, Celine J; Goel, Shikha; Zaman, Naina; Tai, Angela C-S; Walch, Erin M; Smith, Kelly; Page, Alexandra; Shea, Kevin M; Hall, C Dennis; Jishkariani, D; Pillai, Girinath G; Hall, Adam C

2016-06-01

GABAA receptors meet all of the pharmacological requirements necessary to be considered important targets for the action of general anesthetic agents in the mammalian brain. In the following patch-clamp study, the relative modulatory effects of 2,6-dimethylcyclohexanol diastereomers were investigated on human GABAA (α1β3γ2s) receptor currents stably expressed in human embryonic kidney cells. Cis,cis-, trans,trans-, and cis,trans-isomers were isolated from commercially available 2,6-dimethylcyclohexanol and were tested for positive modulation of submaximal GABA responses. For example, the addition of 30 μM cis,cis-isomer resulted in an approximately 2- to 3-fold enhancement of the EC20 GABA current. Coapplications of 30 μM 2,6-dimethylcyclohexanol isomers produced a range of positive enhancements of control GABA responses with a rank order for positive modulation: cis,cis > trans,trans ≥ mixture of isomers > > cis,trans-isomer. In molecular modeling studies, the three cyclohexanol isomers bound with the highest binding energies to a pocket within transmembrane helices M1 and M2 of the β3 subunit through hydrogen-bonding interactions with a glutamine at the 224 position and a tyrosine at the 220 position. The energies for binding to and hydrogen-bond lengths within this pocket corresponded with the relative potencies of the agents for positive modulation of GABAA receptor currents (cis,cis > trans,trans > cis,trans-2,6-dimethylcyclohexanol). In conclusion, the stereochemical configuration within the dimethylcyclohexanols is an important molecular feature in conferring positive modulation of GABAA receptor activity and for binding to the receptor, a consideration that needs to be taken into account when designing novel anesthetics with enhanced therapeutic indices. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

2-Aminoethyl Methylphosphonate, a Potent and Rapidly Acting Antagonist of GABAA-ρ1 Receptors

PubMed Central

Xie, An; Yan, Jun; Yue, Lan; Feng, Feng; Mir, Fozia; Abdel-Halim, Heba; Chebib, Mary; Le Breton, Guy C.; Standaert, Robert F.; Qian, Haohua

2011-01-01

2-Aminoethyl methylphosphonate (2-AEMP), an analog of GABA, has been found to exhibit antagonist activity at GABAA-ρ1 (also known as ρ1 GABAC) receptors. The present study was undertaken to elucidate 2-AEMP's action and to test the activities of 2-AEMP analogs. Whole-cell patch-clamp techniques were used to record membrane currents in neuroblastoma cells stably transfected with human GABAA-ρ1 receptors. The action of 2-AEMP was compared with that of 1,2,5,6-tetrahydropyridin-4-yl methylphosphinic acid (TPMPA), a commonly used GABAA-ρ1 antagonist. With 10 μM GABA, 2-AEMP's IC50 (18 μM) differed by less than 2.5-fold from that of TPMPA (7 μM), and results obtained were consistent with a primarily competitive mode of inhibition by 2-AEMP. Terminating the presentation of 2-AEMP or TPMPA in the presence of GABA produced a release from inhibition. However, the rate of inhibition release upon the termination of 2-AEMP considerably exceeded that determined with termination of TPMPA. Moreover, when presented at concentrations near their respective IC50 values, the preincubation period associated with 2-AEMP's onset of inhibition was much shorter than that for TPMPA. Analogs of 2-AEMP possessing a benzyl or n-butyl rather than a methyl substituent at the phosphorus atom, as well as analogs bearing a C-methyl substituent on the aminoethyl side chain, exhibited reduced potency relative to 2-AEMP. Of these analogs, only (R)-2-aminopropyl methylphosphonate significantly diminished the response to 10 μM GABA. Structure-activity relationships are discussed in the context of molecular modeling of ligand binding to the antagonist binding site of the GABAA-ρ1 receptor. PMID:21810922

Effects of Chronic Ethanol Consumption on Rat GABAA and Strychnine-sensitive Glycine Receptors Expressed by Lateral/Basolateral Amygdala Neurons

PubMed Central

McCool, Brian A.; Frye, Gerald D.; Pulido, Marisa D.; Botting, Shaleen K.

2010-01-01

It is well known that the anxiolytic potential of ethanol is maintained during chronic exposure. We have confirmed this using a light-dark box paradigm following chronic ethanol ingestion via a liquid diet. However, cessation from chronic ethanol exposure is known to cause severe withdrawal anxiety. These opposing effects on anxiety likely result from neuro-adaptations of neurotransmitter systems within the brain regions regulating anxiety. Recent work highlights the importance of amygdala ligand-gated chloride channels in the expression of anxiety. We have therefore examined the effects of chronic ethanol exposure on GABAA and strychnine-sensitive glycine receptors expressed by acutely isolated adult rat lateral/basolateral amygdala neurons. Chronic ethanol exposure increased the functional expression of GABAA receptors in acutely isolated basolateral amygdala neurons without altering strychnine-sensitive glycine receptors. Neither the acute ethanol nor benzodiazepine sensitivity of either receptor system was affected. We explored the likelihood that subunit composition might influence each receptor’s response to chronic ethanol. Importantly, when expressed in a mammalian heterologous system, GABAA receptors composed of unique α subunits were differentially sensitive to acute ethanol. Likewise, the presence of the β subunit appeared to influence the acute ethanol sensitivity of glycine receptors containing the α2 subunit. Our results suggest that the facilitation of GABAA receptors during chronic ethanol exposure may help explain the maintenance of ethanol’s anti-anxiety effects during chronic ethanol exposure. Furthermore, the subunit composition of GABAA and strychnine-sensitive glycine receptors may ultimately influence the response of each system to chronic ethanol exposure. PMID:12560122

β2 Agonists.

PubMed

Billington, Charlotte K; Penn, Raymond B; Hall, Ian P

2017-01-01

History suggests β agonists, the cognate ligand of the β 2 adrenoceptor, have been used as bronchodilators for around 5,000 years, and β agonists remain today the frontline treatment for asthma and chronic obstructive pulmonary disease (COPD). The β agonists used clinically today are the products of significant expenditure and over 100 year's intensive research aimed at minimizing side effects and enhancing therapeutic usefulness. The respiratory physician now has a therapeutic toolbox of long acting β agonists to prophylactically manage bronchoconstriction, and short acting β agonists to relieve acute exacerbations. Despite constituting the cornerstone of asthma and COPD therapy, these drugs are not perfect; significant safety issues have led to a black box warning advising that long acting β agonists should not be used alone in patients with asthma. In addition there are a significant proportion of patients whose asthma remains uncontrolled. In this chapter we discuss the evolution of β agonist use and how the understanding of β agonist actions on their principal target tissue, airway smooth muscle, has led to greater understanding of how these drugs can be further modified and improved in the future. Research into the genetics of the β 2 adrenoceptor will also be discussed, as will the implications of individual DNA profiles on the clinical outcomes of β agonist use (pharmacogenetics). Finally we comment on what the future may hold for the use of β agonists in respiratory disease.

K+ channel TASK-1 knockout mice show enhanced sensitivities to ataxic and hypnotic effects of GABA(A) receptor ligands.

PubMed

Linden, Anni-Maija; Aller, M Isabel; Leppä, Elli; Rosenberg, Per H; Wisden, William; Korpi, Esa R

2008-10-01

TASK two-pore-domain leak K(+) channels occur throughout the brain. However, TASK-1 and TASK-3 knockout (KO) mice have few neurological impairments and only mildly reduced sensitivities to inhalational anesthetics, contrasting with the anticipated functions and importance of these channels. TASK-1/-3 channel expression can compensate for the absence of GABA(A) receptors in GABA(A) alpha6 KO mice. To investigate the converse, we analyzed the behavior of TASK-1 and -3 KO mice after administering drugs with preferential efficacies at GABA(A) receptor subtypes: benzodiazepines (diazepam and flurazepam, active at alpha1betagamma2, alpha2betagamma2, alpha3betagamma2, and alpha5betagamma2 subtypes), zolpidem (alpha1betagamma2 subtype), propofol (beta2-3-containing receptors), gaboxadol (alpha4betadelta and alpha6betadelta subtypes), pregnanolone, and pentobarbital (many subtypes). TASK-1 KO mice showed increased motor impairment in rotarod and beam-walking tests after diazepam and flurazepam administration but not after zolpidem. They also showed prolonged loss of righting reflex induced by propofol and pentobarbital. Autoradiography indicated no change in GABA(A) receptor ligand binding levels. These altered behavioral responses to GABAergic drugs suggest functional up-regulation of alpha2beta2/3gamma2 and alpha3beta2/3gamma2 receptor subtypes in TASK-1 KO mice. In addition, female, but not male, TASK-1 KO mice were more sensitive to gaboxadol, suggesting an increased influence of alpha4betadelta or alpha6betadelta subtypes. The benzodiazepine sensitivity of TASK-3 KO mice was marginally increased. Our results underline that TASK-1 channels perform such key functions in the brain that compensation is needed for their absence. Furthermore, because inhalation anesthetics act partially through GABA(A) receptors, the up-regulation of GABA(A) receptor function in TASK-1 KO mice might mask TASK-1 channel's significance as a target for inhalation anesthetics.

Increased GABA-A receptor binding and reduced connectivity at the motor cortex in children with hemiplegic cerebral palsy: a multimodal investigation using 18F-fluoroflumazenil PET, immunohistochemistry, and MR imaging.

PubMed

Park, Hae-Jeong; Kim, Chul Hoon; Park, Eun Sook; Park, Bumhee; Oh, So Ra; Oh, Maeng-Keun; Park, Chang Il; Lee, Jong Doo

2013-08-01

γ-aminobutyric acid (GABA)-A receptor-mediated neural transmission is important to promote practice-dependent plasticity after brain injury. This study investigated alterations in GABA-A receptor binding and functional and anatomic connectivity within the motor cortex in children with cerebral palsy (CP). We conducted (18)F-fluoroflumazenil PET on children with hemiplegic CP to investigate whether in vivo GABA-A receptor binding is altered in the ipsilateral or contralateral hemisphere of the lesion site. To evaluate changes in the GABA-A receptor subunit after prenatal brain injury, we performed GABA-A receptor immunohistochemistry using rat pups with a diffuse hypoxic ischemic insult. We also performed diffusion tensor MR imaging and resting-state functional MR imaging on the same children with hemiplegic CP to investigate alterations in anatomic and functional connectivity at the motor cortex with increased GABA-A receptor binding. In children with hemiplegic CP, the (18)F-fluoroflumazenil binding potential was increased within the ipsilateral motor cortex. GABA-A receptors with the α1 subunit were highly expressed exclusively within cortical layers III, IV, and VI of the motor cortex in rat pups. The motor cortex with increased GABA-A receptor binding in children with hemiplegic CP had reduced thalamocortical and corticocortical connectivity, which might be linked to increased GABA-A receptor distribution in cortical layers in rats. Increased expression of the GABA-A receptor α1 subunit within the ipsilateral motor cortex may be an important adaptive mechanism after prenatal brain injury in children with CP but may be associated with improper functional connectivity after birth and have adverse effects on the development of motor plasticity.

Driving Human Motor Cortical Oscillations Leads to Behaviorally Relevant Changes in Local GABAA Inhibition: A tACS-TMS Study

PubMed Central

van Ede, Freek

2017-01-01

Beta and gamma oscillations are the dominant oscillatory activity in the human motor cortex (M1). However, their physiological basis and precise functional significance remain poorly understood. Here, we used transcranial magnetic stimulation (TMS) to examine the physiological basis and behavioral relevance of driving beta and gamma oscillatory activity in the human M1 using transcranial alternating current stimulation (tACS). tACS was applied using a sham-controlled crossover design at individualized intensity for 20 min and TMS was performed at rest (before, during, and after tACS) and during movement preparation (before and after tACS). We demonstrated that driving gamma frequency oscillations using tACS led to a significant, duration-dependent decrease in local resting-state GABAA inhibition, as quantified by short interval intracortical inhibition. The magnitude of this effect was positively correlated with the magnitude of GABAA decrease during movement preparation, when gamma activity in motor circuitry is known to increase. In addition, gamma tACS-induced change in GABAA inhibition was closely related to performance in a motor learning task such that subjects who demonstrated a greater increase in GABAA inhibition also showed faster short-term learning. The findings presented here contribute to our understanding of the neurophysiological basis of motor rhythms and suggest that tACS may have similar physiological effects to endogenously driven local oscillatory activity. Moreover, the ability to modulate local interneuronal circuits by tACS in a behaviorally relevant manner provides a basis for tACS as a putative therapeutic intervention. SIGNIFICANCE STATEMENT Gamma oscillations have a vital role in motor control. Using a combined tACS-TMS approach, we demonstrate that driving gamma frequency oscillations modulates GABAA inhibition in the human motor cortex. Moreover, there is a clear relationship between the change in magnitude of GABAA inhibition induced

Enhancement of GABA release through endogenous activation of axonal GABA(A) receptors in juvenile cerebellum.

PubMed

Trigo, Federico F; Chat, Mireille; Marty, Alain

2007-11-14

Recent evidence indicates the presence of presynaptic GABA(A) receptors (GABA(A)Rs) in the axon domain of several classes of central neurons, including cerebellar basket and stellate cells. Here, we investigate the possibility that these receptors could be activated in the absence of electrical or chemical stimulation. We find that low concentrations of GABA increase the frequency of miniature GABAergic synaptic currents. Submaximal concentrations of a GABA(A)R blocker, gabazine, decrease both the miniature current frequency and the probability of evoked GABA release. Zolpidem, an agonist of the benzodiazepine binding site, and NO-711 (1-[2-[[(diphenylmethylene)imino]oxy]ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride), a blocker of GABA uptake, both increase the frequency of miniature currents. These effects occur up to postnatal day 14, but not later. Immunohistochemistry indicates the presence of alpha1-containing GABA(A)Rs in interneuron presynaptic terminals with a similar age dependence. We conclude that, under resting conditions, axonal GABA(A)Rs are significantly activated, that this activation results in enhanced GABA release, and that it can be augmented by increasing the affinity of GABA(A)Rs or reducing GABA uptake. Our findings suggest the existence of a positive-feedback mechanism involving presynaptic GABA(A)Rs that maintains a high release rate and a high local GABA concentration in the immature cerebellar network.

The Memory-Impairing Effects of Septal GABA Receptor Activation Involve GABAergic Septo-Hippocampal Projection Neurons

ERIC Educational Resources Information Center

Krebs-Kraft, Desiree L.; Wheeler, Marina G.; Parent, Marise B.

2007-01-01

Septal infusions of the [gamma]-aminobutyric acid (GABA)[subscript A] agonist muscimol impair memory, and the effect likely involves the hippocampus. GABA[subscript A] receptors are present on the perikarya of cholinergic and GABAergic septo-hippocampal (SH) projections. The current experiments determined whether GABAergic SH projections are…

General, kappa, delta and mu opioid receptor antagonists mediate feeding elicited by the GABA-B agonist baclofen in the ventral tegmental area and nucleus accumbens shell in rats: reciprocal and regional interactions.

PubMed

Miner, Patricia; Shimonova, Lyudmila; Khaimov, Arthur; Borukhova, Yaffa; Ilyayeva, Ester; Ranaldi, Robert; Bodnar, Richard J

2012-03-14

Food intake is significantly increased following administration of agonists of GABA and opioid receptors into the nucleus accumbens shell (NACs) and ventral tegmental area (VTA). GABA-A or GABA-B receptor antagonist pretreatment within the VTA or NACs differentially affects mu-opioid agonist-induced feeding elicited from the same site. Correspondingly, general or selective opioid receptor antagonist pretreatment within the VTA or NACs differentially affects GABA agonist-induced feeding elicited from the same site. Regional interactions have been evaluated in feeding studies by administering antagonists in one site prior to agonist administration in a second site. Thus, opioid antagonist-opioid agonist and GABA antagonist-GABA agonist feeding interactions have been identified between the VTA and NACs. However, pretreatment with GABA-A or GABA-B receptor antagonists in the VTA failed to affect mu opioid agonist-induced feeding elicited from the NACs, and correspondingly, these antagonists administered in the NACs failed to affect mu opioid-induced feeding elicited from the VTA. To evaluate whether regional and reciprocal VTA and NACs feeding interactions occur for opioid receptor modulation of GABA agonist-mediated feeding, the present study examined whether feeding elicited by the GABA-B agonist, baclofen microinjected into the NACs was dose-dependently blocked by pretreatment with general (naltrexone: NTX), mu (beta-funaltrexamine: BFNA), kappa (nor-binaltorphamine: NBNI) or delta (naltrindole: NTI) opioid antagonists in the VTA, and correspondingly, whether VTA baclofen-induced feeding was dose-dependently blocked by NACs pretreatment with NTX, BFNA, NBNI or NTI in rats. Bilateral pairs of cannulae aimed at the VTA and NACs were stereotaxically implanted in rats, and their food intakes were assessed following vehicle and baclofen (200 ng) in each site. Baclofen produced similar magnitudes of increased food intake following VTA and NACs treatment. Baclofen

GABAa excitation and synaptogenesis after Status Epilepticus - A computational study.

PubMed

França, Keite Lira de Almeida; de Almeida, Antônio-Carlos Guimarães; Saddow, Stephen E; Santos, Luiz Eduardo Canton; Scorza, Carla Alessandra; Scorza, Fulvio Alexandre; Rodrigues, Antônio Márcio

2018-03-08

The role of GABAergic neurotransmission on epileptogenesis has been the subject of speculation according to different approaches. However, it is a very complex task to specifically consider the action of the GABAa neurotransmitter, which, in its dependence on the intracellular level of Cl - , can change its effect from inhibitory to excitatory. We have developed a computational model that represents the dentate gyrus and is composed of three different populations of neurons (granule cells, interneurons and mossy cells) that are mutually interconnected. The interconnections of the neurons were based on compensation theory with Hebbian and anti-Hebbian rules. The model also incorporates non-synaptic mechanisms to control the ionic homeostasis and was able to reproduce ictal discharges. The goal of the work was to investigate the hypothesis that the observed aberrant sprouting is promoted by GABAa excitatory action. Conjointly with the abnormal sprouting of the mossy fibres, the simulations show a reduction of the mossy cells connections in the network and an increased inhibition of the interneurons as a response of the neuronal network to control the activity. This finding contributes to increasing the changes in the connectivity of the neuronal circuitry and to increasing the epileptiform activity occurrences.

Effects of chronic ethanol consumption on rat GABA(A) and strychnine-sensitive glycine receptors expressed by lateral/basolateral amygdala neurons.

PubMed

McCool, Brian A; Frye, Gerald D; Pulido, Marisa D; Botting, Shaleen K

2003-02-14

It is well known that the anxiolytic potential of ethanol is maintained during chronic exposure. We have confirmed this using a light-dark box paradigm following chronic ethanol ingestion via a liquid diet. However, cessation from chronic ethanol exposure is known to cause severe withdrawal anxiety. These opposing effects on anxiety likely result from neuro-adaptations of neurotransmitter systems within the brain regions regulating anxiety. Recent work highlights the importance of amygdala ligand-gated chloride channels in the expression of anxiety. We have therefore examined the effects of chronic ethanol exposure on GABA(A) and strychnine-sensitive glycine receptors expressed by acutely isolated adult rat lateral/basolateral amygdala neurons. Chronic ethanol exposure increased the functional expression of GABA(A) receptors in acutely isolated basolateral amygdala neurons without altering strychnine-sensitive glycine receptors. Neither the acute ethanol nor benzodiazepine sensitivity of either receptor system was affected. We explored the likelihood that subunit composition might influence each receptor's response to chronic ethanol. Importantly, when expressed in a mammalian heterologous system, GABA(A) receptors composed of unique alpha subunits were differentially sensitive to acute ethanol. Likewise, the presence of the beta subunit appeared to influence the acute ethanol sensitivity of glycine receptors containing the alpha(2) subunit. Our results suggest that the facilitation of GABA(A) receptors during chronic ethanol exposure may help explain the maintenance of ethanol's anti-anxiety effects during chronic ethanol exposure. Furthermore, the subunit composition of GABA(A) and strychnine-sensitive glycine receptors may ultimately influence the response of each system to chronic ethanol exposure.

A Novel Bifunctional Alkylphenol Anesthetic Allows Characterization of γ-Aminobutyric Acid, Type A (GABAA), Receptor Subunit Binding Selectivity in Synaptosomes.

PubMed

Woll, Kellie A; Murlidaran, Sruthi; Pinch, Benika J; Hénin, Jérôme; Wang, Xiaoshi; Salari, Reza; Covarrubias, Manuel; Dailey, William P; Brannigan, Grace; Garcia, Benjamin A; Eckenhoff, Roderic G

2016-09-23

Propofol, an intravenous anesthetic, is a positive modulator of the GABAA receptor, but the mechanistic details, including the relevant binding sites and alternative targets, remain disputed. Here we undertook an in-depth study of alkylphenol-based anesthetic binding to synaptic membranes. We designed, synthesized, and characterized a chemically active alkylphenol anesthetic (2-((prop-2-yn-1-yloxy)methyl)-5-(3-(trifluoromethyl)-3H-diazirin-3-yl)phenol, AziPm-click (1)), for affinity-based protein profiling (ABPP) of propofol-binding proteins in their native state within mouse synaptosomes. The ABPP strategy captured ∼4% of the synaptosomal proteome, including the unbiased capture of five α or β GABAA receptor subunits. Lack of γ2 subunit capture was not due to low abundance. Consistent with this, independent molecular dynamics simulations with alchemical free energy perturbation calculations predicted selective propofol binding to interfacial sites, with higher affinities for α/β than γ-containing interfaces. The simulations indicated hydrogen bonding is a key component leading to propofol-selective binding within GABAA receptor subunit interfaces, with stable hydrogen bonds observed between propofol and α/β cavity residues but not γ cavity residues. We confirmed this by introducing a hydrogen bond-null propofol analogue as a protecting ligand for targeted-ABPP and observed a lack of GABAA receptor subunit protection. This investigation demonstrates striking interfacial GABAA receptor subunit selectivity in the native milieu, suggesting that asymmetric occupancy of heteropentameric ion channels by alkylphenol-based anesthetics is sufficient to induce modulation of activity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

A Novel Bifunctional Alkylphenol Anesthetic Allows Characterization of γ-Aminobutyric Acid, Type A (GABAA), Receptor Subunit Binding Selectivity in Synaptosomes*

PubMed Central

Woll, Kellie A.; Murlidaran, Sruthi; Pinch, Benika J.; Hénin, Jérôme; Wang, Xiaoshi; Salari, Reza; Covarrubias, Manuel; Dailey, William P.; Brannigan, Grace; Garcia, Benjamin A.; Eckenhoff, Roderic G.

2016-01-01

Propofol, an intravenous anesthetic, is a positive modulator of the GABAA receptor, but the mechanistic details, including the relevant binding sites and alternative targets, remain disputed. Here we undertook an in-depth study of alkylphenol-based anesthetic binding to synaptic membranes. We designed, synthesized, and characterized a chemically active alkylphenol anesthetic (2-((prop-2-yn-1-yloxy)methyl)-5-(3-(trifluoromethyl)-3H-diazirin-3-yl)phenol, AziPm-click (1)), for affinity-based protein profiling (ABPP) of propofol-binding proteins in their native state within mouse synaptosomes. The ABPP strategy captured ∼4% of the synaptosomal proteome, including the unbiased capture of five α or β GABAA receptor subunits. Lack of γ2 subunit capture was not due to low abundance. Consistent with this, independent molecular dynamics simulations with alchemical free energy perturbation calculations predicted selective propofol binding to interfacial sites, with higher affinities for α/β than γ-containing interfaces. The simulations indicated hydrogen bonding is a key component leading to propofol-selective binding within GABAA receptor subunit interfaces, with stable hydrogen bonds observed between propofol and α/β cavity residues but not γ cavity residues. We confirmed this by introducing a hydrogen bond-null propofol analogue as a protecting ligand for targeted-ABPP and observed a lack of GABAA receptor subunit protection. This investigation demonstrates striking interfacial GABAA receptor subunit selectivity in the native milieu, suggesting that asymmetric occupancy of heteropentameric ion channels by alkylphenol-based anesthetics is sufficient to induce modulation of activity. PMID:27462076

Responses to GABA(A) receptor activation are altered in NTS neurons isolated from renal-wrap hypertensive rats.

PubMed

Tolstykh, Gleb; Belugin, Sergei; Tolstykh, Olga; Mifflin, Steve

2003-10-01

The inhibitory amino acid GABA is a potent modulator of the spontaneous discharge and the responses to afferent inputs of neurons in the nucleus of the solitary tract (NTS). To determine if responses to activation of GABA(A) receptors are altered in hypertension, GABA(A) receptor-evoked whole cell currents were measured in enzymatically dispersed NTS neurons from 33 normotensive (NT, 109+/-4 mm Hg, n=7) and 24 hypertensive (HT, 167+/-5 mm Hg, n=24) rats. GABA(A) receptor-evoked currents reversed at the calculated equilibrium potential for chloride and were blocked by bicuculline (n=6). Membrane capacitance was the same in neurons from NT (7.5+/-0.6 pF, n=62) and HT (6.8+/-0.6 pF, n=51) rats. The EC50 for peak GABA-evoked currents cells was significantly greater in neurons from HT (21.0+/-2.6 micromol/L, n=16) compared with NT rats (13.0+/-1.8 micromol/L, n=14, P=0.01). The EC50 of neurons exhibiting DiA labeling of presumptive aortic nerve terminals was no different than that observed in the nonlabeled cells (19.0+/-4.9 micromol/L, n=4). The time constant for desensitization of GABA(A)-evoked currents was the same in neurons from HT (4.5+/-0.3 seconds, n=17) and NT rats (3.8+/-0.3 seconds, n=17, P>0.05). Repetitive pulse application of GABA revealed a more rapid decline in the evoked current in neurons from HT compared with NT rats. The amplitude of the 5th pulse of GABA (5-second duration, 2-second interval) was 21+/-2% the amplitude of the 1st pulse in NT rats (n=10) and 14+/-2% in HT rats (n=11, P<0.05). These alterations in GABAA-receptor evoked currents could render the neurons less sensitive to GABA(A) receptor inhibition and influence afferent integration by NTS neurons in HT.

Decrement of GABAA receptor-mediated inhibitory postsynaptic currents in dentate granule cells in epileptic hippocampus.

PubMed

Isokawa, M

1996-05-01

1. Inhibitory postsynaptic currents (IPSCs) were studied in hippocampal dentate granule cells (DGCs) in the pilocarpine model and human temporal lobe epilepsy, with the use of the whole cell patch-clamp recording technique in slice preparations. 2. In the pilocarpine model, hippocampal slices were prepared from rats that were allowed to experience spontaneous seizures for 2 mo. Human hippocampal specimens were obtained from epileptic patients who underwent surgical treatment for medically intractable seizures. 3. IPSCs were generated by single perforant path stimulation and recorded at a membrane potential (Vm) of 0 mV near the reversal potential of glutamate excitatory postsynaptic currents in the voltage-clamp recording. IPSCs were pharmacologically identified as gamma-aminobutyric acid-A (GABAA) IPSCs by 10 microM bicuculline methiodide. 4. During low-frequency stimulation, IPSCs were not different in amplitude among non-seizure-experienced rat hippocampi, human nonsclerotic hippocampi, seizure-experienced rat hippocampi, and human sclerotic hippocampi. In the last two groups of DGCs, current-clamp recordings indicated the presence of prolonged excitatory postsynaptic potentials (EPSPs) mediated by the N-methyl-D-aspartate (NMDA) receptor. 5. High-frequency stimulation, administered at Vm = -30 mV to activate NMDA currents, reduced GABAA IPSC amplitude specifically in seizure-experienced rat hippocampi (t = 2.5, P < 0.03) and human sclerotic hippocampi (t = 7.7, P < 0.01). This reduction was blocked by an NMDA receptor antagonist, 2-amino-5-phosphonovaleric acid (APV) (50 microM). The time for GABAA IPSCs to recover to their original amplitude was also shortened by the application of APV. 6. I conclude that, when intensively activated, NMDA receptor-mediated excitatory transmission may interact with GABAergic synaptic inhibition in DGCs in seizure-experienced hippocampus to transiently reduce GABA(A) receptor-channel function. Such interactions may contribute to

No association of the GABAA receptor genes on chromosome 5 with alcoholism in the collaborative study on the genetics of alcoholism sample.

PubMed

Dick, Danielle M; Edenberg, Howard J; Xuei, Xiaoling; Goate, Alison; Hesselbrock, Victor; Schuckit, Marc; Crowe, Raymond; Foroud, Tatiana

2005-01-05

A substantial body of literature suggests that gamma-aminobutyric acid (GABA) may be involved in the neurochemical pathways contributing to alcohol use and related disorders. Chromosome 5 contains a cluster of GABA(A) receptor genes, GABRA1, GABRA6, GABRB2, and GABRG2, which have been among the most extensively studied in relation to alcohol use. These studies have yielded mixed results. Using data from large, multiplex alcoholic families collected as part of the Collaborative Study on the Genetics of Alcoholism (COGA), we sought to provide more conclusive evidence regarding the role of the GABA(A) receptor genes on chromosome 5. Multiple single nucleotide polymorphisms (SNPs) were tested in each of the four chromosome 5q GABA(A) receptor genes, and we conducted both classic trio-based association analyzes and extended pedigree analyzes. We found no consistent evidence of association with alcohol dependence or alcohol dependence comorbid with antisocial personality disorder (ASPD) for any of the regions tested in the chromosome 5 GABA(A) receptor genes. These analyses suggest that the GABA(A) receptor genes on chromosome 5 do not play a strong role in alcohol dependence. Future studies are planned to test whether these genes are more important in influencing behavioral endophenotypes related to the risk of alcohol dependence. Copyright 2004 Wiley-Liss, Inc.

Involvement of NMDA receptor in low-frequency magnetic field-induced anxiety in mice.

PubMed

Salunke, Balwant P; Umathe, Sudhir N; Chavan, Jagatpalsingh G

2014-12-01

It had been reported that exposure to extremely low-frequency magnetic field (ELFMF) induces anxiety in human and rodents. Anxiety mediates via the activation of N-methyl-d-aspartate (NMDA) receptor, whereas activation of γ-aminobutyric acid (GABA) receptor attenuates the same. Hence, the present study was carried out to understand the contribution of NMDA and/or GABA receptors modulation in ELFMF-induced anxiety for which Swiss albino mice were exposed to ELFMF (50 Hz, 10 G) by subjecting them to Helmholtz coils. The exposure was for 8 h/day for 7, 30, 60, 90 and 120 days. Anxiety level was assessed in elevated plus maze, open field test and social interaction test, on 7th, 30th, 60th, 90th and 120th exposure day, respectively. Moreover, the role of GABA and glutamate in ELFMF-induced anxiety was assessed by treating mice with muscimol [0.25 mg/kg intraperitoneally (i.p.)], bicuculline (1.0 mg/kg i.p.), NMDA (15 mg/kg i.p.) and MK-801 (0.03 mg/kg i.p.), as a GABAA and NMDA receptor agonist and antagonist, respectively. Glutamate receptor agonist exacerbated while inhibitor attenuated the ELFMF-induced anxiety. In addition, levels of GABA and glutamate were determined in regions of the brain viz, cortex, striatum, hippocampus and hypothalamus. Experiments demonstrated significant elevation of GABA and glutamate levels in the hippocampus and hypothalamus. However, GABA receptor modulators did not produce significant effect on ELFMF-induced anxiety and elevated levels of GABA at tested dose. Together, these findings suggest that ELFMF significantly induced anxiety behavior, and indicated the involvement of NMDA receptor in its effect.

Interaction of H+ and Zn2+ on recombinant and native rat neuronal GABAA receptors

PubMed Central

Krishek, Belinda J; Moss, Stephen J; Smart, Trevor G

1998-01-01

The interaction of Zn2+ and H+ ions with GABAA receptors was examined using Xenopus laevis oocytes expressing recombinant GABAA receptors composed of subunits selected from α1, β1, γ2S and δ types, and by using cultured rat cerebellar granule neurones. The potency of Zn2+ as a non-competitive antagonist of GABA-activated responses on α1β1 receptors was reduced by lowering the external pH from 7.4 to 5.4, increasing the Zn2+ IC50 value from 1.2 to 58.3 μm. Zinc-induced inhibition was largely unaffected by alkaline pH up to pH 9.4. For α1β1δ subunits, concentration-response curves for GABA were displaced laterally by Zn2+ in accordance with a novel mixed/competitive-type inhibition. The Zn2+ IC50 at pH 7.4 was 16.3 μm. Acidification of Ringer solution resulted in a reduced antagonism by Zn2+ (IC50, 49.0 μm) without affecting the type of inhibition. At pH 9.4, Zn2+ inhibition remained unaffected. The addition of the γ2S subunit to the α1β1δ construct caused a marked reduction in the potency of Zn2+ (IC50, 615 μm), comparable to that observed with α1β1γ2S receptors (IC50 639 μm). GABA concentration-response curves were depressed in a mixed/non-competitive fashion. In cultured cerebellar granule neurones, Zn2+ inhibited responses to GABA in a concentration-dependent manner. Lowering external pH from 7.4 to 6.4 increased the IC50 from 139 to 253 μm. The type of inhibition exhibited by Zn2+ on cerebellar granule neurones, previously grown in high K+-containing culture media, was complex, with the GABA concentration-response curves shifting laterally with reduced slopes and similar maxima. The Zn2+-induced shift in the GABA EC50 values was reduced by lowering the external pH from 7.4 to 6.4. The interaction of H+ and Zn2+ ions on GABAA receptors suggests that they share either a common regulatory pathway or coincident binding sites on the receptor protein. The apparent competitive mode of block induced by Zn2+ on α1β1δ receptors is shared by GABAA

Multiple Non-Equivalent Interfaces Mediate Direct Activation of GABAA Receptors by Propofol.

PubMed

Eaton, Megan M; Germann, Allison L; Arora, Ruby; Cao, Lily Q; Gao, Xiaoyi; Shin, Daniel J; Wu, Albert; Chiara, David C; Cohen, Jonathan B; Steinbach, Joe Henry; Evers, Alex S; Akk, Gustav

2016-01-01

Propofol is a sedative agent that at clinical concentrations acts by allosterically activating or potentiating the γ-aminobutyric acid type A (GABAA) receptor. Mutational, modeling, and photolabeling studies with propofol and its analogues have identified potential interaction sites in the transmembrane domain of the receptor. At the &quot;+&quot; of the β subunit, in the β-α interface, meta-azipropofol labels the M286 residue in the third transmembrane domain. Substitution of this residue with tryptophan results in loss of potentiation by propofol. At the &quot;-&quot; side of the β subunit, in the α-β interface (or β-β interface, in the case of homomeric β receptors), ortho-propofol diazirine labels the H267 residue in the second transmembrane domain. Structural modeling indicates that the β(H267) residue lines a cavity that docks propofol with favorable interaction energy. We used two-electrode voltage clamp to determine the functional effects of mutations to the "+" and "-" sides of the β subunit on activation of the α1β3 GABAA receptor by propofol. We found that while the individual mutations had a small effect, the combination of the M286W mutation with tryptophan mutations of selected residues at the α-β interface leads to strong reduction in gating efficacy for propofol. We conclude that α1β3 GABAA receptors can be activated by propofol interactions with the β-β, α-β, and β-α interfaces, where distinct, non-equivalent regions control channel gating. Any interface can mediate activation, hence substitutions at all interfaces are required for loss of activation by propofol.

Multiple Non-Equivalent Interfaces Mediate Direct Activation of GABAA Receptors by Propofol

PubMed Central

Eaton, Megan M.; Germann, Allison L.; Arora, Ruby; Cao, Lily Q.; Gao, Xiaoyi; Shin, Daniel J.; Wu, Albert; Chiara, David C.; Cohen, Jonathan B.; Steinbach, Joe Henry; Evers, Alex S.; Akk, Gustav

2016-01-01

Abstract: Background Propofol is a sedative agent that at clinical concentrations acts by allosterically activating or potentiating the γ-aminobutyric acid type A (GABAA) receptor. Mutational, modeling, and photolabeling studies with propofol and its analogues have identified potential interaction sites in the transmembrane domain of the receptor. At the “+” of the β subunit, in the β-α interface, meta-azipropofol labels the M286 residue in the third transmembrane domain. Substitution of this residue with tryptophan results in loss of potentiation by propofol. At the “-” side of the β subunit, in the α-β interface (or β-β interface, in the case of homomeric β receptors), ortho-propofol diazirine labels the H267 residue in the second transmembrane domain. Structural modeling indicates that the β(H267) residue lines a cavity that docks propofol with favorable interaction energy. Method We used two-electrode voltage clamp to determine the functional effects of mutations to the 
“+” and “-” sides of the β subunit on activation of the α1β3 GABAA receptor by propofol. Results We found that while the individual mutations had a small effect, the combination of the M286W mutation with tryptophan mutations of selected residues at the α-β interface leads to strong reduction in gating efficacy for propofol. Conclusion We conclude that α1β3 GABAA receptors can be activated by propofol interactions with the β-β, α-β, and β-α interfaces, where distinct, non-equivalent regions control channel gating. Any interface can mediate activation, hence substitutions at all interfaces are required for loss of activation by propofol. PMID:26830963

BENZODIAZEPINE-INDUCED SPATIAL LEARNING DEFICITS IN RATS ARE REGULATED BY THE DEGREE OF MODULATION OF α1 GABAA RECEPTORS

PubMed Central

Joksimović, Srđan; Divljaković, Jovana; Van Linn, Michael L.; Varagic, Zdravko; Brajković, Gordana; Milinković, Marija M.; Yin, Wenyuan; Timić, Tamara; Sieghart, Werner; Cook, James M.; Savić, Miroslav M.

2012-01-01

Despite significant advances in understanding the role of benzodiazepine (BZ)-sensitive populations of GABAA receptors, containing the α1, α2, α3 or α5 subunit, factual substrates of BZ-induced learning and memory deficits are not yet fully elucidated. It was shown that α1-subunit affinity-selective antagonist β-CCt almost completely abolished spatial learning deficits induced by diazepam (DZP) in the Morris water maze. We examined a novel, highly (105 fold) α1-subunit selective ligand - WYS8 (0.2, 1 and 10 mg/kg), on its own and in combination with the non-selective agonist DZP (2 mg/kg) or β-CCt (5 mg/kg) in the water maze in rats. The in vitro efficacy study revealed that WYS8 acts as α1-subtype selective weak partial positive modulator (40% potentiation at 100 nM). Measurement of concentrations of WYS8 and DZP in rat serum and brain tissues suggested that they did not substantially cross-influence the respective disposition. In the water maze, DZP impaired spatial learning (acquisition trials) and memory (probe trial). WYS8 caused no effect per se, did not affect the overall influence of DZP on the water-maze performance and was devoid of any activity in this task when combined with β-CCt. Nonetheless, an additional analysis of the latency to reach the platform and the total distance swam suggested that WYS8 addition attenuated the run-down of the spatial impairment induced by DZP at the end of acquisition trials. These results demonstrate a clear difference in the influence of an α1 subtype-selective antagonist and a partial agonist on the effects of DZP on the water-maze acquisition. PMID:22633616

The natural products magnolol and honokiol are positive allosteric modulators of both synaptic and extra-synaptic GABAA receptors

PubMed Central

Alexeev, Mikhail; Grosenbaugh, Denise K.; Mott, David D.; Fisher, Janet L.

2012-01-01

The National Center for Complementary and Alternative Medicine (NCCAM) estimates that nearly 40% of adults in the United States use alternative medicines, often in the form of an herbal supplement. Extracts from the tree bark of magnolia species have been used for centuries in traditional Chinese and Japanese medicines to treat a variety of neurological diseases, including anxiety, depression, and seizures. The active ingredients in the extracts have been identified as the bi-phenolic isomers magnolol and honokiol. These compounds were shown to enhance the activity of GABAA receptors, consistent with their biological effects. The GABAA receptors exhibit substantial subunit heterogeneity, which influences both their functional and pharmacological properties. We examined the activity of magnolol and honokiol at different populations of both neuronal and recombinant GABAA receptors to characterize their mechanism of action and to determine whether sensitivity to modulation was dependent upon the receptor’s subunit composition. We found that magnolol and honokiol enhanced both phasic and tonic GABAergic neurotransmission in hippocampal dentate granule neurons. In addition, all recombinant receptors examined were sensitive to modulation, regardless of the identity of the α, β, or γ subunit subtype, although the compounds showed particularly high efficacy at δ-containing receptors. This direct positive modulation of both synaptic and extra-synaptic populations of GABAA receptors suggests that supplements containing magnolol and/or honokiol would be effective anxiolytics, sedatives, and anti-convulsants. However, significant side-effects and risk of drug interactions would also be expected. PMID:22445602

Changes on auditory physiology in response to the inactivation of amygdala nuclei in high anxiety rats expressing learned fear.

PubMed

Nobre, Manoel Jorge

2013-06-13

The inferior colliculus (IC) is primarily involved in the processing of acoustic stimuli, including those emitted by prey and predators. The role of the central nucleus of the IC (CIC) in fear and anxiety has been suggested based on electrophysiological, behavioral and immunohistochemical studies. The reactivity of high-anxiety rats (HA) to diverse challenges is different from low-anxiety ones (LA). In humans and laboratory animals, pathological anxiety is often accompanied by heightened vigilance and alertness, hyperactivity of the amygdala (AM), and increased amplitude of the auditory evoked potentials (AEP) from the IC. This study aims to evaluate the influence of the inactivation of the central (CEA) and basolateral (BLA) nuclei of the amygdala, after local infusions of the full GABAA agonist muscimol (1nmol/0.2μl), on the AEP elicited in the CIC of rats tested under a learned fear state. Our results showed that both BLA and CEA inactivation change the expression of conditioned fear, in a paradigm using the context as the conditioned stimulus (CS). These changes are correlated to the innate anxiety levels of the animals. It is supposed that this shortcoming is in addition to the imbalance between the regulatory role of the top-down and bottom-up processes in the control of anxiety. Copyright © 2013 Elsevier Inc. All rights reserved.

Angiotensin II and CRF receptors in the central nucleus of the amygdala mediate hemodynamic response variability to cocaine in conscious rats.

PubMed

Watanabe, Mari A; Kucenas, Sarah; Bowman, Tamara A; Ruhlman, Melissa; Knuepfer, Mark M

2010-01-14

Stress or cocaine evokes either a large increase in systemic vascular resistance (SVR) or a smaller increase in SVR accompanied by an increase in cardiac output (designated vascular and mixed responders, respectively) in Sprague-Dawley rats. We hypothesized that the central nucleus of the amygdala (CeA) mediates this variability. Conscious, freely-moving rats, instrumented for measurement of arterial pressure and cardiac output and for drug delivery into the CeA, were given cocaine (5 mg/kg, iv, 4-6 times) and characterized as vascular (n=15) or mixed responders (n=10). Subsequently, we administered cocaine after bilateral microinjections (100 nl) of saline or selective agents in the CeA. Muscimol (80 pmol), a GABA(A) agonist, or losartan (43.4 pmol), an AT(1) receptor antagonist, attenuated the cocaine-induced increase in SVR in vascular responders, selectively, such that vascular responders were no longer different from mixed responders. The corticotropin releasing factor (CRF) antagonist, alpha-helical CRF(9-41) (15.7 pmol), abolished the difference between cardiac output and SVR in mixed and vascular responders. We conclude that greater increases in SVR observed in vascular responders are dependent on AT(1) receptor activation and, to a lesser extent on CRF receptors. Therefore, AT(1) and CRF receptors in the CeA contribute to hemodynamic response variability to intravenous cocaine.

Angiotensin II and CRF Receptors in the Central Nucleus of the Amygdala Mediate Hemodynamic Response Variability to Cocaine in Conscious Rats

PubMed Central

Watanabe, Mari A.; Kucenas, Sarah; Bowman, Tamara A.; Ruhlman, Melissa; Knuepfer, Mark M.

2009-01-01

Stress or cocaine evokes either a large increase in systemic vascular resistance (SVR) or a smaller increase in SVR accompanied by an increase in cardiac output (designated vascular and mixed responders, respectively) in Sprague-Dawley rats. We hypothesized that the central nucleus of the amygdala (CeA) mediates this variability. Conscious, freely-moving rats, instrumented for measurement of arterial pressure and cardiac output and for drug delivery into the CeA, were given cocaine (5 mg/kg, iv, 4-6 times) and characterized as vascular (n=15) or mixed responders (n=10). Subsequently, we administered cocaine after bilateral microinjections (100 nl) of saline or selective agents in the CeA. Muscimol (80 pmol), a GABAA agonist, or losartan (43.4 pmol), an AT1 receptor antagonist, attenuated the cocaine-induced increase in SVR in vascular responders, selectively, such that vascular responders were no longer different from mixed responders. The corticotropin releasing factor (CRF) antagonist, α-helical CRF9-41 (15.7 pmol), abolished the difference between cardiac output and SVR in mixed and vascular responders. We conclude that greater increases in SVR observed in vascular responders are dependent on AT1 receptor activation and, to a lesser extent on CRF receptors. Therefore, AT1 and CRF receptors in the CeA contribute to hemodynamic response variability to intravenous cocaine. PMID:19879859

Posterior insular cortex is necessary for conditioned inhibition of fear

PubMed Central

Foilb, Allison R.; Flyer-Adams, Johanna G.; Maier, Steven F.; Christianson, John P.

2016-01-01

Veridical detection of safety versus danger is critical to survival. Learned signals for safety inhibit fear, and so when presented, reduce fear responses produced by danger signals. This phenomenon is termed conditioned inhibition of fear. Here, we report that CS+/CS− fear discrimination conditioning over 5 days in rats leads the CS− to become a conditioned inhibitor of fear, as measured by the classic tests of conditioned inhibition: summation and retardation of subsequent fear acquisition. We then show that NMDA-receptor antagonist AP5 injected to posterior insular cortex (IC) before training completely prevented the acquisition of a conditioned fear inhibitor, while intra-AP5 to anterior and medial IC had no effect. To determine if the IC contributes to the recall of learned fear inhibition, injections of the GABAA agonist muscimol were made to posterior IC before a summation test. This resulted in fear inhibition per se, which obscured inference to the effect of IC inactivation with recall of the safety cue. Control experiments sought to determine if the role of the IC in conditioned inhibition learning could be reduced to simpler fear discrimination function, but fear discrimination and recall were unaffected by AP5 or muscmiol, respectively, in the posterior IC. These data implicate a role of posterior IC in the learning of conditioned fear inhibitors. PMID:27523750

Residual effect of a 7-amino metabolite of clonazepam on GABAA receptor function in the nucleus reticularis thalami of the rat.

PubMed

Munakata, Mitsutoshi; Tsuchiya, Shigeru

2008-10-01

A considerable amount of 7-aminoclonazepam (ACZP), a major metabolite of clonazepam (CZP), is present in the brain during CZP treatment, yet the pharmacological properties of ACZP remain unknown. We investigated the effects of ACZP on the GABA(A) receptor-mediated currents (I(GABA)) in neurons from the nucleus reticularis thalami (NRT) of the rat, using a nystatin-perforated patch technique. Neurons in which CZP (10 nM) exerted prominent augmentation (>100% augmentation) of I(GABA), which comprised 32% of the neurons tested, were included for the analysis of ACZP. In these neurons, ACZP augmented I(GABA), which was blocked by 10 microM flumazenil, a benzodiazepine receptor (BZR) antagonist. The half-maximal effective concentration of ACZP was 124 nM, whereas that of CZP was 1.8 nM. The maximal enhancements induced by ACZP and CZP were 38% and 170%, respectively. In neurons from the ventrobasal complex of the thalamus, the effect of ACZP was negligible. Our results suggest that ACZP was a weak partial BZR agonist and that ACZP may competitively modify the effect of CZP, leading to clinical consequences for patients with high levels of ACZP.

Lateral Orbitofrontal Cortical Modulation on the Medial Prefrontal Cortex-Amygdala Pathway: Differential Regulation of Intra-Amygdala GABAA and GABAB Receptors.

PubMed

Chang, Chun-Hui

2017-07-01

The basolateral complex of the amygdala receives inputs from neocortical areas, including the medial prefrontal cortex and lateral orbitofrontal cortex. Earlier studies have shown that lateral orbitofrontal cortex activation exerts an inhibitory gating on medial prefrontal cortex-amygdala information flow. Here we examined the individual role of GABAA and GABAB receptors in this process. In vivo extracellular single-unit recordings were done in anesthetized rats. We searched amygdala neurons that fire in response to medial prefrontal cortex activation, tested lateral orbitofrontal cortex gating at different delays (lateral orbitofrontal cortex-medial prefrontal cortex delays: 25, 50, 100, 250, 500, and 1000 milliseconds), and examined differential contribution of GABAA and GABAB receptors with iontophoresis. Relative to baseline, lateral orbitofrontal cortex stimulation exerted an inhibitory modulatory gating on the medial prefrontal cortex-amygdala pathway and was effective up to a long delay of 500 ms (long-delay latencies at 100, 250, and 500 milliseconds). Moreover, blockade of intra-amygdala GABAA receptors with bicuculline abolished the lateral orbitofrontal cortex inhibitory gating at both short- (25 milliseconds) and long-delay (100 milliseconds) intervals, while blockade of GABAB receptors with saclofen reversed the inhibitory gating at long delay (100 milliseconds) only. Among the majority of the neurons examined (8 of 9), inactivation of either GABAA or GABAB receptors during baseline did not change evoked probability per se, suggesting that local feed-forward inhibitory mechanism is pathway specific. Our results suggest that the effect of lateral orbitofrontal cortex inhibitory modulatory gating was effective up to 500 milliseconds and that intra-amygdala GABAA and GABAB receptors differentially modulate the short- and long-delay lateral orbitofrontal cortex inhibitory gating on the medial prefrontal cortex-amygdala pathway. © The Author 2017

Glycosylation of β2 Subunits Regulates GABAA Receptor Biogenesis and Channel Gating*

PubMed Central

Lo, Wen-yi; Lagrange, Andre H.; Hernandez, Ciria C.; Harrison, Rebecca; Dell, Anne; Haslam, Stuart M.; Sheehan, Jonathan H.; Macdonald, Robert L.

2010-01-01

γ-Aminobutyric acid type A (GABAA) receptors are heteropentameric glycoproteins. Based on consensus sequences, the GABAA receptor β2 subunit contains three potential N-linked glycosylation sites, Asn-32, Asn-104, and Asn-173. Homology modeling indicates that Asn-32 and Asn-104 are located before the α1 helix and in loop L3, respectively, near the top of the subunit-subunit interface on the minus side, and that Asn-173 is located in the Cys-loop near the bottom of the subunit N-terminal domain. Using site-directed mutagenesis, we demonstrated that all predicted β2 subunit glycosylation sites were glycosylated in transfected HEK293T cells. Glycosylation of each site, however, produced specific changes in α1β2 receptor surface expression and function. Although glycosylation of Asn-173 in the Cys-loop was important for stability of β2 subunits when expressed alone, results obtained with flow cytometry, brefeldin A treatment, and endo-β-N-acetylglucosaminidase H digestion suggested that glycosylation of Asn-104 was required for efficient α1β2 receptor assembly and/or stability in the endoplasmic reticulum. Patch clamp recording revealed that mutation of each site to prevent glycosylation decreased peak α1β2 receptor current amplitudes and altered the gating properties of α1β2 receptor channels by reducing mean open time due to a reduction in the proportion of long open states. In addition to functional heterogeneity, endo-β-N-acetylglucosaminidase H digestion and glycomic profiling revealed that surface β2 subunit N-glycans at Asn-173 were high mannose forms that were different from those of Asn-32 and N104. Using a homology model of the pentameric extracellular domain of α1β2 channel, we propose mechanisms for regulation of GABAA receptors by glycosylation. PMID:20639197

GABA-A Receptor Modulation and Anticonvulsant, Anxiolytic, and Antidepressant Activities of Constituents from Artemisia indica Linn

PubMed Central

Khan, Imran; Karim, Nasiara; Ahmad, Waqar; Abdelhalim, Abeer; Chebib, Mary

2016-01-01

Artemisia indica, also known as “Mugwort,” has been widely used in traditional medicines. However, few studies have investigated the effects of nonvolatile components of Artemisia indica on central nervous system's function. Fractionation of Artemisia indica led to the isolation of carnosol, ursolic acid, and oleanolic acid which were evaluated for their effects on GABA-A receptors in electrophysiological studies in Xenopus oocytes and were subsequently investigated in mouse models of acute toxicity, convulsions (pentylenetetrazole induced seizures), depression (tail suspension and forced swim tests), and anxiety (elevated plus maze and light/dark box paradigms). Carnosol, ursolic acid, and oleanolic acid were found to be positive modulators of α1β2γ2L GABA-A receptors and the modulation was antagonized by flumazenil. Carnosol, ursolic acid, and oleanolic acid were found to be devoid of any signs of acute toxicity (50–200 mg/kg) but elicited anticonvulsant, antidepressant, and anxiolytic activities. Thus carnosol, ursolic acid, and oleanolic acid demonstrated CNS activity in mouse models of anticonvulsant, antidepressant, and anxiolysis. The anxiolytic activity of all three compounds was ameliorated by flumazenil suggesting a mode of action via the benzodiazepine binding site of GABA-A receptors. PMID:27143980

Acute orexigenic effect of agmatine involves interaction between central α2-adrenergic and GABAergic receptors.

PubMed

Taksande, Brijesh Gulabrao; Sharma, Omi; Aglawe, Manish Manohar; Kale, Mayur Bhimrao; Gawande, Dinesh Yugraj; Umekar, Milind Janraoji; Kotagale, Nandkishor Ramdas

2017-09-01

Agmatine and GABA have been abundantly expressed in brain nuclei involved in regulation of energy homeostasis and promoting stimulation of food intake in rodents. However, their mutual interaction, if any, in the elicitation of feeding behavior is largely remains unclear. The current study provides experimental evidence for the possible interaction of agmatine, adrenergic and GABAergic systems in stimulation of feeding in satiated rats. Satiated rats fitted with intracerebroventricular (i.c.v.) cannulae and were administered agmatine, alone or jointly with (a) GABA A receptor agonist, muscimol, diazepam or antagonist bicuculline and flumazenil, GABA A positive modulator, allopregnanolone or negative modulator of GABA A receptor, dehydroepiandrosterone (b) In view of the high affinity of agmatine for α 2 -adrenoceptors and the close association between α 2 -adrenoceptors and GABAergic system, the effect of their modulators on feeding elicited by agmatine/GABAergic agonists were also examined. I.c.v. administration of agmatine (40-80μg/rat) induces the significant orexigenic effect in satiated rats. The orexigenic effect of agmatine was potentiated by muscimol (25ng/rat, i.c.v.); diazepam (0.5mg/kg, i.p.); allopregnanolone (0.5mg/kg, s.c.) and blocked by bicuculline (1mg/kg, i.p.) and dehydroepiandrosterone (4mg/kg,s.c.). However, it remained unaffected in presence of flumazenil (25ng/rat, i.c.v.). The orexigenic effect of agmatine and GABAergic agonists was potentiated by a α 2 -adrenoceptors agonist, clonidine (10ng/rat, i.c.v.) and blocked by its antagonist, yohimbine (5μg/rat, i.c.v.). Yohimbine also blocked the hyperphagic effect elicited by ineffective dose combination of agmatine (5μg/rat, i.c.v.) with muscimol (25ng/rat, i.c.v.) or diazepam (0.5mg/kg, i.p.) or allopregnanolone (0.5mg/kg,s.c.). The results of the present study suggest that agmatine induced α 2 -adrenoceptors activation might facilitate GABAergic activity to stimulate food intake in

HPLC-based activity profiling for GABAA receptor modulators from the traditional Chinese herbal drug Kushen (Sophora flavescens root)

PubMed Central

2011-01-01

An EtOAc extract from the roots of Sophora flavescens (Kushen) potentiated γ -aminobutyric acid (GABA)-induced chloride influx in Xenopus oocytes transiently expressing GABAA receptors with subunit composition, α1β2γ2S. HPLC-based activity profiling of the extract led to the identification of 8-lavandulyl flavonoids, kushenol I, sophoraflavanone G, (–)-kurarinone, and kuraridine as GABAA receptor modulators. In addition, a series of inactive structurally related flavonoids were characterized. Among these, kushenol Y (4) was identified as a new natural product. The 8-lavandulyl flavonoids are first representatives of a novel scaffold for the target. PMID:21207144

Pharmacological evaluation of the anxiolytic-like effects of EMD 386088, a partial 5-HT6 receptor agonist, in the rat elevated plus-maze and Vogel conflict tests.

PubMed

Jastrzębska-Więsek, Magdalena; Siwek, Agata; Partyka, Anna; Kubacka, Monika; Mogilski, Szczepan; Wasik, Anna; Kołaczkowski, Marcin; Wesołowska, Anna

2014-10-01

The 5-HT6 is one of the most recent additions to the 5-HT receptor family. Its pharmacological profile and anatomical distribution is suggestive of a putative role in mood disorders. Most of preclinical evidence suggests an anxiolytic-like action of 5-HT6 receptor antagonists. Evaluation the anxiolytic-like effects of EMD 386088, a partial 5-HT6receptor agonist, and its putative mechanism of action in rats. EMD 386088, administered intraperitoneally at a dose of 2.5 mg/kg evoked specific anxiolytic-like activity in the automated version of the conflict drinking Vogel and the elevated plus-maze tests visible by increasing all parameters indicating a potential anti-anxiety effect. Its activity was blocked by the selective 5-HT6 receptor antagonist SB 271046, but not by the selective GABAA/benzodiazepine receptor antagonist flumazenil. EMD 386088 did not intensify an anxiolytic-like effect produced by diazepam in the elevated plus-maze test. These findings suggest that EMD 386088, a 5-HT6 receptor agonist, produces anxiolytic-like activity after systemic administration which may result from direct stimulation of 5-HT6 receptors. Copyright © 2014 Elsevier Ltd. All rights reserved.

Estrous cycle variations in GABAA receptor phosphorylation enable rapid modulation by anabolic androgenic steroids in the medial preoptic area

PubMed Central

Oberlander, JG; Porter, DM; Onakomaiya, MM; Penatti, CAA; Vithlani, M; Moss, SJ; Clark, AS; Henderson, LP

2012-01-01

Anabolic androgenic steroids (AAS), synthetic testosterone derivatives that are used for ergogenic purposes, alter neurotransmission and behaviors mediated by GABAA receptors. Some of these effects may reflect direct and rapid action of these synthetic steroids at the receptor. The ability of other natural allosteric steroid modulators to alter GABAA receptor-mediated currents is dependent upon the phosphorylation state of the receptor complex. Here we show that phosphorylation of the GABAA receptor complex immunoprecipitated by β2/β3 subunit-specific antibodies from the medial preoptic area (mPOA) of the mouse varies across the estrous cycle; with levels being significantly lower in estrus. Acute exposure to the AAS, 17α-testosterone (17α-MeT), had no effect on the amplitude or kinetics of inhibitory postsynaptic currents in the mPOA of estrous mice when phosphorylation was low, but increased the amplitude of these currents from mice in diestrus, when it was high. Inclusion of the protein kinase C (PKC) inhibitor, calphostin, in the recording pipette eliminated the ability of 17α-MeT to enhance currents from diestrous animals, suggesting that PKC-receptor phosphorylation is critical for the allosteric modulation elicited by AAS during this phase. In addition, a single injection of 17α-MeT was found to impair an mPOA-mediated behavior (nest-building) in diestrus, but not in estrus. PKC is known to target specific serine residues in the β3 subunit of the GABAA receptor. Although phosphorylation of these β3 serine residues showed a similar profile across the cycle, as did phosphoserine in mPOA lysates immunoprecipitated with β2/β3 antibody (lower in estrus than in diestrus or proestrus), the differences were not significant. These data suggest that the phosphorylation state of the receptor complex regulates both the ability of AAS to modulate receptor function in the mPOA and the expression of a simple mPOA-dependent behavior through PKC-dependent mechanism

Possible effects of depolarizing GABAA conductance on the neuronal input-output relationship: a modeling study.

PubMed

Morita, Kenji; Tsumoto, Kunichika; Aihara, Kazuyuki

2005-06-01

Recent in vitro experiments revealed that the GABAA reversal potential is about 10 mV higher than the resting potential in mature mammalian neocortical pyramidal cells; thus GABAergic inputs could have facilitatory, rather than inhibitory, effects on action potential generation under certain conditions. However, how the relationship between excitatory input conductances and the output firing rate is modulated by such depolarizing GABAergic inputs under in vivo circumstances has not yet been understood. We examine herewith the input-output relationship in a simple conductance-based model of cortical neurons with the depolarized GABAA reversal potential, and show that a tonic depolarizing GABAergic conductance up to a certain amount does not change the relationship between a tonic glutamatergic driving conductance and the output firing rate, whereas a higher GABAergic conductance prevents spike generation. When the tonic glutamatergic and GABAergic conductances are replaced by in vivo-like highly fluctuating inputs, on the other hand, the effect of depolarizing GABAergic inputs on the input-output relationship critically depends on the degree of coincidence between glutamatergic input events and GABAergic ones. Although a wide range of depolarizing GABAergic inputs hardly changes the firing rate of a neuron driven by noncoincident glutamatergic inputs, a certain range of these inputs considerably decreases the firing rate if a large number of driving glutamatergic inputs are coincident with them. These results raise the possibility that the depolarized GABAA reversal potential is not a paradoxical mystery, but is instead a sophisticated device for discriminative firing rate modulation.

Removal of GABAA Receptor γ2 Subunits from Parvalbumin Neurons Causes Wide-Ranging Behavioral Alterations

PubMed Central

Leppä, Elli; Linden, Anni-Maija; Vekovischeva, Olga Y.; Swinny, Jerome D.; Rantanen, Ville; Toppila, Esko; Höger, Harald; Sieghart, Werner; Wulff, Peer; Wisden, William; Korpi, Esa R.

2011-01-01

We investigated the behavioral significance of fast synaptic inhibition by αβγ2-type GABAA receptors on parvalbumin (Pv) cells. The GABAA receptor γ2 subunit gene was selectively inactivated in Pv-positive neurons by Cre/loxP recombination. The resulting Pv-Δγ2 mice were relatively healthy in the first postnatal weeks; but then as Cre started to be expressed, the mice progressively developed wide-ranging phenotypic alterations including low body weight, motor deficits and tremor, decreased anxiety levels, decreased pain sensitivity and deficient prepulse inhibition of the acoustic startle reflex and impaired spatial learning. Nevertheless, the deletion was not lethal, and mice did not show increased mortality even after one year. Autoradiography with t-butylbicyclophosphoro[35S]thionate suggested an increased amount of GABAA receptors with only α and β subunits in central nervous system regions that contained high levels of parvalbumin neurons. Using BAC-transgenesis, we reduced some of the Pv-Δγ2 phenotype by selectively re-expressing the wild-type γ2 subunit back into some Pv cells (reticular thalamic neurons and cerebellar Pv-positive neurons). This produced less severe impairments of motor skills and spatial learning compared with Pv-Δγ2 mice, but all other deficits remained. Our results reveal the widespread significance of fast GABAergic inhibition onto Pv-positive neurons for diverse behavioral modalities, such as motor coordination, sensorimotor integration, emotional behavior and nociception. PMID:21912668

GABAergic control of neostriatal dopamine D2 receptor binding and behaviors in the rat.

PubMed

Nikolaus, Susanne; Beu, Markus; de Souza Silva, Maria Angelica; Huston, Joseph P; Antke, Christina; Müller, Hans-Wilhelm; Hautzel, Hubertus

2017-02-01

The present study assessed the influence of the GABA A receptor agonist muscimol and the GABA A receptor antagonist bicuculline on neostriatal dopamine D 2 receptor binding in relation to motor and exploratory behaviors in the rat. D 2 receptor binding was measured in baseline and after challenge with either 1mg/kg muscimol or 1mg/kg bicuculline. In additional rats, D 2 receptor binding was measured after injection of saline. After treatment with muscimol, bicuculline and saline, motor and exploratory behaviors were assessed for 30min in an open field prior to administration of [ 123 I]S-3-iodo-N-(1-ethyl-2-pyrrolidinyl)methyl-2-hydroxy-6-methoxybenzamide ([ 123 I]IBZM). For baseline and challenges, striatal equilibrium ratios (V 3 ″) were computed as estimation of the binding potential. Muscimol but not bicuculline reduced D 2 receptor binding relative to baseline and to saline. Travelled distance, duration of rearing and frequency of rearing and of head-shoulder motility were lower after muscimol compared to saline. In contrast, duration of rearing and grooming and frequency of rearing, head-shoulder motility and grooming were elevated after bicuculline relative to saline. Moreover, bicuculline decreased duration of sitting and head-shoulder motility. The muscimol-induced decrease of motor/exploratory behaviors can be related to an elevation of striatal dopamine levels. In contrast, bicuculline is likely to elicit a decline of synaptic dopamine, which, however, is compensated by the time of D 2 receptor imaging studies. The results indicate direct GABAergic control over D 2 receptor binding in the neostriatum in relation to behavioral action, and, thus, complement earlier pharmacological studies. Copyright © 2016. Published by Elsevier Inc.

The orthosteric GABAA receptor ligand Thio-4-PIOL displays distinctly different functional properties at synaptic and extrasynaptic receptors

PubMed Central

Hoestgaard-Jensen, K; O'Connor, R M; Dalby, N O; Simonsen, C; Finger, B C; Golubeva, A; Hammer, H; Bergmann, M L; Kristiansen, U; Krogsgaard-Larsen, P; Bräuner-Osborne, H; Ebert, B; Frølund, B; Cryan, J F; Jensen, A A

2013-01-01

BACKGROUND AND PURPOSE Explorations into the heterogeneous population of native GABA type A receptors (GABAARs) and the physiological functions governed by the multiple GABAAR subtypes have for decades been hampered by the lack of subtype-selective ligands. EXPERIMENTAL APPROACH The functional properties of the orthosteric GABAA receptor ligand 5-(4-piperidyl)-3-isothiazolol (Thio-4-PIOL) have been investigated in vitro, ex vivo and in vivo. KEY RESULTS Thio-4-PIOL displayed substantial partial agonist activity at the human extrasynaptic GABAAR subtypes expressed in Xenopus oocytes, eliciting maximal responses of up to ∼30% of that of GABA at α5β3γ2S, α4β3δ and α6β3δ and somewhat lower efficacies at the corresponding α5β2γ2S, α4β2δ and α6β2δ subtypes (maximal responses of 4–12%). In contrast, it was an extremely low efficacious agonist at the α1β3γ2S, α1β2γ2S, α2β2γ2S, α2β3γ2S, α3β2γ2S and α3β3γ2S GABAARs (maximal responses of 0–4%). In concordance with its agonism at extrasynaptic GABAARs and its de facto antagonism at the synaptic receptors, Thio-4-PIOL elicited robust tonic currents in electrophysiological recordings on slices from rat CA1 hippocampus and ventrobasal thalamus and antagonized phasic currents in hippocampal neurons. Finally, the observed effects of Thio-4-PIOL in rat tests of anxiety, locomotion, nociception and spatial memory were overall in good agreement with its in vitro and ex vivo properties. CONCLUSION AND IMPLICATIONS The diverse signalling characteristics of Thio-4-PIOL at GABAARs represent one of the few examples of a functionally subtype-selective orthosteric GABAAR ligand reported to date. We propose that Thio-4-PIOL could be a useful pharmacological tool in future studies exploring the physiological roles of native synaptic and extrasynaptic GABAARs. PMID:23957253

Activity in Prelimbic Cortex Subserves Fear Memory Reconsolidation over Time

ERIC Educational Resources Information Center

Stern, Cristina A. J.; Gazarini, Lucas; Vanvossen, Ana C.; Hames, Mayara S.; Bertoglio, Leandro J.

2014-01-01

The prelimbic cortex has been implicated in the consolidation of previously learned fear. Herein, we report that temporarily inactivating this medial prefrontal cortex subregion with the GABA [subscript A] agonist muscimol (4.0 nmol in 0.2 µL per hemisphere) was able to equally disrupt 1-, 7-, and 21-d-old contextual fear memories after their…

Regulation of /sup 3/H-dopamine release by presynaptic GABA and glutamate heteroreceptors in rat brain nucleus accumbens synaptosomes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kovalev, G.I.; Hetey, L.

1987-06-01

The aim of this investigation was a neurochemical study of the effect of agonists of different types of GABA receptors - muscimol (type A receptor), baclofen (type B receptor), delta-aminolevulinic acid (DALA; GABA autoreceptor), and also of GABA itself - on tritium-labelled dopamine release, stimulated by potassium cations, from synaptosomes of the nuclei accumbenes of the rat brain.

Weaker control of the electrical properties of cerebellar granule cells by tonically active GABAA receptors in the Ts65Dn mouse model of Down’s syndrome

PubMed Central

2013-01-01

Background Down’s syndrome (DS) is caused by triplication of all or part of human chromosome 21 and is characterized by a decrease in the overall size of the brain. One of the brain regions most affected is the cerebellum, in which the number of granule cells (GCs) is markedly decreased. GCs process sensory information entering the cerebellum via mossy fibres and pass it on to Purkinje cells and inhibitory interneurons. How GCs transform incoming signals depends on their input–output relationship, which is adjusted by tonically active GABAA receptor channels. Results We report that in the Ts65Dn mouse model of DS, in which cerebellar volume and GC number are decreased as in DS, the tonic GABAA receptor current in GCs is smaller than in wild-type mice and is less effective in moderating input resistance and raising the minimum current required for action potential firing. We also find that tonically active GABAA receptors curb the height and broaden the width of action potentials in wild-type GCs but not in Ts65Dn GCs. Single-cell real-time quantitative PCR reveals that these electrical differences are accompanied by decreased expression of the gene encoding the GABAA receptor β3 subunit but not genes coding for some of the other GABAA receptor subunits expressed in GCs (α1, α6, β2 and δ). Conclusions Weaker moderation of excitability and action potential waveform in GCs of the Ts65Dn mouse by tonically active GABAA receptors is likely to contribute to atypical transfer of information through the cerebellum. Similar changes may occur in DS. PMID:23870245

An examination of the effects of subthalamic nucleus inhibition or μ-opioid receptor stimulation on food-directed motivation in the non-deprived rat

PubMed Central

Pratt, Wayne E.; Choi, Eugene; Guy, Elizabeth G.

2012-01-01

The subthalamic nucleus (STN) serves important functions in regulating movement, cognition, and motivation and is connected with cortical and basal ganglia circuits that process reward and reinforcement. In order to further examine the role of the STN on motivation toward food in non-deprived rats, these experiments studied the effects of pharmacological inhibition or μ-opioid receptor stimulation of the STN on the 2-hr intake of a sweetened fat diet, the amount of work exerted to earn sucrose on a progressive ratio 2 (PR-2) schedule of reinforcement, and performance on a differential reinforcement of low-rate responding (DRL) schedule for sucrose reward. Separate behavioral groups (N = 6–9) were tested following bilateral inhibition of the STN with the GABAA receptor agonist muscimol (at 0–5 ng/0.5 μl/side) or following μ-opioid receptor stimulation with the agonist D-Ala2, N-MePhe4, Gly-ol-enkephalin (DAMGO; at 0, 0.025 or 0.25 μg/0.5 μl/side). Although STN inhibition increased ambulatory behavior during 2-hr feeding sessions, it did not significantly alter intake of the sweetened fat diet. STN inhibition also did not affect the breakpoint for sucrose pellets during a 1-hr PR-2 reinforcement schedule or impact the number of reinforcers earned on a 1-hr DRL-20 sec reinforcement schedule in non-deprived rats. In contrast, STN μ-opioid receptor stimulation significantly increased feeding on the palatable diet and reduced the reinforcers earned on a DRL-20 schedule, although DAMGO microinfusions had no effect on PR-2 performance. These data suggest that STN inhibition does not enhance incentive motivation for food in the absence of food restriction and that STN μ-opioid receptors play an important and unique role in motivational processes. PMID:22391117

GABAB-receptor activation alters the firing pattern of dopamine neurons in the rat substantia nigra.

PubMed

Engberg, G; Kling-Petersen, T; Nissbrandt, H

1993-11-01

Previous electrophysiological experiments have emphasized the importance of the firing pattern for the functioning of midbrain dopamine (DA) neurons. In this regard, excitatory amino acid receptors appear to constitute an important modulatory control mechanism. In the present study, extracellular recording techniques were used to investigate the significance of GABAB-receptor activation for the firing properties of DA neurons in the substantia nigra (SN) in the rat. Intravenous administration of the GABAB-receptor agonist baclofen (1-16 mg/kg) was associated with a dose-dependent regularization of the firing pattern, concomitant with a reduction in burst firing. At higher doses (16-32 mg/kg), the firing rate of the DA neurons was dose-dependently decreased. Also, microiontophoretic application of baclofen regularized the firing pattern of nigral DA neurons, including a reduction of burst firing. Both the regularization of the firing pattern and inhibition of firing rate produced by systemic baclofen administration was antagonized by the GABAB-receptor antagonist CGP 35348 (200 mg/kg, i.v.). The GABAA-receptor agonist muscimol produced effects on the firing properties of DA neurons that were opposite to those observed following baclofen, i.e., an increase in firing rate accompanied by a decreased regularity. The NMDA receptor antagonist MK 801 (0.4-3.2 mg/kg, i.v.) produced a moderate, dose-dependent increase in the firing rate of the nigral DA neurons as well as a slightly regularized firing pattern. Pretreatment with MK 801 (3.2 mg/kg, i.v., 3-10 min) did neither promote nor prevent the regularization of the firing pattern or inhibition of firing rate on the nigral DA neurons produced by baclofen. The present results clearly show that GABAB-receptors can alter the firing pattern of nigral DA neurons, hereby counterbalancing the previously described ability of glutamate to induce burst firing activity on these neurons.

Relationship between a GABAA alpha 6 Pro385Ser substitution and benzodiazepine sensitivity.

PubMed

Iwata, N; Cowley, D S; Radel, M; Roy-Byrne, P P; Goldman, D

1999-09-01

In humans, interindividual variation in sensitivity to benzodiazepine drugs may correlate with behavioral variation, including vulnerability to disease states such as alcoholism. In the rat, variation in alcohol and benzodiazepine sensitivity has been correlated with an inherited variant of the GABAA alpha 6 receptor. The authors detected a Pro385Ser [1236C > T] amino acid substitution in the human GABAA alpha 6 that may influence alcohol sensitivity. In this pilot study, they evaluated the contribution of this polymorphism to benzodiazepine sensitivity. Sensitivity to diazepam was assessed in 51 children of alcoholics by using two eye movement measures: peak saccadic velocity and average smooth pursuit gain. Association analysis was performed with saccadic velocity and smooth pursuit gain as dependent variables and comparing Pro385/Ser385 heterozygotes and Pro385/Pro385 homozygotes. The Pro385Ser genotype was associated with less diazepam-induced impairment of saccadic velocity but not with smooth pursuit gain. The Pro385Ser genotype may play a role in benzodiazepine sensitivity and conditions, such as alcoholism, that may be correlated with this trait.

Somato-synaptic variation of GABA(A) receptors in cultured murine cerebellar granule cells: investigation of the role of the alpha6 subunit.

PubMed

Mellor, J R; Wisden, W; Randall, A D

2000-07-10

Electrophysiological investigation of cultured cerebellar murine granule cells revealed differences between the GABA(A) receptors at inhibitory synapses and those on the cell body. Specifically, mIPSCs decayed more rapidly than cell body receptors deactivated, the mean single channel conductance at the synapse (32 pS) was greater than that at cell body (21 pS) and only cell body receptors were sensitive to Zn(2+) (150 microM), which depressed response amplitude by 82+/-5% and almost doubled the rate of channel deactivation. The GABA(A) receptor alpha6 subunit is selectively expressed in cerebellar granule cells. Although concentrated at synapses, it is also found on extrasynaptic membranes. Using a mouse line (Deltaalpha6lacZ) lacking this subunit, we investigated its role in the somato-synaptic differences in GABA(A) receptor function. All differences between cell body and synaptic GABA(A) receptors observed in wild-type (WT) granule cells persisted in Deltaalpha6lacZ cells, thus demonstrating that they are not specifically due to the cellular distribution of the alpha6 subunit. However, mIPSCs from WT and Deltaalpha6lacZ cells differed in both their kinetics (faster decay in WT cells) and underlying single channel conductance (32 pS WT, 25 pS Deltaalpha6lacZ). This provides good evidence for a functional contribution of the alpha6 subunit to postsynaptic GABA(A) receptors in these cells. Despite this, deactivation kinetics of mIPSCs in WT and Deltaalpha6lacZ granule cells exhibited similar benzodiazepene (BDZ) sensitivity. This suggests that the enhanced BDZ-induced ataxia seen in Deltaalpha6lacZ mice may reflect physiological activity at extrasynaptic receptors which, unlike those at synapses, display differential BDZ-sensitivity in WT and Deltaalpha6lacZ granule cells (Jones, A.M., Korpi, E.R., McKernan, R.M., Nusser, Z., Pelz, R., Makela, R., Mellor, J.R., Pollard, S., Bahn, S., Stephenson, F.A., Randall, A.D., Sieghart, W., Somogyi, P., Smith, A.J.H., Wisden

Altered GABAA receptor-mediated synaptic transmission disrupts the firing of gonadotropin-releasing hormone neurons in male mice under conditions that mimic steroid abuse

PubMed Central

Penatti, Carlos A A; Davis, Matthew C; Porter, Donna M; Henderson, Leslie P

2010-01-01

Gonadotropin–releasing hormone (GnRH) neurons are the central regulators of reproduction. GABAergic transmission plays a critical role in pubertal activation of pulsatile GnRH secretion. Self-administration of excessive doses of anabolic androgenic steroids (AAS) disrupts reproductive function and may have critical repercussions for pubertal onset in adolescent users. Here, we demonstrate that chronic treatment of adolescent male mice with the AAS, 17α-methyltestosterone (17αMT), significantly decreased action potential frequency in GnRH neurons, reduced the serum gonadotropin levels, and decreased testes mass. AAS treatment did not induce significant changes in GABAA receptor subunit mRNA levels or alter the amplitude or decay kinetics of GABAA receptor-mediated spontaneous postsynaptic currents (sPSC) or tonic currents in GnRH neurons. However, AAS treatment significantly increased action potential frequency in neighboring medial preoptic area (mPOA) neurons and GABAA receptor-mediated sPSC frequency in GnRH neurons. In addition, physical isolation of the more lateral aspects of the mPOA from the medially-localized GnRH neurons abrogated the AAS-induced increase in GABAA receptor-mediated sPSC frequency and the decrease in action potential firing in the GnRH cells. Our results indicate that AAS act predominantly on steroid-sensitive presynaptic neurons within the mPOA to impart significant increases in GABAA receptor-mediated inhibitory tone onto downstream GnRH neurons resulting in diminished activity of these pivotal mediators of reproductive function. These AAS-induced changes in central GABAergic circuits of the forebrain may significantly contribute to the disruptive actions of these drugs on pubertal maturation and the development of reproductive competence in male steroid abusers. PMID:20463213

Discrimination between olfactory receptor agonists and non-agonists.

PubMed

Topin, Jérémie; de March, Claire A; Charlier, Landry; Ronin, Catherine; Antonczak, Serge; Golebiowski, Jérôme

2014-08-11

A joint approach combining free-energy calculations and calcium-imaging assays on the broadly tuned human 1G1 olfactory receptor is reported. The free energy of binding of ten odorants was computed by means of molecular-dynamics simulations. This state function allows separating the experimentally determined eight agonists from the two non-agonists. This study constitutes a proof-of-principle for the computational deorphanization of olfactory receptors. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Regulation of (/sup 3/H)GABA release from strips of guinea pig urinary bladder

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shirakawa, J.; Taniyama, K.; Iwai, S.

1988-12-01

The presence of receptors that regulate the release of gamma-aminobutyric acid (GABA) was studied in strips of the guinea pig urinary bladder. GABA (10(-8)-10(-5) M) and muscimol (10(-8)-10(-5) M), but not baclofen (10(-5) M), reduced the Ca2+-dependent, tetrodotoxin-resistant release of (/sup 3/H)GABA evoked by high K+ from the urinary bladder strips preloaded with (/sup 3/H)GABA. The inhibitory effect of muscimol was antagonized by bicuculline and potentiated by diazepam, clonazepam, and pentobarbital sodium. The potentiating effect of clonazepam was antagonized by Ro 15-1788. Acetylcholine (ACh) inhibited the high K+-evoked release of (/sup 3/H)GABA. The inhibitory effect of ACh was antagonized bymore » atropine sulfate and pirenzepine but not by hexamethonium. Norepinephrine (NE) inhibited the evoked release of (/sup 3/H)GABA. The inhibitory effect of NE was mimicked by clonidine, but not by phenylephrine, and was antagonized by yohimbine but not by prazosin. These results provide evidence that the release of GABA from strips of guinea pig urinary bladder is regulated via the bicuculline-sensitive GABAA receptor, M1-muscarinic, and alpha 2-adrenergic receptors.« less

GABAB receptor modulation of the release of substance P from capsaicin-sensitive neurones in the rat trachea in vitro.

PubMed Central

Ray, N. J.; Jones, A. J.; Keen, P.

1991-01-01

1. The role of gamma-aminobutyric acid (GABA) as an inhibitory transmitter in the central nervous system is well documented. Recently, GABAA and GABAB receptors have been identified in the peripheral nervous system, notably on primary afferent neurones (PAN). We have utilised a multi-superfusion system to investigate the effect of selective GABA receptor agonists and antagonists on the release of substance P (SP) from the rat trachea in vitro. 2. GABA (1-100 microM) did not affect spontaneous release of SP-like immunoreactivity (LI) but caused dose-related inhibition of calcium-dependent potassium (60 mM)-stimulated SP-LI release. The greatest inhibition of 77.7 +/- 18.8% was observed at 100 microM. 3. The inhibitory effect of GABA was mimicked by the GABAB receptor agonist, (+/-)-baclofen (1-100 microM), but not the GABAA receptor agonist, 3-amino-1-propane-sulphonic acid (3-APS, 1-100 microM). Baclofen (100 microM) had no effect on SP-LI release stimulated by capsaicin (1 microM). 4. The inhibitory effect of baclofen (30 microM) was significantly reduced by prior and concomitant exposure to the GABAB receptor antagonist, phacolofen (100 microM) but not the GABAA receptor antagonist, bicuculline (10 microM). Neither antagonist, alone, affected spontaneous or potassium-stimulated SP-LI release. 5. We conclude that activation of pre-synaptic GABAB receptors on the peripheral termini of PANs in the rat trachea inhibits SP-LI release and suggest that GABAB receptor agonists may be of value in the therapeutic treatment of asthma. PMID:1713105

GABA(A) receptor antagonism in the ventrocaudal periaqueductal gray increases anxiety in the anxiety-resistant postpartum rat.

PubMed

Miller, Stephanie M; Piasecki, Christopher C; Peabody, Mitchell F; Lonstein, Joseph S

2010-06-01

Postpartum mammals show suppressed anxiety, which is necessary for their ability to appropriately care for offspring. It is parsimonious to suggest that the neurobiological basis of this reduced anxiety is similar to that of non-parturient animals, involving GABA(A) receptor activity in sites including the midbrain periaqueductal gray (PAG). In Experiment 1, postpartum and diestrous virgin female rats received an intraperitoneal injection of the GABA(A) receptor antagonist (+)-bicuculline (0, 2 and 4 mg/kg) and anxiety-related behavior was assessed with an elevated plus maze. The 4 mg/kg dose of (+)-bicuculline significantly increased anxiety-related behavior, particularly in the postpartum females. Experiment 2 revealed that bicuculline's action was within the central nervous system, because anxiety in neither dams nor virgins was significantly affected by intraperitoneal injection of bicuculline methiodide (0, 2 and 6 mg/kg), which does not readily cross the blood-brain-barrier. In Experiment 3, bicuculline methiodide (2.5 ng/side) was directly infused into the ventrocaudal PAG (cPAGv) and significantly increased dams' anxiety compared to saline-infused controls. These studies expand our knowledge of how GABA(A) receptor modulators affect anxiety behaviors in postpartum rats to the widely-used elevated plus maze, and indicate that the postpartum suppression of anxiety is in part a consequence of elevated GABAergic neurotransmission in the cPAGv. Copyright 2010 Elsevier Inc. All rights reserved.

RNAi screening of subtracted transcriptomes reveals tumor suppression by taurine-activated GABAA receptors involved in volume regulation

PubMed Central

van Nierop, Pim; Vormer, Tinke L.; Foijer, Floris; Verheij, Joanne; Lodder, Johannes C.; Andersen, Jesper B.; Mansvelder, Huibert D.; te Riele, Hein

2018-01-01

To identify coding and non-coding suppressor genes of anchorage-independent proliferation by efficient loss-of-function screening, we have developed a method for enzymatic production of low complexity shRNA libraries from subtracted transcriptomes. We produced and screened two LEGO (Low-complexity by Enrichment for Genes shut Off) shRNA libraries that were enriched for shRNA vectors targeting coding and non-coding polyadenylated transcripts that were reduced in transformed Mouse Embryonic Fibroblasts (MEFs). The LEGO shRNA libraries included ~25 shRNA vectors per transcript which limited off-target artifacts. Our method identified 79 coding and non-coding suppressor transcripts. We found that taurine-responsive GABAA receptor subunits, including GABRA5 and GABRB3, were induced during the arrest of non-transformed anchor-deprived MEFs and prevented anchorless proliferation. We show that taurine activates chloride currents through GABAA receptors on MEFs, causing seclusion of cell volume in large membrane protrusions. Volume seclusion from cells by taurine correlated with reduced proliferation and, conversely, suppression of this pathway allowed anchorage-independent proliferation. In human cholangiocarcinomas, we found that several proteins involved in taurine signaling via GABAA receptors were repressed. Low GABRA5 expression typified hyperproliferative tumors, and loss of taurine signaling correlated with reduced patient survival, suggesting this tumor suppressive mechanism operates in vivo. PMID:29787571

Changes of body temperature and thermoregulatory responses of freely moving rats during GABAergic pharmacological stimulation to the preoptic area and anterior hypothalamus in several ambient temperatures.

PubMed

Ishiwata, Takayuki; Saito, Takehito; Hasegawa, Hiroshi; Yazawa, Toru; Kotani, Yasunori; Otokawa, Minoru; Aihara, Yasutsugu

2005-06-28

Action of gamma-aminobutyric acid (GABA) in the preoptic area and anterior hypothalamus (PO/AH) has been implicated to regulate body temperature (T(b)). However, its precise role in thermoregulation remains unclear. Moreover, little is known about its release pattern in the PO/AH during active thermoregulation. Using microdialysis and telemetry techniques, we measured several parameters related to thermoregulation of freely moving rats during pharmacological stimulation of GABA in normal (23 degrees C), cold (5 degrees C), and hot (35 degrees C) ambient temperatures. We also measured extracellular GABA levels in the PO/AH during cold (5 degrees C) and heat (35 degrees C) exposure combined with microdialysis and high performance liquid chromatography (HPLC). Perfusion of GABA(A) agonist muscimol into the PO/AH increased T(b), which is associated with increased heart rate (HR), as an index of heat production in all ambient temperatures. Although tail skin temperature (T(tail)) as an index of heat loss increased only under normal ambient temperatures, its response was relatively delayed in comparison with HR and T(b), suggesting that the increase in T(tail) was a secondary response to increased HR and T(b). Locomotor activity also increased in all ambient temperatures, but its response was not extraordinary. Interestingly, thermoregulatory responses were different after perfusion of GABA(A) antagonist bicuculline at each ambient temperature. In normal ambient temperature conditions, perfusion of bicuculline had no effect on any parameter. However, under cold ambient temperature, the procedure induced significant hypothermia concomitant with a decrease in HR in spite of hyperactivity and increase of T(tail). It induced hyperthermia with the increase of HR but no additional change of T(tail) in hot ambient temperature conditions. Furthermore, the extracellular GABA level increased significantly during cold exposure. Its release was lower during heat exposure than in a

GABA-A receptors in mPOAH simultaneously regulate sleep and body temperature in freely moving rats.

PubMed

Jha, S K; Yadav, V; Mallick, B N

2001-09-01

Sleep-wakefulness and body temperature are two circadian rhythmic biological phenomena. The role of GABAergic inputs in the medial preoptico-anterior hypothalamus (mPOAH) on simultaneous regulation of those phenomena was investigated in freely moving normally behaving rats. The GABA-A receptors were blocked by microinjecting picrotoxin, and the effects on electrophysiological parameters signifying sleep-wakefulness, rectal temperature and brain temperature were recorded simultaneously. The results suggest that, normally, GABA in the medial preoptic area acts through GABA-A receptor that induces sleep and prevents an excessive rise in body temperature. However, the results do not allow us to comment on the cause and effect relationship, if any, between changes in sleep-wakefulness and body temperature. The changes in brain and rectal temperatures showed a positive correlation, however, the former varied within a narrower range than that of the latter.

Negative cooperativity in binding of muscarinic receptor agonists and GDP as a measure of agonist efficacy

PubMed Central

Jakubík, J; Janíčková, H; El-Fakahany, EE; Doležal, V

2011-01-01

BACKGROUND AND PURPOSE Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5′-γ−thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M2 muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative measure of agonist efficacy. EXPERIMENTAL APPROACH Filtration and scintillation proximity assays measured equilibrium binding as well as binding kinetics of [35S]GTPγS and [3H]GDP to a mixture of G-proteins as well as individual classes of G-proteins upon binding of structurally different agonists to the M2 muscarinic acetylcholine receptor. KEY RESULTS Agonists displayed biphasic competition curves with the antagonist [3H]-N-methylscopolamine. GTPγS (1 µM) changed the competition curves to monophasic with low affinity and 50 µM GDP produced a similar effect. Depletion of membrane-bound GDP increased the proportion of agonist high-affinity sites. Carbachol accelerated the dissociation of [3H]GDP from membranes. The inverse agonist N-methylscopolamine slowed GDP dissociation and GTPγS binding without changing affinity for GDP. Carbachol affected both GDP association with and dissociation from Gi/o G-proteins but only its dissociation from Gs/olf G-proteins. CONCLUSIONS AND IMPLICATIONS These findings suggest the existence of a low-affinity agonist-receptor conformation complexed with GDP-liganded G-protein. Also the negative cooperativity between GDP and agonist binding at the receptor/G-protein complex determines agonist efficacy. GDP binding reveals differences in action of agonists versus inverse agonists as well as differences in activation of Gi/o versus Gs/olf G-proteins that are not identified by conventional GTPγS binding. PMID:20958290

Negative cooperativity in binding of muscarinic receptor agonists and GDP as a measure of agonist efficacy.

PubMed

Jakubík, J; Janíčková, H; El-Fakahany, E E; Doležal, V

2011-03-01

Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5'-γ-thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M₂ muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative measure of agonist efficacy. Filtration and scintillation proximity assays measured equilibrium binding as well as binding kinetics of [³⁵S]GTPγS and [³H]GDP to a mixture of G-proteins as well as individual classes of G-proteins upon binding of structurally different agonists to the M₂ muscarinic acetylcholine receptor. Agonists displayed biphasic competition curves with the antagonist [³H]-N-methylscopolamine. GTPγS (1 µM) changed the competition curves to monophasic with low affinity and 50 µM GDP produced a similar effect. Depletion of membrane-bound GDP increased the proportion of agonist high-affinity sites. Carbachol accelerated the dissociation of [³H]GDP from membranes. The inverse agonist N-methylscopolamine slowed GDP dissociation and GTPγS binding without changing affinity for GDP. Carbachol affected both GDP association with and dissociation from G(i/o) G-proteins but only its dissociation from G(s/olf) G-proteins. These findings suggest the existence of a low-affinity agonist-receptor conformation complexed with GDP-liganded G-protein. Also the negative cooperativity between GDP and agonist binding at the receptor/G-protein complex determines agonist efficacy. GDP binding reveals differences in action of agonists versus inverse agonists as well as differences in activation of G(i/o) versus G(s/olf) G-proteins that are not identified by conventional GTPγS binding. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Modulation by bicuculline and penicillin of the block by t-butyl-bicyclo-phosphorothionate (TBPS) of GABAA-receptor mediated Cl−-current responses in rat striatal neurones

PubMed Central

Behrends, Jan C

2000-01-01

T-butyl-bicyclo-phosphorothionate (TBPS) is a prototypical representative of the cage-convulsants which act through a use-dependent block of the GABAA-receptor-ionophore complex. Using current recordings from cultured neurones of rat striatum the manner was investigated in which two antagonists, bicuculline and penicillin, presumably acting at the agonist binding site and in the ionic channel, respectively, modify the rate of block by TBPS. Penicillin (5 or 10 mM) did not slow the rate of block by TBPS, but produced a significant enhancement of block rate, which, however, was inversely related to the degree of antagonism by penicillin of the GABA-induced current. Bicuculline (10 μM) reduced the rate of block by TBPS. However, this effect was 3 fold weaker than its GABA-antagonistic action. The slowing of block rate and the current antagonism exhibited a biphasic, positive-negative relationship. Co-application of bicuculline (100 μM) in a concentration that produced nearly complete antagonism and TBPS (10 μM) resulted in a marked (∼40%) reduction of subsequent GABA response amplitudes compatible with a direct, bicuculline-induced conformational change in the receptor required for the binding of and block by TBPS. The lack of protection afforded by the channel blocker penicillin as well as the lack of correlation between bicuculline antagonism of the Cl−-current and its efficiency in protecting against TBPS block is evidence against an open channel blocking mechanism for TBPS. TBPS does, therefore, not appear to gain access to its binding site via the open pore but through alternative routes regulated from the agonist binding site. PMID:10694249

Insights into structure–activity relationship of GABAA receptor modulating coumarins and furanocoumarins

PubMed Central

Singhuber, Judith; Baburin, Igor; Ecker, Gerhard F.; Kopp, Brigitte; Hering, Steffen

2011-01-01

The coumarins imperatorin and osthole are known to exert anticonvulsant activity. We have therefore analyzed the modulation of GABA-induced chloride currents (IGABA) by a selection of 18 coumarin derivatives on recombinant α1β2γ2S GABAA receptors expressed in Xenopus laevis oocytes by means of the two-microelectrode voltage clamp technique. Osthole (EC50=14±1 μM) and oxypeucedanin (EC50=25±8 μM) displayed the highest efficiency with IGABA potentiation of 116±4% and 547±56%, respectively. IGABA enhancement by osthole and oxypeucedanin was not inhibited by flumazenil (1 μM) indicating an interaction with a binding site distinct from the benzodiazepine binding site. In general, prenyl residues are essential for the positive modulatory activity, while longer side chains or bulkier residues (e.g. geranyl residues) diminish IGABA modulation. Generation of a binary classification tree revealed the importance of polarisability, which is sufficient to distinguish actives from inactives. A 4-point pharmacophore model based on oxypeucedanin – comprising three hydrophobic and one aromatic feature – identified 6 out of 7 actives as hits. In summary, (oxy-)prenylated coumarin derivatives from natural origin represent new GABAA receptor modulators. PMID:21749864

Long-term administration with levonorgestrel decreases allopregnanolone levels and alters GABA(A) receptor subunit expression and anxiety-like behavior.

PubMed

Porcu, Patrizia; Mostallino, Maria Cristina; Sogliano, Cristiana; Santoru, Francesca; Berretti, Roberta; Concas, Alessandra

2012-08-01

Fluctuations in the concentrations of the neuroactive steroid allopregnanolone are thought to influence γ-amino-butyric acid type A (GABA(A)) receptor gene expression and function. Long-term treatment with ethinyl estradiol (EE) plus levonorgestrel (LNG), two of the most widely used steroids in the hormonal contraceptive pill, decreases allopregnanolone levels in rat cerebral cortex and plasma, alters GABA(A) receptor expression and induces anxiety-like behavior. We evaluated which component of the hormonal contraceptive pill is responsible for the aforementioned changes. Female rats were injected subcutaneously (s.c.) with EE (0.030 mg) or LNG (0.125 mg) once a day for 4 weeks. Compared to the respective vehicle-treated control groups, EE decreased cerebral cortical levels of allopregnanolone, progesterone and pregnenolone by 76, 72 and 33%, respectively and hippocampal levels by 52, 56 and 50%, respectively. Likewise, LNG decreased cerebral cortical levels of allopregnanolone, progesterone and pregnenolone by 75, 68 and 33%, respectively, and hippocampal levels by 55, 65 and 60%, respectively. Administration of LNG, but not EE, increased the abundance of the γ2 subunit peptide in cerebral cortex and hippocampus by 38 and 59%, respectively. Further, LNG, but not EE, decreased the time spent and the number of entries into the open arms of the elevated plus maze by 56 and 43%, respectively, an index of anxiety-like behavior. These results suggest that alterations in GABA(A) receptor subunit expression and anxiety-like behavior induced by long-term treatment with combined EE/LNG appear to be caused by LNG. Given that both EE and LNG decrease allopregnanolone levels in a similar manner, these results further suggest that changes in allopregnanolone levels are not associated with GABA(A) receptor expression. Copyright © 2012 Elsevier Inc. All rights reserved.

Estrous Cycle Regulation of Extrasynaptic δ-Containing GABAA Receptor-Mediated Tonic Inhibition and Limbic Epileptogenesis

PubMed Central

Wu, Xin; Gangisetty, Omkaram; Carver, Chase Matthew

2013-01-01

The ovarian cycle affects susceptibility to behavioral and neurologic conditions. The molecular mechanisms underlying these changes are poorly understood. Deficits in cyclical fluctuations in steroid hormones and receptor plasticity play a central role in physiologic and pathophysiologic menstrual conditions. It has been suggested that synaptic GABAA receptors mediate phasic inhibition in the hippocampus and extrasynaptic receptors mediate tonic inhibition in the dentate gyrus. Here we report a novel role of extrasynaptic δ-containing GABAA receptors as crucial mediators of the estrous cycle–related changes in neuronal excitability in mice, with hippocampus subfield specificity. In molecular and immunofluorescence studies, a significant increase occurred in δ-subunit, but not α4- and γ2-subunits, in the dentate gyrus during diestrus. However, δ-subunit upregulation was not evident in the CA1 region. The δ-subunit expression was undiminished by age and ovariectomy and in mice lacking progesterone receptors, but it was significantly reduced by finasteride, a neurosteroid synthesis inhibitor. Electrophysiologic studies confirmed greater potentiation of GABA currents by progesterone-derived neurosteroid allopregnanolone in dissociated dentate gyrus granule cells in diestrus than in CA1 pyramidal cells. The baseline conductance and allopregnanolone potentiation of tonic currents in dentate granule cells from hippocampal slices were higher than in CA1 pyramidal cells. In behavioral studies, susceptibility to hippocampus kindling epileptogenesis was lower in mice during diestrus. These results demonstrate the estrous cycle–related plasticity of neurosteroid-sensitive, δ-containing GABAA receptors that mediate tonic inhibition and seizure susceptibility. These findings may provide novel insight on molecular cascades of menstrual disorders like catamenial epilepsy, premenstrual syndrome, and migraine. PMID:23667248

Insulin-Independent GABAA Receptor-Mediated Response in the Barrel Cortex of Mice with Impaired Met Activity

PubMed Central

Lo, Fu-Sun; Erzurumlu, Reha S.

2016-01-01

Autism spectrum disorder (ASD) is a neurodevelopmental disorder caused by genetic variants, susceptibility alleles, and environmental perturbations. The autism associated gene MET tyrosine kinase has been implicated in many behavioral domains and endophenotypes of autism, including abnormal neural signaling in human sensory cortex. We investigated somatosensory thalamocortical synaptic communication in mice deficient in Met activity in cortical excitatory neurons to gain insights into aberrant somatosensation characteristic of ASD. The ratio of excitation to inhibition is dramatically increased due to decreased postsynaptic GABAA receptor-mediated inhibition in the trigeminal thalamocortical pathway of mice lacking active Met in the cerebral cortex. Furthermore, in contrast to wild-type mice, insulin failed to increase GABAA receptor-mediated response in the barrel cortex of mice with compromised Met signaling. Thus, lacking insulin effects may be a risk factor in ASD pathogenesis. SIGNIFICANCE STATEMENT A proposed common cause of neurodevelopmental disorders is an imbalance in excitatory neural transmission, provided by the glutamatergic neurons, and the inhibitory signals from the GABAergic interneurons. Many genes associated with autism spectrum disorders impair synaptic transmission in the expected cell type. Previously, inactivation of the autism-associated Met tyrosine kinase receptor in GABAergic interneurons led to decreased inhibition. In thus report, decreased Met signaling in glutamatergic neurons had no effect on excitation, but decimated inhibition. Further experiments indicate that loss of Met activity downregulates GABAA receptors on glutamatergic neurons in an insulin independent manner. These data provide a new mechanism for the loss of inhibition and subsequent abnormal excitation/inhibition balance and potential molecular candidates for treatment or prevention. PMID:27030755

Dose-dependent EEG effects of zolpidem provide evidence for GABA(A) receptor subtype selectivity in vivo.

PubMed

Visser, S A G; Wolters, F L C; van der Graaf, P H; Peletier, L A; Danhof, M

2003-03-01

Zolpidem is a nonbenzodiazepine GABA(A) receptor modulator that binds in vitro with high affinity to GABA(A) receptors expressing alpha(1) subunits but with relatively low affinity to receptors expressing alpha(2), alpha(3), and alpha(5) subunits. In the present study, it was investigated whether this subtype selectivity could be detected and quantified in vivo. Three doses (1.25, 5, and 25 mg) of zolpidem were administered to rats in an intravenous infusion over 5 min. The time course of the plasma concentrations was determined in conjunction with the change in the beta-frequency range of the EEG as pharmacodynamic endpoint. The concentration-effect relationship of the three doses showed a dose-dependent maximum effect and a dose-dependent potency. The data were analyzed for one- or two-site binding using two pharmacodynamic models based on 1) the descriptive model and 2) a novel mechanism-based pharmacokinetic/pharmacodynamic (PK/PD) model for GABA(A) receptor modulators that aims to separates drug- and system-specific properties, thereby allowing the estimation of in vivo affinity and efficacy. The application of two-site models significantly improved the fits compared with one-site models. Furthermore, in contrast to the descriptive model, the mechanism-based PK/PD model yielded dose-independent estimates for affinity (97 +/- 40 and 33,100 +/- 14,800 ng x ml(-1)). In conclusion, the mechanism-based PK/PD model is able to describe and explain the observed dose-dependent EEG effects of zolpidem and suggests the subtype selectivity of zolpidem in vivo.

Selective labelling of diazepam-insensitive GABAA receptors in vivo using [3H]Ro 15-4513.

PubMed

Pym, Luanda J; Cook, Susan M; Rosahl, Thomas; McKernan, Ruth M; Atack, John R

2005-11-01

Classical benzodiazepines (BZs), such as diazepam, bind to GABAA receptors containing alpha1, alpha2, alpha3 or alpha5 subunits that are therefore described as diazepam-sensitive (DS) receptors. However, the corresponding binding site of GABAA receptors containing either an alpha4 or alpha6 subunit do not bind the classical BZs and are therefore diazepam-insensitive (DIS) receptors; a difference attributable to a single amino acid (histidine in alpha1, alpha2, alpha3 and alpha5 subunits and arginine in alpha4 and alpha6). Unlike classical BZs, the imidazobenzodiazepines Ro 15-4513 and bretazenil bind to both DS and DIS populations of GABAA receptors. In the present study, an in vivo assay was developed using lorazepam to fully occupy DS receptors such that [3H]Ro 15-4513 was then only able to bind to DIS receptors. When dosed i.v., [3H]Ro 15-4513 rapidly entered and was cleared from the brain, with approximately 70% of brain radioactivity being membrane-bound. Essentially all membrane binding to DS+DIS receptors could be displaced by unlabelled Ro 15-4513 or bretazenil, with respective ID50 values of 0.35 and 1.2 mg kg(-1). A dose of 30 mg kg(-1) lorazepam was used to block all DS receptors in a [3H]Ro 15-1788 in vivo binding assay. When predosed in a [3H]Ro 15-4513 binding assay, lorazepam blocked [3H]Ro 15-4513 binding to DS receptors, with the remaining binding to DIS receptors accounting for 5 and 23% of the total (DS plus DIS) receptors in the forebrain and cerebellum, respectively. The in vivo binding of [3H]Ro 15-4513 to DIS receptors in the presence of lorazepam was confirmed using alpha1H101R knock-in mice, in which alpha1-containing GABAA receptors are rendered diazepam insensitive by mutation of the histidine that confers diazepam sensitivity to arginine. In these mice, and in the presence of lorazepam, there was an increase of in vivo [3H]Ro 15-4513 binding in the forebrain and cerebellum from 4 and 15% to 36 and 59% of the total (i.e. DS plus DIS) [3H

Mechanisms of anabolic androgenic steroid inhibition of mammalian ɛ-subunit-containing GABAA receptors

PubMed Central

Jones, Brian L; Whiting, Paul J; Henderson, Leslie P

2006-01-01

GABAergic transmission regulates the activity of gonadotrophin-releasing hormone (GnRH) neurons in the preoptic area/hypothalamus that control the onset of puberty and the expression of reproductive behaviours. One of the hallmarks of illicit use of anabolic androgenic steroids (AAS) is disruption of behaviours under neuroendocrine control. GnRH neurons are among a limited population of cells that express high levels of the ɛ-subunit of the GABAA receptor. To better understand the actions of AAS on neuroendocrine mechanisms, we have characterized modulation of GABAA receptor-mediated currents in mouse native GnRH neurons and in heterologous cells expressing recombinant α2β3ɛ-receptors. GnRH neurons exhibited robust currents in response to millimolar concentrations of GABA and a picrotoxin (PTX)-sensitive, bicuculline-insensitive current that probably arises from spontaneous openings of GABAA receptors. The AAS 17α-methyltestosterone (17α-MeT) inhibited spontaneous and GABA-evoked currents in GnRH neurons. For recombinant α2β3ɛ-receptors, 17α-MeT inhibited phasic and tonic GABA-elicited responses, accelerated desensitization and slowed paired pulse response recovery. Single channel analysis indicated that GABA-evoked events could be described by three open dwell components and that 17α-MeT enhanced residence in the intermediate dwell state. This AAS also inhibited a PTX-sensitive, spontaneous current (open probability, ∼0.15–0.2) in a concentration-dependent fashion (IC50 ≈ 9 μm). Kinetic modelling indicated that the inhibition induced by 17α-MeT occurs by an allosteric block in which the AAS interacts preferentially with a closed state and promotes accumulation in that state. Finally, studies with a G302S mutant ɛ-subunit suggest that this residue within the transmembrane domain TM2 plays a role in mediating AAS binding and modulation. In sum, our results indicate that inclusion of the ɛ-subunit significantly alters the profile of AAS

MT-7716, a novel selective nonpeptidergic NOP receptor agonist, effectively blocks ethanol-induced increase in GABAergic transmission in the rat central amygdala

PubMed Central

Kallupi, Marsida; Oleata, Christopher S.; Luu, George; Teshima, Koji; Ciccocioppo, Roberto; Roberto, Marisa

2014-01-01

The GABAergic system in the central amygdala (CeA) plays a major role in ethanol dependence and the anxiogenic-like response to ethanol withdrawal. A large body of evidence shows that Nociceptin/Orphanin FQ (N/OFQ) regulates ethanol intake and anxiety-like behavior. In the rat, ethanol significantly augments CeA GABA release, whereas N/OFQ diminishes it. Using electrophysiological techniques in an in vitro slice preparation, in this study we investigated the effects of a nonpeptidergic NOP receptor agonist, MT-7716 [(R)-2-3-[1-(Acenaphthen-1-yl)piperidin-4-yl]-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl-N-methylacetamide hydrochloride hydrate], and its interaction with ethanol on GABAergic transmission in CeA slices of naïve rats. We found that MT-7716 dose-dependently (100–1000 nM) diminished evoked GABAA receptor-mediated inhibitory postsynaptic potentials (IPSPs) and increased paired-pulse facilitation (PPF) ratio of these evoked IPSPs, suggesting a presynaptic site of action of the MT-7716 by decreasing GABA release at CeA synapses. The presynaptic action of MT-7716 was also supported by the significant decrease in the frequency of miniature inhibitory postsynaptic currents (mIPSCs) induced by the nociceptin receptor (NOP) agonist. Interestingly, MT-7716 prevented the ethanol-induced augmentation of evoked IPSPs. A putative selective NOP antagonist, [Nphe1]Nociceptin(1–13)NH2, totally prevented the MT-7716-induced inhibition of IPSP amplitudes indicating that MT-7716 exerts its effect through NOPs. These data provide support for an interaction between the nociceptin and GABAergic systems in the CeA and for the anti-alcohol properties of the NOP activation. The development of a synthetic nonpeptidergic NOP receptor agonist such as MT-7716 may represent a useful therapeutic target for alcoholism. PMID:24600360

Enhanced GABAA-Mediated Tonic Inhibition in Auditory Thalamus of Rats with Behavioral Evidence of Tinnitus.

PubMed

Sametsky, Evgeny A; Turner, Jeremy G; Larsen, Deb; Ling, Lynne; Caspary, Donald M

2015-06-24

Accumulating evidence suggests a role for inhibitory neurotransmitter dysfunction in the pathology of tinnitus. Opposing hypotheses proposed either a pathologic decrease or increase of GABAergic inhibition in medial geniculate body (MGB). In thalamus, GABA mediates fast synaptic inhibition via synaptic GABAA receptors (GABAARs) and persistent tonic inhibition via high-affinity extrasynaptic GABAARs. Given that extrasynaptic GABAARs control the firing mode of thalamocortical neurons, we examined tonic GABAAR currents in MGB neurons in vitro, using the following three groups of adult rats: unexposed control (Ctrl); sound exposed with behavioral evidence of tinnitus (Tin); and sound exposed with no behavioral evidence of tinnitus (Non-T). Tonic GABAAR currents were evoked using the selective agonist gaboxadol. Months after a tinnitus-inducing sound exposure, gaboxadol-evoked tonic GABAAR currents showed significant tinnitus-related increases contralateral to the sound exposure. In situ hybridization studies found increased mRNA levels for GABAAR δ-subunits contralateral to the sound exposure. Tin rats showed significant increases in the number of spikes per burst evoked using suprathreshold-injected current steps. In summary, we found little evidence of tinnitus-related decreases in GABAergic neurotransmission. Tinnitus and chronic pain may reflect thalamocortical dysrhythmia, which results from abnormal theta-range resonant interactions between thalamus and cortex, due to neuronal hyperpolarization and the initiation of low-threshold calcium spike bursts (Walton and Llinás, 2010). In agreement with this hypothesis, we found tinnitus-related increases in tonic extrasynaptic GABAAR currents, in action potentials/evoked bursts, and in GABAAR δ-subunit gene expression. These tinnitus-related changes in GABAergic function may be markers for tinnitus pathology in the MGB. Copyright © 2015 the authors 0270-6474/15/359369-12$15.00/0.

Role of α1- and α2-GABAA receptors in mediating the respiratory changes associated with benzodiazepine sedation

PubMed Central

Masneuf, S; Buetler, J; Koester, C; Crestani, F

2012-01-01

BACKGROUND AND PURPOSE The molecular substrates underlying the respiratory changes associated with benzodiazepine sedation are unknown. We examined the effects of different doses of diazepam and alprazolam on resting breathing in wild-type (WT) mice and clarified the contribution of α1- and α2-GABAA receptors, which mediate the sedative and muscle relaxant action of diazepam, respectively, to these drug effects using point-mutated mice possessing either α1H101R- or α2H101R-GABAA receptors insensitive to benzodiazepine. EXPERIMENTAL APPROACH Room air breathing was monitored using whole-body plethysmography. Different groups of WT mice were injected i.p. with diazepam (1–100 mg·kg−1), alprazolam (0.3, 1 or 3 mg·kg−1) or vehicle. α1H101R and α2H101R mice received 1 or 10 mg·kg−1 diazepam or 0.3 or 3 mg·kg−1 alprazolam. Respiratory frequency, tidal volume, time of expiration and time of inspiration before and 20 min after drug injection were analysed. KEY RESULTS Diazepam (10 mg·kg−1) decreased the time of expiration, thereby increasing the resting respiratory frequency, in WT and α2H101R mice, but not in α1H101R mice. The time of inspiration was shortened in WT and α1H101R mice, but not in α2H101R mice. Alprazolam (1–3 mg·kg−1) stimulated the respiratory frequency by shortening expiration and inspiration duration in WT mice. This tachypnoeic effect was partially conserved in α1H101R mice while absent in α2H101R mice. CONCLUSIONS AND IMPLICATIONS These results identify a specific role for α1-GABAA receptors and α2-GABAA receptors in mediating the shortening by benzodiazepines of the expiratory and inspiratory phase of resting breathing respectively. PMID:22044283

Mechanisms Underlying Tolerance after Long-Term Benzodiazepine Use: A Future for Subtype-Selective GABAA Receptor Modulators?

PubMed Central

Vinkers, Christiaan H.; Olivier, Berend

2012-01-01

Despite decades of basic and clinical research, our understanding of how benzodiazepines tend to lose their efficacy over time (tolerance) is at least incomplete. In appears that tolerance develops relatively quickly for the sedative and anticonvulsant actions of benzodiazepines, whereas tolerance to anxiolytic and amnesic effects probably does not develop at all. In light of this evidence, we review the current evidence for the neuroadaptive mechanisms underlying benzodiazepine tolerance, including changes of (i) the GABAA receptor (subunit expression and receptor coupling), (ii) intracellular changes stemming from transcriptional and neurotrophic factors, (iii) ionotropic glutamate receptors, (iv) other neurotransmitters (serotonin, dopamine, and acetylcholine systems), and (v) the neurosteroid system. From the large variance in the studies, it appears that either different (simultaneous) tolerance mechanisms occur depending on the benzodiazepine effect, or that the tolerance-inducing mechanism depends on the activated GABAA receptor subtypes. Importantly, there is no convincing evidence that tolerance occurs with α subunit subtype-selective compounds acting at the benzodiazepine site. PMID:22536226

Proton modulation of recombinant GABAA receptors: influence of GABA concentration and the β subunit TM2–TM3 domain

PubMed Central

Wilkins, Megan E; Hosie, Alastair M; Smart, Trevor G

2005-01-01

Regulation of GABAA receptors by extracellular pH exhibits a dependence on the receptor subunit composition. To date, the molecular mechanism responsible for the modulation of GABAA receptors at alkaline pH has remained elusive. We report here that the GABA-activated current can be potentiated at pH 8.4 for both αβ and αβγ subunit-containing receptors, but only at GABA concentrations below the EC40. Site-specific mutagenesis revealed that a single lysine residue, K279 in the β subunit TM2–TM3 linker, was critically important for alkaline pH to modulate the function of both α1β2 and α1β2γ2 receptors. The ability of low concentrations of GABA to reveal different pH titration profiles for GABAA receptors was also examined at acidic pH. At pH 6.4, GABA activation of αβγ receptors was enhanced at low GABA concentrations. This effect was ablated by the mutation H267A in the β subunit. Decreasing the pH further to 5.4 inhibited GABA responses via αβγ receptors, whereas those responses recorded from αβ receptors were potentiated. Inserting homologous β subunit residues into the γ2 subunit to recreate, in αβγ receptors, the proton modulatory profile of αβ receptors, established that in the presence of β2H267, the mutation γ2T294K was necessary to potentiate the GABA response at pH 5.4. This residue, T294, is homologous to K279 in the β subunit and suggests that a lysine at this position is an important residue for mediating the allosteric effects of both acidic and alkaline pH changes, rather than forming a direct site for protonation within the GABAA receptor. PMID:15946973

Neurodevelopmental disorders among individuals with duplication of 4p13 to 4p12 containing a GABAA receptor subunit gene cluster

PubMed Central

Polan, Michelle B; Pastore, Matthew T; Steingass, Katherine; Hashimoto, Sayaka; Thrush, Devon L; Pyatt, Robert; Reshmi, Shalini; Gastier-Foster, Julie M; Astbury, Caroline; McBride, Kim L

2014-01-01

Recent studies have shown that certain copy number variations (CNV) are associated with a wide range of neurodevelopmental disorders, including autism spectrum disorders (ASD), bipolar disorder and intellectual disabilities. Implicated regions and genes have comprised a variety of post synaptic complex proteins and neurotransmitter receptors, including gamma-amino butyric acid A (GABAA). Clusters of GABAA receptor subunit genes are found on chromosomes 4p12, 5q34, 6q15 and 15q11-13. Maternally inherited 15q11-13 duplications among individuals with neurodevelopmental disorders are well described, but few case reports exist for the other regions. We describe a family with a 2.42 Mb duplication at chromosome 4p13 to 4p12, identified in the index case and other family members by oligonucleotide array comparative genomic hybridization, that contains 13 genes including a cluster of four GABAA receptor subunit genes. Fluorescent in-situ hybridization was used to confirm the duplication. The duplication segregates with a variety of neurodevelopmental disorders in this family, including ASD (index case), developmental delay, dyspraxia and ADHD (brother), global developmental delays (brother), learning disabilities (mother) and bipolar disorder (maternal grandmother). In addition, we identified and describe another individual unrelated to this family, with a similar duplication, who was diagnosed with ASD, ADHD and borderline intellectual disability. The 4p13 to 4p12 duplication appears to confer a susceptibility to a variety of neurodevelopmental disorders in these two families. We hypothesize that the duplication acts through a dosage effect of GABAA receptor subunit genes, adding evidence for alterations in the GABAergic system in the etiology of neurodevelopmental disorders. PMID:23695283

Structural basis for ligand recognition at the benzodiazepine binding site of GABAA alpha 3 receptor, and pharmacophore-based virtual screening approach.

PubMed

Vijayan, R S K; Ghoshal, Nanda

2008-10-01

Given the heterogeneity of GABA(A) receptor, the pharmacological significance of identifying subtype selective modulators is increasingly being recognized. Thus, drugs selective for GABA(A) alpha(3) receptors are expected to display fewer side effects than the drugs presently in clinical use. Hence we carried out 3D QSAR (three-dimensional quantitative structure-activity relationship) studies on a series of novel GABA(A) alpha(3) subtype selective modulators to gain more insight into subtype affinity. To identify the 3D functional attributes required for subtype selectivity, a chemical feature-based pharmacophore, primarily based on selective ligands representing diverse structural classes was generated. The obtained pseudo receptor model of the benzodiazepine binding site revealed a binding mode akin to "Message-Address" concept. Scaffold hopping was carried out across multi-conformational May Bridge database for the identification of novel chemotypes. Further a focused data reduction approach was employed to choose a subset of enriched compounds based on "Drug likeness" and "Similarity-based" methods. These results taken together could provide impetus for rational design and optimization of more selective and high affinity leads with a potential to have decreased adverse effects.

Lindane blocks GABAA-mediated inhibition and modulates pyramidal cell excitability in the rat hippocampal slice.

PubMed

Joy, R M; Walby, W F; Stark, L G; Albertson, T E

1995-01-01

An in vitro paired-pulse orthodromic stimulation technique was used to examine the effects of lindane on excitatory afferent terminals, CA1 pyramidal cells and recurrent collateral evoked inhibition in the rat hippocampal slice. This was done to establish simultaneous effects on a simple neural network and to develop procedures for more detailed analyses of the effects of lindane. Hippocampal slices 400 microns thick were perfused with oxygenated artificial cerebrospinal fluid. Electrodes were placed in the CA1 region to record extracellular population spike (PS) or excitatory postsynaptic potential (EPSP) responses to stimulation of Schaffer collateral/commissural (SC/C) fibers. Gamma-aminobutyric acid (GABA)-mediated recurrent inhibition was measured using a paired-pulse technique. Perfusion with lindane produced both time and dose dependent changes in a number of the responses measured. The most striking effect produced by lindane was the loss of GABAA-mediated recurrent collateral inhibition. This tended to occur rapidly, often before changes in EPSP or PS responses could be detected. With longer exposures to lindane, repetitive discharge of pyramidal cells developed resulting in multiple PSs to single stimuli. Lindane (50 microM) also completely reversed the effects of the injectable anesthetic, propofol, a compound known to potentiate GABAA-mediated inhibition via a direct action on the GABAA receptor-chloride channel complex. An analysis of input/output relationships at varying stimulus intensities showed that lindane increased EPSP and PS response amplitudes at any given stimulus intensity resulting in a leftward shift in the EPSP amplitude/stimulus intensity, PS amplitude/stimulus intensity and PS amplitude/EPSP amplitude relationships. This effect was most noticeable with low intensity stimuli and became progressively less so as stimulus intensities approached those yielding maximal responses. In addition lindane significantly increased paired pulse

Alcohol use disorders and current pharmacological therapies: the role of GABAA receptors

PubMed Central

Liang, Jing; Olsen, Richard W

2014-01-01

Alcohol use disorders (AUD) are defined as alcohol abuse and alcohol dependence, which create large problems both for society and for the drinkers themselves. To date, no therapeutic can effectively solve these problems. Understanding the underlying mechanisms leading to AUD is critically important for developing effective and safe pharmacological therapies. Benzodiazepines (BZs) are used to reduce the symptoms of alcohol withdrawal syndrome. However, frequent use of BZs causes cross-tolerance, dependence, and cross-addiction to alcohol. The FDA-approved naltrexone and acamprosate have shown mixed results in clinical trials. Naltrexone is effective to treat alcohol dependence (decreased length and frequency of drinking bouts), but its severe side effects, including withdrawal symptoms, are difficult to overcome. Acamprosate showed efficacy for treating alcohol dependence in European trials, but two large US trials have failed to confirm the efficacy. Another FDA-approved medication, disulfiram, does not diminish craving, and it causes a peripheral neuropathy. Kudzu is the only natural medication mentioned by the National Institute on Alcohol Abuse and Alcoholism, but its mechanisms of action are not yet established. It has been recently shown that dihydromyricetin, a flavonoid purified from Hovenia, has unique effects on GABAA receptors and blocks ethanol intoxication and withdrawal in alcoholic animal models. In this article, we review the role of GABAA receptors in the treatment of AUD and currently available and potentially novel pharmacological agents. PMID:25066321

Astrocytes Modulate a Postsynaptic NMDA–GABAA-Receptor Crosstalk in Hypothalamic Neurosecretory Neurons

PubMed Central

Potapenko, Evgeniy S.; Biancardi, Vinicia C.; Zhou, Yiqiang

2013-01-01

A dynamic balance between the excitatory and inhibitory neurotransmitters glutamate and GABA is critical for maintaining proper neuronal activity in the brain. This balance is partly achieved via presynaptic interactions between glutamatergic and GABAAergic synapses converging into the same targets. Here, we show that in hypothalamic magnocellular neurosecretory neurons (MNCs), a direct crosstalk between postsynaptic NMDA receptors (NMDARs) and GABAA receptors (GABAARs) contributes to the excitatory/inhibitory balance in this system. We found that activation of NMDARs by endogenous glutamate levels controlled by astrocyte glutamate transporters, evokes a transient and reversible potentiation of postsynaptic GABAARs. This inter-receptor crosstalk is calcium-dependent and involves a kinase-dependent phosphorylation mechanism, but does not require nitric oxide as an intermediary signal. Finally, we found the NMDAR–GABAAR crosstalk to be blunted in rats with heart failure, a pathological condition in which the hypothalamic glutamate–GABA balance is tipped toward an excitatory predominance. Together, our findings support a novel form of glutamate–GABA interactions in MNCs, which involves crosstalk between NMDA and GABAA postsynaptic receptors, whose strength is controlled by the activity of local astrocytes. We propose this inter-receptor crosstalk to act as a compensatory, counterbalancing mechanism to dampen glutamate-mediated overexcitation. Finally, we propose that an uncoupling between NMDARs and GABAARs may contribute to exacerbated neuronal activity and, consequently, sympathohumoral activation in such disease conditions as heart failure. PMID:23303942

Emamectin is a non-selective allosteric activator of nicotinic acetylcholine receptors and GABAA/C receptors

PubMed Central

Xu, Xiaojun; Sepich, Caraline; Lukas, Ronald J; Zhu, Guonian; Chang, Yongchang

2016-01-01

Avermectins are a group of compounds isolated from a soil-dwelling bacterium. They have been widely used as parasiticides and insecticides, acting by relatively irreversible activation of invertebrate chloride channels. Emamectin is a soluble derivative of an avermectin. It is an insecticide, which persistently activates glutamate-gated chloride channels. However, its effects on mammalian ligand-gated ion channels are unknown. To this end, we tested the effect of emamectin on two cation selective nicotinic receptors and two GABA-gated chloride channels expressed in Xenopus oocytes using two-electrode voltage clamp. Our results demonstrate that emamectin could directly activate α7 nAChR, α4β2 nAChR, α1β2γ2 GABAA receptor and ρ1 GABAC receptor concentration dependently, with similar potencies for each channel. However, the potencies for it to activate these channels were at least two orders of magnitude lower than its potency of activating invertebrate glutamate-gated chloride channel. In contrast, ivermectin only activated the α1β2γ2 GABAA receptor. PMID:27049309

Emamectin is a non-selective allosteric activator of nicotinic acetylcholine receptors and GABAA/C receptors.

PubMed

Xu, Xiaojun; Sepich, Caraline; Lukas, Ronald J; Zhu, Guonian; Chang, Yongchang

2016-05-13

Avermectins are a group of compounds isolated from a soil-dwelling bacterium. They have been widely used as parasiticides and insecticides, acting by relatively irreversible activation of invertebrate chloride channels. Emamectin is a soluble derivative of an avermectin. It is an insecticide, which persistently activates glutamate-gated chloride channels. However, its effects on mammalian ligand-gated ion channels are unknown. To this end, we tested the effect of emamectin on two cation selective nicotinic receptors and two GABA-gated chloride channels expressed in Xenopus oocytes using two-electrode voltage clamp. Our results demonstrate that emamectin could directly activate α7 nAChR, α4β2 nAChR, α1β2γ2 GABAA receptor and ρ1 GABAC receptor concentration dependently, with similar potencies for each channel. However, the potencies for it to activate these channels were at least two orders of magnitude lower than its potency of activating invertebrate glutamate-gated chloride channel. In contrast, ivermectin only activated the α1β2γ2 GABAA receptor. Copyright © 2016 Elsevier Inc. All rights reserved.

Inhaled Anesthetic Responses of Recombinant Receptors and Knockin Mice Harboring α2(S270H/L277A) GABAA Receptor Subunits That Are Resistant to Isoflurane

PubMed Central

Werner, D. F.; Swihart, A.; Rau, V.; Jia, F.; Borghese, C. M.; McCracken, M. L.; Iyer, S.; Fanselow, M. S.; Oh, I.; Sonner, J. M.; Eger, E. I.; Harrison, N. L.; Harris, R. A.

2011-01-01

The mechanism by which the inhaled anesthetic isoflurane produces amnesia and immobility is not understood. Isoflurane modulates GABAA receptors (GABAA-Rs) in a manner that makes them plausible targets. We asked whether GABAA-R α2 subunits contribute to a site of anesthetic action in vivo. Previous studies demonstrated that Ser270 in the second transmembrane domain is involved in the modulation of GABAA-Rs by volatile anesthetics and alcohol, either as a binding site or a critical allosteric residue. We engineered GABAA-Rs with two mutations in the α2 subunit, changing Ser270 to His and Leu277 to Ala. Recombinant receptors with these mutations demonstrated normal affinity for GABA, but substantially reduced responses to isoflurane. We then produced mutant (knockin) mice in which this mutated subunit replaced the wild-type α2 subunit. The adult mutant mice were overtly normal, although there was evidence of enhanced neonatal mortality and fear conditioning. Electrophysiological recordings from dentate granule neurons in brain slices confirmed the decreased actions of isoflurane on mutant receptors contributing to inhibitory synaptic currents. The loss of righting reflex EC50 for isoflurane did not differ between genotypes, but time to regain the righting reflex was increased in N2 generation knockins. This effect was not observed at the N4 generation. Isoflurane produced immobility (as measured by tail clamp) and amnesia (as measured by fear conditioning) in both wild-type and mutant mice, and potencies (EC50) did not differ between the strains for these actions of isoflurane. Thus, immobility or amnesia does not require isoflurane potentiation of the α2 subunit. PMID:20807777

Tyrosine Phosphorylation of GABAA Receptor γ2-Subunit Regulates Tonic and Phasic Inhibition in the Thalamus

PubMed Central

Nani, Francesca; Bright, Damian P.; Revilla-Sanchez, Raquel; Tretter, Verena; Moss, Stephen J.

2013-01-01

GABA-mediated tonic and phasic inhibition of thalamic relay neurons of the dorsal lateral geniculate nucleus (dLGN) was studied after ablating tyrosine (Y) phosphorylation of receptor γ2-subunits. As phosphorylation of γ2 Y365 and Y367 reduces receptor internalization, to understand their importance for inhibition we created a knock-in mouse in which these residues are replaced by phenylalanines. On comparing wild-type (WT) and γ2Y365/367F+/− (HT) animals (homozygotes are not viable in utero), the expression levels of GABAA receptor α4-subunits were increased in the thalamus of female, but not male mice. Raised δ-subunit expression levels were also observed in female γ2Y365/367F +/− thalamus. Electrophysiological analyses revealed no difference in the level of inhibition in male WT and HT dLGN, while both the spontaneous inhibitory postsynaptic activity and the tonic current were significantly augmented in female HT relay cells. The sensitivity of tonic currents to the δ-subunit superagonist THIP, and the blocker Zn2+, were higher in female HT relay cells. This is consistent with upregulation of extrasynaptic GABAA receptors containing α4- and δ-subunits to enhance tonic inhibition. In contrast, the sensitivity of GABAA receptors mediating inhibition in the female γ2Y356/367F +/− to neurosteroids was markedly reduced compared with WT. We conclude that disrupting tyrosine phosphorylation of the γ2-subunit activates a sex-specific increase in tonic inhibition, and this most likely reflects a genomic-based compensation mechanism for the reduced neurosteroid sensitivity of inhibition measured in female HT relay neurons. PMID:23904608

Responses to GABA(A) receptor activation are altered in NTS neurons isolated from chronic hypoxic rats.

PubMed

Tolstykh, Gleb; Belugin, Sergei; Mifflin, Steve

2004-04-23

The inhibitory amino acid GABA is released within the nucleus of the solitary tract (NTS) during hypoxia and modulates the respiratory response to hypoxia. To determine if responses of NTS neurons to activation of GABA(A) receptors are altered following exposure to chronic hypoxia, GABA(A) receptor-evoked whole cell currents were measured in enzymatically dispersed NTS neurons from normoxic and chronic hypoxic rats. Chronic hypoxic rats were exposed to 10% O(2) for 9-12 days. Membrane capacitance was the same in neurons from normoxic (6.9+/-0.5 pF, n=16) and hypoxic (6.3+/-0.5 pF, n=15) rats. The EC(50) for peak GABA-evoked current density was significantly greater in neurons from hypoxic (21.7+/-2.2 microM) compared to normoxic rats (12.2+/-0.9 microM) (p<0.001). Peak and 5-s adapted GABA currents evoked by 1, 3 and 10 microM were greater in neurons from normoxic compared to hypoxic rats (p<0.05) whereas peak and 5-s adapted responses to 30 and 100 microM GABA were not different comparing normoxic to hypoxic rats. Desensitization of GABA(A)-evoked currents was observed at concentrations greater than 3 microM and, measured as the ratio of the current 5 s after the onset of 100 microM GABA application to the peak GABA current, was the same in neurons from normoxic (0.37+/-0.03) and hypoxic rats (0.33+/-0.04). Reduced sensitivity to GABA(A) receptor-evoked inhibition in chronic hypoxia could influence chemoreceptor afferent integration by NTS neurons.

Enhancement of extinction memory consolidation: the role of the noradrenergic and GABAergic systems within the basolateral amygdala.

PubMed

Berlau, Daniel J; McGaugh, James L

2006-09-01

Evidence from previous studies indicates that the noradrenergic and GABAergic influences within the basolateral amygdala (BLA) modulate the consolidation of memory for fear conditioning. The present experiments investigated whether the same modulatory influences are involved in regulating the extinction of fear-based learning. To investigate this issue, male Sprague Dawley rats implanted with unilateral or bilateral cannula aimed at the BLA were trained on a contextual fear conditioning (CFC) task and 24 and 48 h later were given extinction training. Immediately following each extinction session they received intra-BLA infusions of the GABAergic antagonist bicuculline (50 ng), the beta-adrenocepter antagonist propranolol (500 ng), bicuculline with propranolol, norepinephrine (NE) (0.3, 1.0, and 3.0 microg), the GABAergic agonist muscimol (125 ng), NE with muscimol or a control solution. To investigate the involvement of the dorsal hippocampus (DH) as a possible target of BLA activation during extinction, other animals were given infusions of muscimol (500 ng) via an ipsilateral cannula implanted in the DH. Bilateral BLA infusions of bicuculline significantly enhanced extinction, as did infusions into the right, but not left BLA. Propranolol infused into the right BLA together with bicuculline blocked the bicuculline-induced memory enhancement. Norepinephrine infused into the right BLA also enhanced extinction, and this effect was not blocked by co-infusions of muscimol. Additionally, muscimol infused into the DH did not attenuate the memory enhancing effects of norepinephrine infused into the BLA. These findings provide evidence that, as with original CFC learning, noradrenergic activation within the BLA modulates the consolidation of CFC extinction. The findings also suggest that the BLA influence on extinction is not mediated by an interaction with the dorsal hippocampus.

Expression of specific ionotropic glutamate and GABA-A receptor subunits is decreased in central amygdala of alcoholics.

PubMed

Jin, Zhe; Bhandage, Amol K; Bazov, Igor; Kononenko, Olga; Bakalkin, Georgy; Korpi, Esa R; Birnir, Bryndis

2014-01-01

The central amygdala (CeA) has a role for mediating fear and anxiety responses. It is also involved in emotional imbalance caused by alcohol abuse and dependence and in regulating relapse to alcohol abuse. Growing evidences suggest that excitatory glutamatergic and inhibitory γ-aminobutyric acid-ergic (GABAergic) transmissions in the CeA are affected by chronic alcohol exposure. Human post-mortem CeA samples from male alcoholics (n = 9) and matched controls (n = 9) were assayed for the expression level of ionotropic glutamate and GABA-A receptors subunit mRNAs using quantitative real-time reverse transcription-PCR (RT-qPCR). Our data revealed that out of the 16 ionotropic glutamate receptor subunits, mRNAs encoding two AMPA [2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid] receptor subunits GluA1 and GluA4; one kainate receptor subunit GluK2; one NMDA (N-methyl-D-aspartate) receptor subunit GluN2D and one delta receptor subunit GluD2 were significantly decreased in the CeA of alcoholics. In contrast, of the 19 GABA-A receptor subunits, only the mRNA encoding the α2 subunit was significantly down-regulated in the CeA of the alcoholics as compared with control subjects. Our findings imply that the down-regulation of specific ionotropic glutamate and GABA-A receptor subunits in the CeA of alcoholics may represent one of the molecular substrates underlying the new balance between excitatory and inhibitory neurotransmission in alcohol dependence.

Expression of specific ionotropic glutamate and GABA-A receptor subunits is decreased in central amygdala of alcoholics

PubMed Central

Jin, Zhe; Bhandage, Amol K.; Bazov, Igor; Kononenko, Olga; Bakalkin, Georgy; Korpi, Esa R.; Birnir, Bryndis

2014-01-01

The central amygdala (CeA) has a role for mediating fear and anxiety responses. It is also involved in emotional imbalance caused by alcohol abuse and dependence and in regulating relapse to alcohol abuse. Growing evidences suggest that excitatory glutamatergic and inhibitory γ-aminobutyric acid-ergic (GABAergic) transmissions in the CeA are affected by chronic alcohol exposure. Human post-mortem CeA samples from male alcoholics (n = 9) and matched controls (n = 9) were assayed for the expression level of ionotropic glutamate and GABA-A receptors subunit mRNAs using quantitative real-time reverse transcription-PCR (RT-qPCR). Our data revealed that out of the 16 ionotropic glutamate receptor subunits, mRNAs encoding two AMPA [2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid] receptor subunits GluA1 and GluA4; one kainate receptor subunit GluK2; one NMDA (N-methyl-D-aspartate) receptor subunit GluN2D and one delta receptor subunit GluD2 were significantly decreased in the CeA of alcoholics. In contrast, of the 19 GABA-A receptor subunits, only the mRNA encoding the α2 subunit was significantly down-regulated in the CeA of the alcoholics as compared with control subjects. Our findings imply that the down-regulation of specific ionotropic glutamate and GABA-A receptor subunits in the CeA of alcoholics may represent one of the molecular substrates underlying the new balance between excitatory and inhibitory neurotransmission in alcohol dependence. PMID:25278838

The epileptogenic spectrum of opiate agonists.

PubMed

Snead, O C; Bearden, L J

1982-11-01

The present authors gave mu, delta, kappa, epsilon and sigma opiate receptor agonists intracerebroventricularly to rats both singly and in combination while monitoring the electroencephalogram from cortical and depth electrodes. Dose-response curves were plotted with naloxone against the changes produced by each agonist, and the effect of a number of anticonvulsant drugs on agonist-induced seizures was ascertained. Each opiate agonist produced a different seizure pattern with a different naloxone dose-response curve and anticonvulsant profile. The order of convulsive potency was epsilon greater than delta greater than mu greater than sigma much greater than kappa. Petit mal-like seizure activity was unique to the delta agonist, leucine-enkephalin, while only the mu agonist, morphine produced generalized convulsive seizures. These experiments raise the possibility that opiate systems in the brain may be involved in the pathogenesis of a wide spectrum of seizure disorders.

Human α1β3γ2L gamma-aminobutyric acid type A receptors: High-level production and purification in a functional state.

PubMed

Dostalova, Zuzana; Zhou, Xiaojuan; Liu, Aiping; Zhang, Xi; Zhang, Yinghui; Desai, Rooma; Forman, Stuart A; Miller, Keith W

2014-02-01

Gamma-aminobutyric acid type A receptors (GABA(A)Rs) are the most important inhibitory chloride ion channels in the central nervous system and are major targets for a wide variety of drugs. The subunit compositions of GABA(A)Rs determine their function and pharmacological profile. GABAA Rs are heteropentamers of subunits, and (α1)2 (β3)2 (γ2L)1 is a common subtype. Biochemical and biophysical studies of GABA(A)Rs require larger quantities of receptors of defined subunit composition than are currently available. We previously reported high-level production of active human α1β3 GABA(A)R using tetracycline-inducible stable HEK293 cells. Here we extend the strategy to receptors containing three different subunits. We constructed a stable tetracycline-inducible HEK293-TetR cell line expressing human (N)-FLAG-α1β3γ2L-(C)-(GGS)3 GK-1D4 GABA(A)R. These cells achieved expression levels of 70-90 pmol [(3)H]muscimol binding sites/15-cm plate at a specific activity of 15-30 pmol/mg of membrane protein. Incorporation of the γ2 subunit was confirmed by the ratio of [(3)H]flunitrazepam to [(3)H]muscimol binding sites and sensitivity of GABA-induced currents to benzodiazepines and zinc. The α1β3γ2L GABA(A)Rs were solubilized in dodecyl-D-maltoside, purified by anti-FLAG affinity chromatography and reconstituted in CHAPS/asolectin at an overall yield of ∼ 30%. Typical purifications yielded 1.0-1.5 nmoles of [(3)H]muscimol binding sites/60 plates. Receptors with similar properties could be purified by 1D4 affinity chromatography with lower overall yield. The composition of the purified, reconstituted receptors was confirmed by ligand binding, Western blot, and proteomics. Allosteric interactions between etomidate and [(3)H]muscimol binding were maintained in the purified state. © 2013 The Protein Society.

GABA(A) receptor modulation during adolescence alters adult ethanol intake and preference in rats.

PubMed

Hulin, Mary W; Amato, Russell J; Winsauer, Peter J

2012-02-01

To address the hypothesis that GABA(A) receptor modulation during adolescence may alter the abuse liability of ethanol during adulthood, the effects of adolescent administration of both a positive and negative GABA(A) receptor modulator on adult alcohol intake and preference were assessed. Three groups of adolescent male rats received 12 injections of lorazepam (3.2 mg/kg), dehydroepiandrosterone (DHEA, 56 mg/kg), or vehicle on alternate days starting on postnatal day (PD) 35. After this time, the doses were increased to 5.6 and 100 mg/kg, respectively, for 3 more injections on alternate days. Subjects had access to 25 to 30 g of food daily, during the period of the first 6 injections, and 18 to 20 g thereafter. Food intake of each group was measured 60 minutes after food presentation, which occurred immediately after drug administration on injection days or at the same time of day on noninjection days. When subjects reached adulthood (PD 88), ethanol preference was determined on 2 separate occasions, an initial 3-day period and a 12-day period, in which increasing concentrations of ethanol were presented. During each preference test, intake of water, saccharin, and an ethanol/saccharin solution was measured after each 23-hour access period. During adolescence, lorazepam increased 60-minute food intake, and this effect was enhanced under the more restrictive feeding schedule. DHEA had the opposite effect on injection days, decreasing food intake compared with noninjection days. In adulthood, the lorazepam-treated group preferred the 2 lowest concentrations of ethanol/saccharin more than saccharin alone compared with vehicle-treated subjects, which showed no preference for any concentration of ethanol/saccharin over saccharin. DHEA-treated subjects showed no preference among the 3 solutions. These data demonstrate that GABA(A) receptor modulation during adolescence can alter intake and preference for ethanol in adulthood and highlights the importance of drug history

Region-specific roles of the prelimbic cortex, the dorsal CA1, the ventral DG and ventral CA1 of the hippocampus in the fear return evoked by a sub-conditioning procedure in rats.

PubMed

Fu, Juan; Xing, Xiaoli; Han, Mengfi; Xu, Na; Piao, Chengji; Zhang, Yue; Zheng, Xigeng

2016-02-01

The return of learned fear is an important issue in anxiety disorder research since an analogous process may contribute to long-term fear maintenance or clinical relapse. A number of studies demonstrate that mPFC and hippocampus are important in the modulation of post-extinction re-expression of fear memory. However, the region-specific role of these structures in the fear return evoked by a sub-threshold conditioning (SC) is not known. In the present experiments, we first examined specific roles of the prelimbic cortex (PL), the dorsal hippocampus (DH, the dorsal CA1 area in particular), the ventral hippocampus (the ventral dentate gyrus (vDG) and the ventral CA1 area in particular) in this fear return process. Then we examined the role of connections between PL and vCA1 with this behavioral approach. Rats were subjected to five tone-shock pairings (1.0-mA shock) to induce conditioned fear (freezing), followed by three fear extinction sessions (25 tone-alone trials each session). After a post-test for extinction memory, some rats were retrained with the SC procedure to reinstate tone-evoked freezing. Rat groups were injected with low doses of the GABAA agonist muscimol to selectively inactivate PL, DH, vDG, or vCA1 120 min before the fear return test. A disconnection paradigm with ipsilateral or contralateral muscimol injection of the PL and the vCA1 was used to examine the role of this pathway in the fear return. We found that transient inactivation of these areas significantly impaired fear return (freezing): inactivation of the prelimbic cortex blocked SC-evoked fear return in particular but did not influence fear expression in general; inactivation of the DH area impaired fear return, but had no effect on the extinction retrieval process; both ventral DG and ventral CA1 are required for the return of extinguished fear whereas only ventral DG is required for the extinction retrieval. These findings suggest that PL, DH, vDG, and vCA1 all contribute to the fear

Proton sensitivity of rat cerebellar granule cell GABAA receptors: dependence on neuronal development

PubMed Central

Krishek, Belinda J; Smart, Trevor G

2001-01-01

The effect of GABAA receptor development in culture on the modulation of GABA-induced currents by external H+ was examined in cerebellar granule cells using whole-cell and single-channel recording. Equilibrium concentration-response curves revealed a lower potency for GABA between 11 and 12 days in vitro (DIV) resulting in a shift of the EC50 from 10.7 to 2.4 μM. For granule cells before 11 DIV, the peak GABA-activated current was inhibited at low external pH and enhanced at high pH with a pKa of 6.65. For the steady-state response, low pH was inhibitory with a pKa of 5.56. After 11 DIV, the peak GABA-activated current was largely pH insensitive; however, the steady-state current was potentiated at low pH with a pKa of 6.84. Single GABA-activated ion channels were recorded from outside-out patches of granule cell bodies. At pH 5.4-9.4, single GABA channels exhibited multiple conductance states occurring at 22-26, 16-17 and 12-14 pS. The conductance levels were not significantly altered over the time period of study, nor by changing the external H+ concentration. Two exponential functions were required to fit the open-time frequency histograms at both early (< 11 DIV) and late (> 11 DIV) development times at each H+ concentration. The short and long open time constants were unaffected either by the extracellular H+ concentration or by neuronal development. The distribution of all shut times was fitted by the sum of three exponentials designated as short, intermediate and long. At acidic pH, the long shut time constant decreased with development as did the relative contribution of these components to the overall distribution. This was concurrent with an increase in the mean probability of channel opening. In conclusion, this study demonstrates in cerebellar granule cells that external pH can either reduce, have no effect on, or enhance GABA-activated responses depending on the stage of development, possibly related to the subunit composition of the GABAA receptors

Evaluation of partial beta-adrenoceptor agonist activity.

PubMed

Lipworth, B J; Grove, A

1997-01-01

A partial beta-adrenoceptor (beta-AR) agonist will exhibit opposite agonist and antagonist activity depending on the prevailing degree of adrenergic tone or the presence of a beta-AR agonist with higher intrinsic activity. In vivo partial beta-AR agonist activity will be evident at rest with low endogenous adrenergic tone, as for example with chronotropicity (beta 1/beta 2), inotropicity (beta 1) or peripheral vasodilatation and finger tremor (beta 2). beta-AR blocking drugs which have partial agonist activity may exhibit a better therapeutic profile when used for hypertension because of maintained cardiac output without increased systemic vascular resistance, along with an improved lipid profile. In the presence of raised endogenous adrenergic tone such as exercise or an exogenous full agonist, beta-AR subtype antagonist activity will become evident in terms of effects on exercise induced heart rate (beta 1) and potassium (beta 2) responses. Reduction of exercise heart rate will occur to a lesser degree in the case of a beta-adrenoceptor blocker with partial beta 1-AR agonist activity compared with a beta-adrenoceptor blocker devoid of partial agonist activity. This may result in reduced therapeutic efficacy in the treatment of angina on effort when using beta-AR blocking drugs with partial beta 1-AR agonist activity. Effects on exercise hyperkalaemia are determined by the balance between beta 2-AR partial agonist activity and endogenous adrenergic activity. For predominantly beta 2-AR agonist such as salmeterol and salbutamol, potentiation of exercise hyperkalaemia occurs. For predominantly beta 2-AR antagonists such as carteolol, either potentiation or attenuation of exercise hyperkalaemia occurs at low and high doses respectively. beta 2-AR partial agonist activity may also be expressed as antagonism in the presence of an exogenous full agonist, as for example attenuation of fenoterol induced responses by salmeterol. Studies are required to investigate whether

Effects of GABAA receptor inhibition on response properties of barrel cortical neurons in C-fiber-depleted rats.

PubMed

Farazifard, Rasoul; Kiani, Roozbeh; Esteky, Hossein

2005-07-19

C-fiber depletion results in expansion of low threshold somatosensory mechanoreceptive fields. In this study, we investigated the role of intact C-fibers in GABAA-mediated inhibition in barrel cortical neurons. We used electronically controlled mechanical stimulation of whiskers to quantitatively examine the responses of barrel cells to whisker displacements. After systemic injection of picrotoxin neuronal responses were recorded at 5 min intervals for 20 min and then at 10 min intervals for 100 min. Picrotoxin injection caused a 3-fold increase in response magnitude of adjacent whisker stimulation and 1.4-fold increase in response magnitude of principal whisker stimulation with a maximum enhancement 50 min after the injection. There was no significant change in spontaneous activity following picrotoxin injection. The response enhancement and receptive field expansion observed in normal rats were completely absent in the C-fiber-depleted rats. These results suggest that the GABAA-mediated inhibition that modulates the receptive field functional organization of the barrel cortex depends on intact C-fibers.

Perimenstrual-Like Hormonal Regulation of Extrasynaptic δ-Containing GABAA Receptors Mediating Tonic Inhibition and Neurosteroid Sensitivity

PubMed Central

Carver, Chase Matthew; Wu, Xin; Gangisetty, Omkaram

2014-01-01

Neurosteroids are endogenous regulators of neuronal excitability and seizure susceptibility. Neurosteroids, such as allopregnanolone (AP; 3α-hydroxy-5α-pregnan-20-one), exhibit enhanced anticonvulsant activity in perimenstrual catamenial epilepsy, a neuroendocrine condition in which seizures are clustered around the menstrual period associated with neurosteroid withdrawal (NSW). However, the molecular mechanisms underlying such enhanced neurosteroid sensitivity remain unclear. Neurosteroids are allosteric modulators of both synaptic (αβγ2-containing) and extrasynaptic (αβδ-containing) GABAA receptors, but they display greater sensitivity toward δ-subunit receptors in dentate gyrus granule cells (DGGCs). Here we report a novel plasticity of extrasynaptic δ-containing GABAA receptors in the dentate gyrus in a mouse perimenstrual-like model of NSW. In molecular and immunofluorescence studies, a significant increase occurred in δ subunits, but not α1, α2, β2, and γ2 subunits, in the dentate gyrus of NSW mice. Electrophysiological studies confirmed enhanced sensitivity to AP potentiation of GABA-gated currents in DGGCs, but not in CA1 pyramidal cells, in NSW animals. AP produced a greater potentiation of tonic currents in DGGCs of NSW animals, and such enhanced AP sensitivity was not evident in δ-subunit knock-out mice subjected to a similar withdrawal paradigm. In behavioral studies, mice undergoing NSW exhibited enhanced seizure susceptibility to hippocampus kindling. AP has enhanced anticonvulsant effects in fully kindled wild-type mice, but not δ-subunit knock-out mice, undergoing NSW-induced seizures, confirming δ-linked neurosteroid sensitivity. These results indicate that perimenstrual NSW is associated with striking upregulation of extrasynaptic, δ-containing GABAA receptors that mediate tonic inhibition and neurosteroid sensitivity in the dentate gyrus. These findings may represent a molecular rationale for neurosteroid therapy of catamenial

GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

PubMed

Reddy, Doodipala Samba

2018-01-01

Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

Differential agonist and inverse agonist profile of antipsychotics at D2L receptors coupled to GIRK potassium channels.

PubMed

Heusler, Peter; Newman-Tancredi, Adrian; Castro-Fernandez, Annabelle; Cussac, Didier

2007-03-01

The D(2) dopaminergic receptor represents a major target of antipsychotic drugs. Using the coupling of the human D(2long) (hD(2L)) receptor to G protein-coupled inward rectifier potassium (GIRK) channels in Xenopus laevis oocytes, we examined the activity of antipsychotic agents of different classes - typical, atypical, and a "new generation" of compounds, exhibiting a preferential D(2) and 5-HT(1A) receptor profile. When the hD(2L) receptor was coexpressed with GIRK channels, a series of reference compounds exhibited full agonist (dopamine, and quinpirole), partial agonist (apomorphine, (-)3-PPP, and (+)-UH232) or inverse agonist (raclopride, and L741626) properties. Sarizotan exhibited only very weak partial agonist action. At higher levels of receptor cRNA injected per oocyte, both partial agonist activity and inverse agonist properties were generally more pronounced. The inverse agonist action of L741626 was reversed by interaction with sarizotan, thus confirming the constitutive activity of wild-type hD(2L) receptors in the oocyte expression system. When antipsychotic agents were tested for their actions at the hD(2L) receptor, typical (haloperidol) as well as atypical (nemonapride, ziprasidone, and clozapine) compounds acted as inverse agonists. In contrast, among D(2)/5-HT(1A) antipsychotics, only SLV313 and F15063 behaved as inverse agonists, whilst the other members of this group (bifeprunox, SSR181507 and the recently marketed antipsychotic, aripiprazole) exhibited partial agonist properties. Thus, the X. laevis oocyte expression system highlights markedly different activity of antipsychotics at the hD(2L) receptor. These differential properties may translate to distinct therapeutic potential of these compounds.

Acute sleep deprivation preconditions the heart against ischemia/ reperfusion injury: the role of central GABA-A receptors

PubMed Central

Parsa, Hoda; Imani, Alireza; Faghihi, Mahdieh; Riahi, Esmail; Badavi, Mohammad; Shakoori, Abbas; Rastegar, Tayebeh; Aghajani, Marjan; Rajani, Sulail Fatima

2017-01-01

Objective(s): Central γ-aminobutyric acid (GABA) neurotransmission modulates cardiovascular functions and sleep. Acute sleep deprivation (ASD) affects functions of various body organs via different mechanisms. Here, we evaluated the effect of ASD on cardiac ischemia/reperfusion injury (IRI), and studied the role of GABA-A receptor inhibition in central nucleus of amygdala (CeA) by assessing nitric oxide (NO) and oxidative stress. Materials and Methods: The CeA in sixty male Wistar rats was cannulated for saline or bicuculline (GABA-A receptor antagonist) administration. All animals underwent 30 min of coronary occlusion (ischemia), followed by 2 hr reperfusion (IR). The five experimental groups (n=12) included are as follows: IR: received saline; BIC+IR: received Bicuculline; MLP+IR: received saline, followed by the placement of animals in an aquarium with multiple large platforms; ASD+IR: underwent ASD in an aquarium with multiple small platforms; and BIC+ASD+IR: received bicuculline prior to ASD. Results: Bicuculline administration increased the malondialdehyde levels and infarct size, and decreased the NO metabolites levels and endothelial nitric oxide synthase (eNOS) gene expression in infarcted and non-infarcted areas in comparison to IR group. ASD reduced malondialdehyde levels and infarct size and increased NO metabolites, corticosterone levels and eNOS expression in infarcted and non-infarcted areas as compared to the IR group. Levels of malondialdehyde were increased while levels of NO metabolites, corticosterone and eNOS expression in infarcted and non-infarcted areas were reduced in the BIC+ASD+IR as compared to the ASD+IR group. Conclusion: Blockade of GABA-A receptors in the CeA abolishes ASD-induced cardioprotection by suppressing oxidative stress and NO production. PMID:29299201

Acute sleep deprivation preconditions the heart against ischemia/ reperfusion injury: the role of central GABA-A receptors.

PubMed

Parsa, Hoda; Imani, Alireza; Faghihi, Mahdieh; Riahi, Esmail; Badavi, Mohammad; Shakoori, Abbas; Rastegar, Tayebeh; Aghajani, Marjan; Rajani, Sulail Fatima

2017-11-01

Central γ-aminobutyric acid (GABA) neurotransmission modulates cardiovascular functions and sleep. Acute sleep deprivation (ASD) affects functions of various body organs via different mechanisms. Here, we evaluated the effect of ASD on cardiac ischemia/reperfusion injury (IRI), and studied the role of GABA-A receptor inhibition in central nucleus of amygdala (CeA) by assessing nitric oxide (NO) and oxidative stress. The CeA in sixty male Wistar rats was cannulated for saline or bicuculline (GABA-A receptor antagonist) administration. All animals underwent 30 min of coronary occlusion (ischemia), followed by 2 hr reperfusion (IR). The five experimental groups (n=12) included are as follows: IR: received saline; BIC+IR: received Bicuculline; MLP+IR: received saline, followed by the placement of animals in an aquarium with multiple large platforms; ASD+IR: underwent ASD in an aquarium with multiple small platforms; and BIC+ASD+IR: received bicuculline prior to ASD. Bicuculline administration increased the malondialdehyde levels and infarct size, and decreased the NO metabolites levels and endothelial nitric oxide synthase (eNOS) gene expression in infarcted and non-infarcted areas in comparison to IR group. ASD reduced malondialdehyde levels and infarct size and increased NO metabolites, corticosterone levels and eNOS expression in infarcted and non-infarcted areas as compared to the IR group. Levels of malondialdehyde were increased while levels of NO metabolites, corticosterone and eNOS expression in infarcted and non-infarcted areas were reduced in the BIC+ASD+IR as compared to the ASD+IR group. Blockade of GABA-A receptors in the CeA abolishes ASD-induced cardioprotection by suppressing oxidative stress and NO production.

Modulation of spinal nociception by GluR5 kainate receptor ligands in acute and hyperalgesic states and the role of gabaergic mechanisms.

PubMed

Mascias, Paula; Scheede, Manuela; Bloms-Funke, Petra; Chizh, Boris

2002-09-01

GluR5 receptors modulate spinal nociception, however, their role in nociceptive hypersensitivity remains unclear. Using behavioural and electrophysiological approaches, we have investigated several GluR5 ligands in acute and hyperalgesic states. Furthermore, as the GABAergic system plays a role in GluR5 mediated effects in the brain, we also analysed the interaction between GluR5 agonists and GABA(A) antagonists in the spinal cord. In young rats in vivo, the GluR5 selective agonist ATPA was antinociceptive and antihyperalgesic in a model of inflammatory hyperalgesia (ED(50) approximately 4.6 and approximately 5.2 mg/kg, respectively), whereas the GluR5/GluR6 agonist SYM2081 was only antihyperalgesic. ATPA, but not SYM2081, was also able to inhibit nociceptive motoneurone responses in anaesthetised adult rats after intrathecal administration. In hemisected spinal cords in vitro, SYM2081 was inactive, whereas ATPA and another GluR5 agonist, (S)-5-iodowillardiine, inhibited nociceptive reflexes (EC(50) 1.1+/-0.4 micro M and 0.36+/-0.05 micro M, respectively). Both GluR5 agonists also inhibited motoneurone responses to repetitive dorsal root stimulation and their cumulative depolarisation, a correlate of wind-up. The GABA(A) antagonists bicuculline (10 micro M) and SR95531 (1 micro M) enhanced polysynaptic responses to single stimuli but abolished the cumulative depolarisation. Both bicuculline and SR95531 significantly attenuated the inhibition of nociceptive responses by 1 micro M ATPA (by approximately 50%). We conclude that selective GluR5 kainate receptor activation inhibits spinal nociception and its sensitisation caused by ongoing peripheral nociceptive drive. GABA(A) receptors are involved in tonic inhibition of segmental responses, but contribute to their sensitisation by repetitive primary afferent stimulation. Furthermore, there is a cross-talk between the two systems, presumably due to GluR5-mediated activation of GABAergic inhibitory interneurones in the

Differential effects of chronic lorazepam and alprazolam on benzodiazepine binding and GABAA-receptor function.

PubMed Central

Galpern, W. R.; Miller, L. G.; Greenblatt, D. J.; Shader, R. I.

1990-01-01

1. Chronic benzodiazepine administration has been associated with tolerance and with downregulation of gamma-aminobutyric acidA (GABAA)-receptor binding and function. However, effects of individual benzodiazepines on brain regions have varied. 2. To compare the effects of chronic lorazepam and alprazolam, we have administered these drugs to mice for 1 and 7 days (2 mg kg-1 day-1) and determined benzodiazepine receptor binding in vivo with and without administration of CL 218,872, 25 mg kg-1 i.p., and GABA-dependent chloride uptake in 3 brain regions at these time points. 3. Benzodiazepine binding was decreased in the cortex and hippocampus at day 7 compared to day 1 of lorazepam, with an increase in CL 218,872-resistant (Type 2) sites in both regions. Maximal GABA-dependent chloride uptake was also decreased in the cortex and hippocampus at day 7. 4. Binding was decreased only in the cortex after 7 days of alprazolam, with no significant change in Type 2 binding. Maximal GABA-dependent chloride uptake was also decreased only in the cortex. 5. These data suggest that the effects of chronic benzodiazepine administration on the GABAA-receptor may be both region-specific and receptor subtype-specific. PMID:1964820

Prodepressant- and anxiogenic-like effects of serotonin-selective, but not noradrenaline-selective, antidepressant agents in mice lacking α2-containing GABAA receptors.

PubMed

Benham, Rebecca S; Hewage, Nishani B; Suckow, Raymond F; Engin, Elif; Rudolph, Uwe

2017-08-14

Deficits in neuronal inhibition via gamma-aminobutyric acid (GABA) type A receptors (GABAA-Rs) are implicated in the pathophysiology of major depressive disorder and the therapeutic effects of current antidepressant treatments, however, the relevant GABAA-R subtype as defined by its alpha subunit is still unknown. We previously reported anxiety- and depressive-like behavior in alpha2+/- and alpha2-/- mice, respectively (Vollenweider, 2011). We sought to determine whether this phenotype could be reversed by chronic antidepressant treatment. Adult male mice received 4 or 8mg/kg fluoxetine or 53mg/kg desipramine in their drinking water for four weeks before undergoing behavioral testing. In the novelty suppressed feeding test, desipramine had anxiolytic-like effects reducing the latencies to bite and to eat the pellet in both wild-type and alpha2+/- mice. Surprisingly, 4mg/kg fluoxetine had anxiogenic-like effects in alpha2+/- mice increasing latency to bite and to eat while 8mg/kg fluoxetine increased the latency to eat in both wild-type and alpha2+/- mice. In the forced swim and tail suspension tests, chronic desipramine treatment increased latency to immobility in wild-type and alpha2-/- mice. In contrast, chronic fluoxetine treatment increased immobility in alpha2-/- mice in both tasks while generally having no effect in wild-type mice. These findings suggest that in preclinical paradigms of anxiety and behavioral despair the antidepressant-like effects of desipramine are independent of alpha2-containing GABAA-Rs, while a reduction in alpha2 expression leads to an increased sensitivity to anxiogenic- and prodepressant-like effects with chronic fluoxetine treatment, pointing to a potential role of alpha2-containing GABAA-Rs in the response to serotonin-selective antidepressants. Copyright © 2017 Elsevier B.V. All rights reserved.

Encephalitis with refractory seizures, status epilepticus, and antibodies to the GABAA receptor: a case series, characterisation of the antigen, and analysis of the effects of antibodies.

PubMed

Petit-Pedrol, Mar; Armangue, Thaís; Peng, Xiaoyu; Bataller, Luis; Cellucci, Tania; Davis, Rebecca; McCracken, Lindsey; Martinez-Hernandez, Eugenia; Mason, Warren P; Kruer, Michael C; Ritacco, David G; Grisold, Wolfgang; Meaney, Brandon F; Alcalá, Carmen; Sillevis-Smitt, Peter; Titulaer, Maarten J; Balice-Gordon, Rita; Graus, Francesc; Dalmau, Josep

2014-03-01

Increasing evidence suggests that seizures and status epilepticus can be immune-mediated. We aimed to describe the clinical features of a new epileptic disorder, and to establish the target antigen and the effects of patients' antibodies on neuronal cultures. In this observational study, we selected serum and CSF samples for antigen characterisation from 140 patients with encephalitis, seizures or status epilepticus, and antibodies to unknown neuropil antigens. The samples were obtained from worldwide referrals of patients with disorders suspected to be autoimmune between April 28, 2006, and April 25, 2013. We used samples from 75 healthy individuals and 416 patients with a range of neurological diseases as controls. We assessed the samples using immunoprecipitation, mass spectrometry, cell-based assay, and analysis of antibody effects in cultured rat hippocampal neurons with confocal microscopy. Neuronal cell-membrane immunoprecipitation with serum of two index patients revealed GABAA receptor sequences. Cell-based assay with HEK293 expressing α1/β3 subunits of the GABAA receptor showed high titre serum antibodies (>1:160) and CSF antibodies in six patients. All six patients (age 3-63 years, median 22 years; five male patients) developed refractory status epilepticus or epilepsia partialis continua along with extensive cortical-subcortical MRI abnormalities; four patients needed pharmacologically induced coma. 12 of 416 control patients with other diseases, but none of the healthy controls, had low-titre GABAA receptor antibodies detectable in only serum samples, five of them also had GAD-65 antibodies. These 12 patients (age 2-74 years, median 26.5 years; seven male patients) developed a broader spectrum of symptoms probably indicative of coexisting autoimmune disorders: six had encephalitis with seizures (one with status epilepticus needing pharmacologically induced coma; one with epilepsia partialis continua), four had stiff-person syndrome (one with seizures

Anticonflict effect of alpidem as compared with the benzodiazepine alprazolam in rats.

PubMed

Hascoët, M; Bourin, M

1997-02-01

A comparative study between two drugs acting on the GABAA receptor, alprazolam and alpidem was undertaken, using simple tests such as measurement of spontaneous locomotor activity, four plates test and rotarod in mice. Additional conflict test was further performed using a new conflict paradigm where the opportunity existed for rats to choose during punished periods between immediate, punished reinforcement and delayed non-punished reinforcement. The benzodiazepine alprazolam, demonstrated, as expected, strong anxiolytic effects in mice and increased punished response in rats at non sedative doses (0.5, 1 mg/kg). High doses of alprazolam decreased spontaneous locomotor activity and induced myorelaxant effects in mice. Alpidem, an imidazopyridine derivative, induced motor impairment in mice and only very weak anxiolytic effects in the four plates test in mice (4 mg/kg) and in punished procedure in rats (32 mg/kg). As alprazolam is a full agonist for the GABAA receptor complex and alpidem is a partial agonist acting with specificity on omega 1 GABAA receptor subtypes, the results are discussed. Activity on omega 1 receptor subtypes is perhaps not sufficient in order to obtain a true anti-conflict effect and compounds such as alpidem only relieve some of the symptoms of anxiety disorders.

[Dopamine agonists--clinical applications beyond Parkinson's disease].

PubMed

Kuran, Włodzimierz

2007-01-01

Contemporary experience and results of clinical trials concerning dopamine agonist application in the treatment of many different diseases (apart from Parkinson's disease) are presented in the paper. A basic clinical recommendation for agonists is restless legs syndrome. In this syndrome almost all agonists give a considerable subjective and objective improvement. Treatment of atypical parkinsonism (MSA, PSP, CBD) in the majority of patients is ineffective. The author also presents promising results of treatment with agonists in such diverse diseases as hyperkinetic syndromes, cocaine dependence, drug-resistant depression and erectile dysfunction (apomorphine). Dopamine partial agonists (e.g. aripiprazol) are recommended in the modern treatment of schizophrenia.

Caudal Nucleus Accumbens Core Is Critical in the Regulation of Cue-Elicited Approach-Avoidance Decisions

PubMed Central

Hamel, Laurie; Thangarasa, Tharshika; Samadi, Osai

2017-01-01

The nucleus accumbens (NAc) is thought to be a site of integration of positively and negatively valenced information and action selection. Functional differentiation in valence processing has previously been found along the rostrocaudal axis of the shell region of the NAc in assessments of unconditioned motivation. Given that the core region of the NAc has been implicated in the elicitation of motivated behavior in response to conditioned cues, we sought to assess the role of caudal, intermediate, and rostral sites within this subregion in cue-elicited approach-avoidance decisions. Rats were trained to associate visuo-tactile cues with appetitive, aversive, and neutral outcomes. Following the successful acquisition of the cue-outcome associations, rats received microinfusions of GABAA and GABAB receptor agonists (muscimol/baclofen) or saline into the caudal, intermediate, or rostral NAc core and were then exposed to a superimposition of appetitively and aversively valenced cues versus neutral cues in a “conflict test,” as well as to the appetitive versus neutral cues, and aversive cues versus neutral cues, in separate conditioned preference/avoidance tests. Disruption of activity in the intermediate to caudal parts of the NAc core resulted in a robust avoidance bias in response to motivationally conflicting cues, as well as a potentiated avoidance of aversive cues as compared with control animals, coupled with an attenuated conditioned preference for the appetitive cue. These results suggest that the caudal NAc core may have the capacity to exert bidirectional control over appetitively and aversively motivated responses to valence signals. PMID:28275709

Excitatory amino acid receptors in the dorsomedial hypothalamus mediate prostaglandin-evoked thermogenesis in brown adipose tissue.

PubMed

Madden, C J; Morrison, S F

2004-02-01

We determined whether the dorsomedial hypothalamus (DMH) plays a role in the thermogenic, metabolic, and cardiovascular effects evoked by centrally administered PGE2. Microinjection of PGE2 (170 pmol/60 nl) into the medial preoptic area of the hypothalamus in urethane-chloralose-anesthetized, artificially ventilated rats increased brown adipose tissue (BAT) sympathetic nerve activity (SNA; +207 +/- 18% of control), BAT temperature (1.5 +/- 0.2 degrees C), expired CO2 (0.9 +/- 0.1%), heart rate (HR; 106 +/- 12 beats/min), and mean arterial pressure (22 +/- 4 mmHg). Within 5 min of subsequent bilateral microinjections of the GABAA receptor agonist muscimol (120 pmol.60 nl-1.side-1) or the ionotropic excitatory amino acid antagonist kynurenate (6 nmol.60 nl-1.side-1) into the DMH, the PGE2-evoked increases were, respectively, attenuated by 91 +/- 3% and 108 +/- 7% for BAT SNA, by 73 +/- 12% and 102 +/- 28% for BAT temperature, by 100 +/- 4% and 125 +/- 21% for expired CO2, by 72 +/- 11% and 70 +/- 16% for HR, and by 84 +/- 19% and 113 +/- 16% for mean arterial pressure. Microinjections outside the DMH within the dorsal hypothalamic area adjacent to the mamillothalamic tracts or within the ventromedial hypothalamus were less effective for attenuating the PGE2-evoked thermogenic, metabolic, and cardiovascular responses. These results demonstrate that activation of excitatory amino acid receptors within the DMH is necessary for the thermogenic, metabolic, and cardiovascular responses evoked by microinjection of PGE2 into the medial preoptic area.

The Role of the Medial Prefrontal Cortex in Regulating Social Familiarity-Induced Anxiolysis

PubMed Central

Lungwitz, Elizabeth A; Stuber, Garret D; Johnson, Philip L; Dietrich, Amy D; Schartz, Nicole; Hanrahan, Brian; Shekhar, Anantha; Truitt, William A

2014-01-01

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. PMID:24157502

The nucleus raphe magnus OFF-cells are involved in diffuse noxious inhibitory controls.

PubMed

Chebbi, R; Boyer, N; Monconduit, L; Artola, A; Luccarini, P; Dallel, R

2014-06-01

Diffuse noxious inhibitory controls (DNIC) are very powerful long-lasting descending inhibitory controls which are pivotal in modulating the activity of spinal and trigeminal nociceptive neurons. DNIC are subserved by a loop involving supraspinal structures such as the lateral parabrachial nucleus and the subnucleus reticularis dorsalis. Surprisingly, though, whether the nucleus raphe magnus (NRM), another supraspinal area which is long known to be important in pain modulation, is involved in DNIC is still a matter of discussion. Here, we reassessed the role of the NRM neurons in DNIC by electrophysiologically recording from wide dynamic range (WDR) neurons in the trigeminal subnucleus oralis and pharmacologically manipulating the NRM OFF- and ON-cells. In control conditions, C-fiber-evoked responses in trigeminal WDR neurons are inhibited by a conditioning noxious heat stimulation applied to the hindpaw. We show that inactivating the NRM by microinjecting the GABAA receptor agonist, muscimol, both facilitates C-fiber-evoked responses of trigeminal WDR neurons and strongly attenuates their inhibition by heat applied to the hindpaw. Interestingly, selective blockade of ON-cells by microinjecting the broad-spectrum excitatory amino acid antagonist, kynurenate, into the NRM neither affects C-fiber-evoked responses nor attenuates DNIC of trigeminal WDR neurons. These results indicate that the NRM tonically inhibits trigeminal nociceptive inputs and is involved in the neuronal network underlying DNIC. Moreover, within NRM, OFF-cells might be more specifically involved in both the tonic and phasic descending inhibitory controls of trigeminal nociception. Copyright © 2014 Elsevier Inc. All rights reserved.

The bed nucleus of the stria terminalis regulates ethanol-seeking behavior in mice.

PubMed

Pina, Melanie M; Young, Emily A; Ryabinin, Andrey E; Cunningham, Christopher L

2015-12-01

Drug-associated stimuli are considered important factors in relapse to drug use. In the absence of drug, these cues can trigger drug craving and drive subsequent drug seeking. One structure that has been implicated in this process is the bed nucleus of the stria terminalis (BNST), a chief component of the extended amygdala. Previous studies have established a role for the BNST in cue-induced cocaine seeking. However, it is unclear if the BNST underlies cue-induced seeking of other abused drugs such as ethanol. In the present set of experiments, BNST involvement in ethanol-seeking behavior was assessed in male DBA/2J mice using the conditioned place preference procedure (CPP). The BNST was inhibited during CPP expression using electrolytic lesions (Experiment 1), co-infusion of GABAA and GABAB receptor agonists muscimol and baclofen (M+B; Experiment 2), and activation of inhibitory designer receptors exclusively activated by designer drugs (hM4Di-DREADD) with clozapine-N-oxide (CNO; Experiment 3). The magnitude of ethanol CPP was reduced significantly by each of these techniques. Notably, infusion of M+B (Exp. 2) abolished CPP altogether. Follow-up studies to Exp. 3 showed that ethanol cue-induced c-Fos immunoreactivity in the BNST was reduced by hM4Di activation (Experiment 4) and in the absence of hM4Di, CNO did not affect ethanol CPP (Experiment 5). Combined, these findings demonstrate that the BNST is involved in the modulation of cue-induced ethanol-seeking behavior. Copyright © 2015 Elsevier Ltd. All rights reserved.

The pharmacokinetic and pharmacodynamic effects of SL65.1498, a GABA-A alpha2,3 selective agonist, in comparison with lorazepam in healthy volunteers.

PubMed

de Haas, S L; Franson, K L; Schmitt, J A J; Cohen, A F; Fau, J B; Dubruc, C; van Gerven, J M A

2009-08-01

Benzodiazepines are effective short-term treatments for anxiety disorders, but their use is limited by undesirable side effects related to Central Nervous System impairment and tolerance development. SL65.1498 is a new compound that acts in vitro as a full agonist at the gamma-aminobutyric acid(A) 2 and 3 receptor and as a partial agonist at the 1 and 5 receptor subtypes. It is thought that the compound could be anxiolytic by its activation at the alpha2 and alpha3 receptor subtypes, without causing unfavourable side effects, which are believed to be mediated by the alpha1 and alpha5 subtypes. This study was a double-blind, five-way cross-over study to investigate the effects of three doses of SL65.1498 in comparison with placebo and lorazepam 2 mg in healthy volunteers. The objective was to select a dose level (expected to be therapeutically active), free of any significant deleterious effect. Psychomotor and cognitive effects were measured using a validated battery of measurements, including eye movements, body sway, memory tests, reaction-time assessments, and visual analogue scales. The highest dose of SL65.1498 showed slight effects on saccadic peak velocity and smooth pursuit performance, although to a much lesser extent than lorazepam. In contrast to lorazepam, none of the SL65.1498 doses affected body sway, visual analogue scale alertness, attention, or memory tests. This study showed that the three doses of SL65.1498 were well tolerated and induced no impairments on memory, sedation, psychomotor, and cognitive functions.

Duration of treatment and activation of α1-containing GABAA receptors variably affect the level of anxiety and seizure susceptibility after diazepam withdrawal in rats

PubMed Central

Kovačević, Jovana; Timić, Tamara; Tiruveedhula, Veera V.; Batinić, Bojan; Namjoshi, Ojas A.; Milić, Marija; Joksimović, Srđan; Cook, James M.; Savić, Miroslav M.

2014-01-01

Long-term use of benzodiazepine-type drugs may lead to physical dependence, manifested by withdrawal syndrome after abrupt cessation of treatment. The aim of the present study was to investigate the influence of duration of treatment, as well as the role of α1-containing GABAA receptors, in development of physical dependence to diazepam, assessed through the level of anxiety and susceptibility to pentylenetetrazole (PTZ)-induced seizures, 24 h after withdrawal from protracted treatment in rats. Withdrawal of 2 mg/kg diazepam after 28, but not after 14 or 21 days of administration led to an anxiety-like behavior in the elevated plus maze. Antagonism of the diazepam effects at α1-containing GABAA receptors, achieved by daily administration of the neutral modulator βCCt (5 mg/kg), did not affect the anxiety level during withdrawal. An increased susceptibility to PTZ-induced seizures was observed during diazepam withdrawal after 21 and 28 days of treatment. Daily co-administration of βCCt further decreased the PTZ-seizure threshold after 21 days of treatment, whilst it prevented the diazepam withdrawal-elicited decrease of the PTZ threshold after 28 days of treatment. In conclusion, the current study suggests that the role of α1-containing GABAA receptors in mediating the development of physical dependence may vary based on the effect being studied and duration of protracted treatment. Moreover, the present data supports previous findings that the lack of activity at α1-containing GABAA receptors is not sufficient to eliminate physical dependence liability of ligands of the benzodiazepine type. PMID:24695241

Decreased GABA-A binding on FMZ-PET in succinic semialdehyde dehydrogenase deficiency.

PubMed

Pearl, P L; Gibson, K M; Quezado, Z; Dustin, I; Taylor, J; Trzcinski, S; Schreiber, J; Forester, K; Reeves-Tyer, P; Liew, C; Shamim, S; Herscovitch, P; Carson, R; Butman, J; Jakobs, C; Theodore, W

2009-08-11

Succinic semialdehyde dehydrogenase (SSADH) deficiency is an autosomal recessive disorder of GABA metabolism characterized by elevated levels of GABA and gamma-hydroxybutyric acid. Clinical findings include intellectual impairment, hypotonia, hyporeflexia, hallucinations, autistic behaviors, and seizures. Autoradiographic labeling and slice electrophysiology studies in the murine model demonstrate use-dependent downregulation of GABA(A) receptors. We studied GABA(A) receptor activity in human SSADH deficiency utilizing [(11)C]-flumazenil (FMZ)-PET. FMZ binding was measured in 7 patients, 10 unaffected parents, and 8 healthy controls. Data analysis was performed using a reference region compartmental model, with time-activity curve from pons as the input function. Relative parametric binding potential (BP(ND)) was derived, with MRI-based pixel by pixel partial volume correction, in regions of interest drawn on coregistered MRI. In amygdala, hippocampus, cerebellar vermis, frontal, parietal, and occipital cortex, patients with SSADH deficiency had significant reductions in FMZ BP(ND) compared to parents and controls. Mean cortical values were 6.96 +/- 0.79 (controls), 6.89 +/- 0.71 (parents), and 4.88 +/- 0.77 (patients) (F ratio 16.1; p < 0.001). There were no differences between controls and parents in any cortical region. Succinic semialdehyde dehydrogenase (SSADH) deficient patients show widespread reduction in BZPR binding on [(11)C]-flumazenil-PET. Our results suggest that high endogenous brain GABA levels in SSADH deficiency downregulate GABA(A)-BZPR binding site availability. This finding suggests a potential mechanism for neurologic dysfunction in a serious neurodevelopmental disorder, and suggests that PET may be useful to translate studies in animal models to human disease.

The insecticide fipronil and its metabolite fipronil sulphone inhibit the rat α1β2γ2L GABAA receptor

PubMed Central

Li, P; Akk, G

2008-01-01

Background and purpose: Fipronil is the active ingredient in a number of widely used insecticides. Human exposure to fipronil leads to symptoms (headache, nausea and seizures) typically associated with the antagonism of GABAA receptors in the brain. In this study, we have examined the modulation of the common brain GABAA receptor subtype by fipronil and its major metabolite, fipronil sulphone. Experimental approach: Whole-cell and single-channel recordings were made from HEK 293 cells transiently expressing rat α1β2γ2L GABAA receptors. Key results: The major effect of fipronil was to increase the rate of current decay in macroscopic recordings. In single-channel recordings, the presence of fipronil resulted in shorter cluster durations without affecting the intracluster open and closed time distributions or the single-channel conductance. The α1V256S mutation, previously shown alleviate channel inhibition by inhibitory steroids and several insecticides, had a relatively small effect on channel block by fipronil. The mode of action of fipronil sulphone was similar to that of its parent compound but the metabolite was less potent at inhibiting the α1β2γ2L receptor. Conclusions and implications: We conclude that exposure to fipronil induces accumulation of receptors in a novel, long-lived blocked state. This process proceeds in parallel with and independently of, channel desensitization. The lower potency of fipronil sulphone indicates that the conversion serves as a detoxifying process in mammalian brain. PMID:18660823

Mechanism of the cardiovascular effects of the GABAA receptors of the ventral tegmental area of the rat brain.

PubMed

Yeganeh, Fahimeh; Ranjbar, Afsaneh; Hatam, Masoumeh; Nasimi, Ali

2015-07-23

The ventral tegmental area (VTA) contains GABA terminals involved in the regulation of the cardiovascular system. Previously, we demonstrated that blocking GABAA but not GABAB receptors produced a pressor response accompanied by marked bradycardia. This study was performed to find the possible mechanisms involved in these responses by blocking ganglionic nicotinic receptors, peripheral muscarinic receptors or peripheral V1 vasopressin receptors. Experiments were performed on urethane anesthetized male Wistar rats. Drugs were microinjected unilaterally into the VTA (100 nl). The average changes in mean arterial pressure (MAP) and heart rate (HR) were compared between pre- and post-treatment using paired t-test. Injection of bicuculline methiodide (BMI), a GABAA antagonist, into the VTA caused a significant increase in MAP and a decrease in HR. Administration (i.v.) of the nicotinic receptor blocker, hexamethonium, enhanced the pressor response but abolished the bradycardic response to BMI, which ruled out involvement of the sympathetic nervous system. Blockade of the peripheral muscarinic receptors by homatropine (i.v.) abolished the bradycardic effect of BMI, but had no effect on the pressor response, indicating that bradycardia was produced by the parasympathetic outflow to the heart. Both the pressor and bradycardic responses to BMI were blocked by V1 receptor antagonist (i.v.), indicating that administration of BMI in the VTA disinhibited the release of vasopressin into the circulation. In conclusion, we demonstrated that GABAergic mechanism of the VTA exerts a tonic inhibition on vasopressin release through activation of GABAA receptors. The sympathetic system is not involved in the decrease of blood pressure by GABA of the VTA. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Kampo Medicine: Evaluation of the Pharmacological Activity of 121 Herbal Drugs on GABAA and 5-HT3A Receptors

PubMed Central

Hoffmann, Katrin M.; Herbrechter, Robin; Ziemba, Paul M.; Lepke, Peter; Beltrán, Leopoldo; Hatt, Hanns; Werner, Markus; Gisselmann, Günter

2016-01-01

Kampo medicine is a form of Japanese phytotherapy originating from traditional Chinese medicine (TCM). During the last several decades, much attention has been paid to the pharmacological effects of these medical plants and their constituents. However, in many cases, a systematic screening of Kampo remedies to determine pharmacologically relevant targets is still lacking. In this study, a broad screening of Kampo remedies was performed to look for pharmacologically relevant 5-HT3A and GABAA receptor ligands. Several of the Kampo remedies are currently used for symptoms such as nausea, emesis, gastrointestinal motility disorders, anxiety, restlessness, or insomnia. Therefore, the pharmacological effects of 121 herbal drugs from Kampo medicine were analyzed as ethanol tinctures on heterologously expressed 5-HT3A and GABAA receptors, due to the involvement of these receptors in such pathophysiological processes. The tinctures of Lindera aggregata (radix) and Leonurus japonicus (herba) were the most effective inhibitory compounds on the 5-HT3A receptor. Further investigation of known ingredients in these compounds led to the identification of leonurine from Leonurus as a new natural 5-HT3A receptor antagonist. Several potentiating herbs (e.g., Magnolia officinalis (cortex), Syzygium aromaticum (flos), and Panax ginseng (radix)) were also identified for the GABAA receptor, which are all traditionally used for their sedative or anxiolytic effects. A variety of tinctures with antagonistic effects Salvia miltiorrhiza (radix) were also detected. Therefore, this study reveals new insights into the pharmacological action of a broad spectrum of herbal drugs from Kampo, allowing for a better understanding of their physiological effects and clinical applications. PMID:27524967

Kappa Opioid Receptor Agonist and Brain Ischemia

PubMed Central

Chunhua, Chen; Chunhua, Xi; Megumi, Sugita; Renyu, Liu

2014-01-01

Opioid receptors, especially Kappa opioid receptor (KOR) play an important role in the pathophysiological process of cerebral ischemia reperfusion injury. Previously accepted KOR agonists activity has included anti-nociception, cardiovascular, anti-pruritic, diuretic, and antitussive effects, while compelling evidence from various ischemic animal models indicate that KOR agonist have neuroprotective effects through various mechanisms. In this review, we aimed to demonstrate the property of KOR agonist and its role in global and focal cerebral ischemia. Based on current preclinical research, the KOR agonists may be useful as a neuroprotective agent. The recent discovery of salvinorin A, highly selective non-opioid KOR agonist, offers a new tool to study the role of KOR in brain HI injury and the protective effects of KOR agonist. The unique pharmacological profile of salvinorin A along with the long history of human usage provides its high candidacy as a potential alternative medication for brain HI injury. PMID:25574482

The effects of serotonin1A receptor on female mice body weight and food intake are associated with the differential expression of hypothalamic neuropeptides and the GABAA receptor.

PubMed

Butt, Isma; Hong, Andrew; Di, Jing; Aracena, Sonia; Banerjee, Probal; Shen, Chang-Hui

2014-10-01

Both common eating disorders anorexia nervosa and bulimia nervosa are characteristically diseases of women. To characterize the role of the 5-HT1A receptor (5-HT1A-R) in these eating disorders in females, we investigated the effect of saline or 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) treatment on feeding behavior and body weight in adult WT female mice and in adult 5-HT1A-R knockout (KO) female mice. Our results showed that KO female mice have lower food intake and body weight than WT female mice. Administration of 8-OH-DPAT decreased food intake but not body weight in WT female mice. Furthermore, qRT-PCR was employed to analyze the expression levels of neuropeptides, γ-aminobutyric acid A receptor subunit β (GABAA β subunits) and glutamic acid decarboxylase in the hypothalamic area. The results showed the difference in food intake between WT and KO mice was accompanied by differential expression of POMC, CART and GABAA β2, and the difference in body weight between WT and KO mice was associated with significantly different expression levels of CART and GABAA β2. As such, our data provide new insight into the role of 5-HT1A-R in both feeding behavior and the associated expression of neuropeptides and the GABAA receptor. Copyright © 2014 Elsevier Ltd. All rights reserved.

Adverse Effects of GLP-1 Receptor Agonists

PubMed Central

Filippatos, Theodosios D.; Panagiotopoulou, Thalia V.; Elisaf, Moses S.

2014-01-01

Glucagon-like peptide-1 (GLP-1) receptor agonists are a class of injective anti-diabetic drugs that improve glycemic control and many other atherosclerosis-related parameters in patients with type 2 diabetes (T2D). However, the use of this relatively new class of drugs may be associated with certain adverse effects. Concerns have been expressed regarding the effects of these drugs on pancreatic and thyroid tissue, since animal studies and analyses of drug databases indicate an association of GLP-1 receptor agonists with pancreatitis, pancreatic cancer, and thyroid cancer. However, several meta-analyses failed to confirm a cause-effect relation between GLP-1 receptor agonists and the development of these adverse effects. One benefit of GLP-1 receptor agonists is that they do not cause hypoglycemia when combined with metformin or thiazolidinediones, but the dose of concomitant sulphonylurea or insulin may have to be decreased to reduce the risk of hypoglycemic episodes. On the other hand, several case reports have linked the use of these drugs, mainly exenatide, with the occurrence of acute kidney injury, primarily through hemodynamic derangement due to nausea, vomiting, and diarrhea. The most common symptoms associated with the use of GLP-1 receptor agonists are gastrointestinal symptoms, mainly nausea. Other common adverse effects include injection site reactions, headache, and nasopharyngitis, but these effects do not usually result in discontinuation of the drug. Current evidence shows that GLP-1 receptor agonists have no negative effects on the cardiovascular risk of patients with T2D. Thus, GLP-1 receptor agonists appear to have a favorable safety profile, but ongoing trials will further assess their cardiovascular effects. The aim of this review is to analyze critically the available data regarding adverse events of GLP-1 receptor agonists in different anatomic systems published in Pubmed and Scopus. Whenever possible, certain differences between GLP-1

Involvement of posterior cingulate cortex in ketamine-induced psychosis relevant behaviors in rats.

PubMed

Ma, Jingyi; Leung, L Stan

2018-02-15

The involvement of posterior cingulate cortex (PCC) on ketamine-induced psychosis relevant behaviors was investigated in rats. Bilateral infusion of muscimol, a GABA A receptor agonist, into the PCC significantly antagonized ketamine-induced deficit in prepulse inhibition of a startle reflex (PPI), deficit in gating of hippocampal auditory evoked potentials, and behavioral hyperlocomotion in a dose dependent manner. Local infusion of ketamine directly into the PCC also induced a PPI deficit. Systemic injection of ketamine (3mg/kg,s.c.) induced an increase in power of electrographic activity in the gamma band (30-100Hz) in both the PCC and the hippocampus; peak theta (4-10Hz) power was not significantly altered, but peak theta frequency was increased by ketamine. In order to exclude volume conduction from the hippocampus to PCC, inactivation of the hippocampus was made by local infusion of muscimol into the hippocampus prior to ketamine administration. Muscimol in the hippocampus effectively blocked ketamine-induced increase of gamma power in the hippocampus but not in the PCC, suggesting independent generation of gamma waves in PCC and hippocampus. It is suggested that the PCC is part of the brain network mediating ketamine-induced psychosis related behaviors. Copyright © 2017 Elsevier B.V. All rights reserved.

Recruitment of GABA(A) receptors and fearfulness in chicks: modulation by systemic insulin and/or epinephrine.

PubMed

Cid, Mariana Paula; Toledo, Carolina Maribel; Salvatierra, Nancy Alicia

2013-02-01

One-day-old chicks were individually assessed on their latency to peck pebbles, and categorized as low latency (LL) or high latency (HL) according to fear. Interactions between acute stress and systemic insulin and epinephrine on GABA(A) receptor density in the forebrain were studied. At 10 days of life, LL and HL chicks were intraperitoneally injected with insulin, epinephrine or saline, and immediately after stressed by partial water immersion for 15 min and killed by decapitation. Forebrains were dissected and the GABA(A) receptor density was measured ex vivo by the (3)[H]-flunitrazepam binding assay in synaptosomes. In non-stressed chicks, insulin (non-hypoglycemic dose) at 2.50 IU/kg of body weight incremented the Bmax by 40.53% in the HL chicks compared to saline group whereas no significant differences were observed between individuals in the LL subpopulation. Additionally, insulin increased the Bmax (23.48%) in the HL group with respect to the LL ones, indicating that the insulin responses were different according to the anxiety of each category. Epinephrine administration (0.25 and 0.50mg/kg) incremented the Bmax in non-stressed chicks, in the LL group by about 37% and 33%, respectively, compared to ones injected with saline. In the stressed chicks, 0.25mg/kg bw epinephrine increased the Bmax significantly in the HL group by about 24% compared to saline, suggesting that the effect of epinephrine was only observed in the HL group under acute stress conditions. Similarly, the same epinephrine doses co-administered with insulin increased the receptor density in both subpopulations and also showed that the highest dose of epinephrine did not further increase the maximum density of GABA(A)R in HL chicks. These results suggest that systemic epinephrine, perhaps by evoking central norepinephrine release, modulated the increase in the forebrain GABA(A) receptor recruitment induced by both insulin and stress in different ways depending on the subpopulation

Beta-agonists and animal welfare

USDA-ARS?s Scientific Manuscript database

The use of beta-agonists in animal feed is a high profile topic within the U.S. as consumers and activist groups continue to question its safety. The only beta-agonist currently available for use in swine is ractopamine hydrochloride (RAC). This is available as Paylean™ (Elanco Animal Health – FDA a...

The insecticide fipronil and its metabolite fipronil sulphone inhibit the rat alpha1beta2gamma2L GABA(A) receptor.

PubMed

Li, P; Akk, G

2008-11-01

Fipronil is the active ingredient in a number of widely used insecticides. Human exposure to fipronil leads to symptoms (headache, nausea and seizures) typically associated with the antagonism of GABA(A) receptors in the brain. In this study, we have examined the modulation of the common brain GABA(A) receptor subtype by fipronil and its major metabolite, fipronil sulphone. Whole-cell and single-channel recordings were made from HEK 293 cells transiently expressing rat alpha1beta2gamma2L GABA(A) receptors. The major effect of fipronil was to increase the rate of current decay in macroscopic recordings. In single-channel recordings, the presence of fipronil resulted in shorter cluster durations without affecting the intracluster open and closed time distributions or the single-channel conductance. The alpha1V256S mutation, previously shown alleviate channel inhibition by inhibitory steroids and several insecticides, had a relatively small effect on channel block by fipronil. The mode of action of fipronil sulphone was similar to that of its parent compound but the metabolite was less potent at inhibiting the alpha1beta2gamma2L receptor. We conclude that exposure to fipronil induces accumulation of receptors in a novel, long-lived blocked state. This process proceeds in parallel with and independently of, channel desensitization. The lower potency of fipronil sulphone indicates that the conversion serves as a detoxifying process in mammalian brain.

Separate and combined effects of the GABAA positive allosteric modulator diazepam and Δ⁹-THC in humans discriminating Δ⁹-THC.

PubMed

Lile, Joshua A; Kelly, Thomas H; Hays, Lon R

2014-10-01

Our previous research suggested the involvement of γ-aminobutyric acid (GABA), in particular the GABAB receptor subtype, in the interoceptive effects of Δ(9)-tetrahydrocannabinol (Δ(9)-THC). The aim of the present study was to determine the potential involvement of the GABAA receptor subtype by assessing the separate and combined effects of the GABAA positive allosteric modulator diazepam and Δ(9)-THC using pharmacologically selective drug-discrimination procedures. Ten cannabis users learned to discriminate 30 mg oral Δ(9)-THC from placebo and then received diazepam (5 and 10mg), Δ(9)-THC (5, 15 and 30 mg) and placebo, alone and in combination. Self-report, task performance and physiological measures were also collected. Δ(9)-THC functioned as a discriminative stimulus, produced subjective effects typically associated with cannabinoids (e.g., High, Stoned, Like Drug) and elevated heart rate. Diazepam alone impaired performance on psychomotor performance tasks and increased ratings on a limited number of self-report questionnaire items (e.g., Any Effect, Sedated), but did not substitute for the Δ(9)-THC discriminative stimulus or alter the Δ(9)-THC discrimination dose-response function. Similarly, diazepam had limited impact on the other behavioral effects of Δ(9)-THC. These results suggest that the GABAA receptor subtype has minimal involvement in the interoceptive effects of Δ(9)-THC, and by extension cannabis, in humans. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

A Residue in Loop 9 of the β2-Subunit Stabilizes the Closed State of the GABAA Receptor*

PubMed Central

Williams, Carrie A.; Bell, Shannon V.; Jenkins, Andrew

2010-01-01

In γ-aminobutyric acid type A (GABAA) receptors, the structural elements that couple ligand binding to channel opening remain poorly defined. Here, site-directed mutagenesis was used to determine if Loop 9 on the non-GABA binding site interface of the β2-subunit may be involved in GABAA receptor activation. Specifically, residues Gly170-Gln185 of the β2-subunit were mutated to alanine, co-expressed with wild-type α1- and γ2S-subunits in human embryonic kidney (HEK) 293 cells and assayed for their activation by GABA, the intravenous anesthetic propofol and the endogenous neurosteroid pregnanolone using whole cell macroscopic recordings. Three mutants, G170A, V175A, and G177A, produced 2.5-, 6.7-, and 5.6-fold increases in GABA EC50 whereas one mutant, Q185A, produced a 5.2-fold decrease in GABA EC50. None of the mutations affected the ability of propofol or pregnanolone to potentiate a submaximal GABA response, but the Q185A mutant exhibited 8.3- and 3.5-fold increases in the percent direct activation by propofol and pregnanolone, respectively. Mutant Q185A receptors also had an increased leak current that was sensitive to picrotoxin, indicating an increased gating efficiency. Further Q185E, Q185L, and Q185W substitutions revealed a strong correlation between the hydropathy of the amino acid at this position and the GABA EC50. Taken together, these results indicate that β2 Loop 9 is involved in receptor activation by GABA, propofol, and pregnanolone and that β2(Q185) participates in hydrophilic interactions that are important for stabilizing the closed state of the GABAA receptor. PMID:20007704

Changes in the sensitivity of GABAA current rundown to drug treatments in a model of temporal lobe epilepsy

PubMed Central

Cifelli, Pierangelo; Palma, Eleonora; Roseti, Cristina; Verlengia, Gianluca; Simonato, Michele

2013-01-01

The pharmacological treatment of mesial temporal lobe epilepsy (mTLE), the most common epileptic syndrome in adults, is still unsatisfactory, as one-third of the patients are or become refractory to antiepileptic agents. Refractoriness may depend upon drug-induced alterations, but the disease per se may also undergo a progressive evolution that affects the sensitivity to drugs. mTLE has been shown to be associated with a dysfunction of the inhibitory signaling mediated by GABAA receptors. In particular, the repetitive activation of GABAA receptors produces a use-dependent decrease (rundown) of the evoked currents (IGABA), which is markedly enhanced in the hippocampus and cortex of drug-resistant mTLE patients. This phenomenon has been also observed in the pilocarpine model, where the increased IGABA rundown is observed in the hippocampus at the time of the first spontaneous seizure, then extends to the cortex and remains constant in the chronic phase of the disease. Here, we examined the sensitivity of IGABA to pharmacological modulation. We focused on the antiepileptic agent levetiracetam (LEV) and on the neurotrophin brain-derived neurotrophic factor (BDNF), which were previously reported to attenuate mTLE-induced increased rundown in the chronic human tissue. In the pilocarpine model, BDNF displayed a paramount effect, decreasing rundown in the hippocampus at the time of the first seizure, as well as in the hippocampus and cortex in the chronic period. In contrast, LEV did not affect rundown in the hippocampus, but attenuated it in the cortex. Interestingly, this effect of LEV was also observed on the still unaltered rundown observed in the cortex at the time of the first spontaneous seizure. These data suggest that the sensitivity of GABAA receptors to pharmacological interventions undergoes changes during the natural history of mTLE, implicating that the site of seizure initiation and the timing of treatment may highly affect the therapeutic outcome. PMID

The tuberal lateral hypothalamus is a major target for GABAA--but not GABAB-mediated control of food intake.

PubMed

Turenius, Christine I; Charles, Jonathan R; Tsai, Donna H; Ebersole, Priscilla L; Htut, Myat H; Ngo, Phuong T; Lara, Raul N; Stanley, B Glenn

2009-08-04

The lateral hypothalamus (LH) is a site of integration for control mechanisms of feeding behavior as it has extensive reciprocal connections with multiple intrahypothalamic and extrahypothalamic brain areas. Evidence suggests that blockade of ionotropric gamma-aminobutyric acid (GABA) receptors in the LH elicits eating in satiated rats. To determine whether this GABA(A) receptor antagonist effect is specific to the LH, the antagonist picrotoxin was injected into one of six nearby sites and food intake was measured. Picrotoxin at 133 pmol elicited eating in the LH, but not in surrounding sites (thalamus, lateral preoptic area, ventral tegmental area, dorsomedial hypothalamus, and entopeduncular nucleus). More specifically, picrotoxin injected into the tuberal LH (tLH) elicited eating, but was ineffective when injected into the anterior or posterior LH. We also investigated whether GABA(B) receptors in the LH participated in the control of food intake and found that neither blockade nor activation of these receptors under multiple conditions changed food intake. Collectively, our findings suggest that GABA(A) but not GABA(B) receptors in the tLH act to suppress feeding behavior.

Aspects of the homeostaic plasticity of GABAA receptor-mediated inhibition

PubMed Central

Mody, Istvan

2005-01-01

Plasticity of ligand-gated ion channels plays a critical role in nervous system development, circuit formation and refinement, and pathological processes. Recent advances have mainly focused on the plasticity of channels gated by excitatory amino acids, including their acclaimed role in learning and memory. These receptors, together with voltage-gated ion channels, have also been known to be subjected to a homeostatic form of plasticity that prevents destabilization of the neurone's function and that of the network during various physiological processes. To date, the plasticity of GABAA receptors has been examined mainly from a developmental and a pathological point of view. Little is known about homeostatic mechanisms governing their plasticity. This review summarizes some of the findings on the homeostatic plasticity of tonic and phasic inhibitory activity. PMID:15528237

GABAA Receptors Containing ρ1 Subunits Contribute to In Vivo Effects of Ethanol in Mice

PubMed Central

Blednov, Yuri A.; Benavidez, Jillian M.; Black, Mendy; Leiter, Courtney R.; Osterndorff-Kahanek, Elizabeth; Johnson, David; Borghese, Cecilia M.; Hanrahan, Jane R.; Johnston, Graham A. R.; Chebib, Mary; Harris, R. Adron

2014-01-01

GABAA receptors consisting of ρ1, ρ2, or ρ3 subunits in homo- or hetero-pentamers have been studied mainly in retina but are detected in many brain regions. Receptors formed from ρ1 are inhibited by low ethanol concentrations, and family-based association analyses have linked ρ subunit genes with alcohol dependence. We determined if genetic deletion of ρ1 in mice altered in vivo ethanol effects. Null mutant male mice showed reduced ethanol consumption and preference in a two-bottle choice test with no differences in preference for saccharin or quinine. Null mutant mice of both sexes demonstrated longer duration of ethanol-induced loss of righting reflex (LORR), and males were more sensitive to ethanol-induced motor sedation. In contrast, ρ1 null mice showed faster recovery from acute motor incoordination produced by ethanol. Null mutant females were less sensitive to ethanol-induced development of conditioned taste aversion. Measurement of mRNA levels in cerebellum showed that deletion of ρ1 did not change expression of ρ2, α2, or α6 GABAA receptor subunits. (S)-4-amino-cyclopent-1-enyl butylphosphinic acid (“ρ1” antagonist), when administered to wild type mice, mimicked the changes that ethanol induced in ρ1 null mice (LORR and rotarod tests), but the ρ1 antagonist did not produce these effects in ρ1 null mice. In contrast, (R)-4-amino-cyclopent-1-enyl butylphosphinic acid (“ρ2” antagonist) did not change ethanol actions in wild type but produced effects in mice lacking ρ1 that were opposite of the effects of deleting (or inhibiting) ρ1. These results suggest that ρ1 has a predominant role in two in vivo effects of ethanol, and a role for ρ2 may be revealed when ρ1 is deleted. We also found that ethanol produces similar inhibition of function of recombinant ρ1 and ρ2 receptors. These data indicate that ethanol action on GABAA receptors containing ρ1/ρ2 subunits may be important for specific effects of ethanol in vivo. PMID:24454882

Long-term studies of dopamine agonists.

PubMed

Hubble, Jean P

2002-02-26

Dopamine agonists have long been used as adjunctive therapy for the treatment of Parkinson's disease (PD). In more recent years these drugs have also been proved safe and effective as initial therapy in lieu of levodopa in the treatment of PD. Long-term levodopa therapy is associated with motor complications, including fluctuating response patterns and dyskinesia. By initially introducing a dopamine agonist as symptomatic drug therapy, it may be possible to postpone the use of levodopa and delay or prevent the development of motor complications. Recently, four clinical trials have explored this hypothesis by comparing the long-term response and side effects of levodopa with dopamine agonist therapy. The drugs studied have included ropinirole, pramipexole, cabergoline, and pergolide. In each of these projects, the occurrence of motor complications, such as wearing off and dyskinesia, was significantly less in the subjects assigned to initiation of therapy with a dopamine agonist. The addition of levodopa could be postponed by many months or even several years. Therefore, these long-term studies of dopamine agonists support the initiation of a dopamine agonist instead of levodopa in an effort to postpone levodopa-related motor complications. This therapeutic approach may be particularly appropriate in PD patients with a long treatment horizon on the basis of age and general good health. The extension phase of the long-term study comparing pramipexole with levodopa is ongoing, and follow-up information may help to establish the value of this treatment strategy.

Evaluation of AhR-agonists and AhR-agonist activity in sediments of Liaohe River protected areas, China.

PubMed

Zhang, Yun; Ke, Xin; Gui, Shaofeng; Wu, Xiaoqiong; Wang, Chunyong; Zhang, Haijun

2017-02-15

A total of 9 sediment samples of Liaohe River protected areas were collected to evaluate aryl hydrocarbon receptor agonists (AhR-agonists) and AhR-agonist activity via chemical analysis and in vitro H4IIE cell bioassay. Results indicated that bioassay-derived 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (Bio-TEQs) ranged from 89.1 to 251.1pg/g dry weight. Concentrations of 16 EPA polycyclic aromatic hydrocarbons (PAHs), 12 dioxin-like polychlorinated biphenyls (PCBs), and polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) ranged from 256.8 to 560.1ng/g, 79.2 to 416.2pg/g, and 199.6 to 538.4pg/g, respectively. According to potency balance analysis, TEQ chem s based on PAHs, PCBs, and PCDD/Fs could contribute 16.56% to 26.11% of Bio-TEQs. This could be explained by the potential existence of unidentified AhR-agonists and the potential non-additive interactions among AhR-agonists in sediment extracts. Through the different contributions to Bio-TEQs, this study confirms that PCDD/Fs were the main pollutants that induced significantly AhR-agonist activity in sediments of Liaohe River protected areas. Copyright © 2016. Published by Elsevier Ltd.

Modification of kindled amygdaloid seizures by opiate agonists and antagonists.

PubMed

Albertson, T E; Joy, R M; Stark, L G

1984-03-01

The effects of 19 opiate agonists and antagonists on kindled amygdaloid seizures in the rat were studied. The mu agonists tended to reduce the length of elicited afterdischarges and behavioral ranks, while markedly increasing postictal electroencephalogram spikes and behavioral arrest time. These effects were reversed by naloxone. The kappa agonists reduced behavioral rank and variably reduced afterdischarge length with a concomitant lengthening of postictal behavioral arrest time and number of electroencephalogram spikes. The putative sigma agonist, SKF 10,047, reduced afterdischarge durations only at the higher doses tested. The decreases found after the sigma agonists in postictal electroencephalogram spiking and time of behavioral arrest were not reversed by naloxone. Only the lower doses of normeperidine were found to decrease seizure thresholds. The mixed agonist/antagonists (MAA) cyclazocine and cyclorphan markedly increased seizure threshold and reduced afterdischarge duration and behavioral rank. Only the MAA pentazocine tended to increase threshold but not suprathreshold afterdischarge durations. The order of ability to modify the ictal events was MAA (selected) greater than kappa agonists greater than mu agonists greater than sigma agonists. The increase in postictal events (behavior arrest and spikes) was caused most effectively by pretreatment with mu agonist greater than kappa agonist greater than selected MAA greater than sigma agonists.(ABSTRACT TRUNCATED AT 250 WORDS)

Variation in GABA-A subunit gene copy number in an autistic patient with mosaic 4 p duplication (p12p16).

PubMed

Kakinuma, Hiroaki; Ozaki, Mamoru; Sato, Hitoshi; Takahashi, Hiroaki

2008-09-05

Autism has been associated with chromosomal aberrations, including duplications at chromosome 4, and the identification of genetic factors contributing to the etiology of this disease is the focus of much research. Here we report a Japanese girl with mosaic of chromosome 4p duplication, mos 46,XX,dup(4)(p12p16)[54]/46,XX[6], who was diagnosed with autism at 3 years of age. Fluorescence in situ hybridization (FISH) with probes covering the region spanning a cluster of the gamma aminobutyric acid A (GABA-A) receptor subunit genes in the proximal short arm of chromosome 4 demonstrated total three signals for the GABRG1, GABRA4, and GABRA2 genes, but only two signals for GABRB1. This suggests that aberrant copy number of the GABA-A receptor subunit genes may contribute to the etiology of autism in this patient. 2007 Wiley-Liss, Inc.

Dopamine agonists in the treatment of Parkinson's disease.

PubMed

Bonuccelli, Ubaldo; Pavese, Nicola

2006-01-01

Dopamine agonists are highly effective as adjunctive therapy to levodopa in advanced Parkinson's disease and have rapidly gained popularity as a monotherapy in the early stages of Parkinson's disease for patients less than 65-70 years old. In the latter case, dopamine agonists are about as effective as levodopa but patients demonstrate a lower tendency to develop motor complications. However, dopamine agonists lose efficacy over time and the number of patients remaining on agonist monotherapy decreases to less than 50% after 3 years of treatment. Thus, after a few years of treatment the majority of patients who started on dopamine agonists will be administered levodopa, in a combined dopaminergic therapy, in order to achieve a better control of motor symptoms.

Dopamine agonist withdrawal syndrome: implications for patient care.

PubMed

Nirenberg, Melissa J

2013-08-01

Dopamine agonists are effective treatments for a variety of indications, including Parkinson's disease and restless legs syndrome, but may have serious side effects, such as orthostatic hypotension, hallucinations, and impulse control disorders (including pathological gambling, compulsive eating, compulsive shopping/buying, and hypersexuality). The most effective way to alleviate these side effects is to taper or discontinue dopamine agonist therapy. A subset of patients who taper a dopamine agonist, however, develop dopamine agonist withdrawal syndrome (DAWS), which has been defined as a severe, stereotyped cluster of physical and psychological symptoms that correlate with dopamine agonist withdrawal in a dose-dependent manner, cause clinically significant distress or social/occupational dysfunction, are refractory to levodopa and other dopaminergic medications, and cannot be accounted for by other clinical factors. The symptoms of DAWS include anxiety, panic attacks, dysphoria, depression, agitation, irritability, suicidal ideation, fatigue, orthostatic hypotension, nausea, vomiting, diaphoresis, generalized pain, and drug cravings. The severity and prognosis of DAWS is highly variable. While some patients have transient symptoms and make a full recovery, others have a protracted withdrawal syndrome lasting for months to years, and therefore may be unwilling or unable to discontinue DA therapy. Impulse control disorders appear to be a major risk factor for DAWS, and are present in virtually all affected patients. Thus, patients who are unable to discontinue dopamine agonist therapy may experience chronic impulse control disorders. At the current time, there are no known effective treatments for DAWS. For this reason, providers are urged to use dopamine agonists judiciously, warn patients about the risks of DAWS prior to the initiation of dopamine agonist therapy, and follow patients closely for withdrawal symptoms during dopamine agonist taper.

Structure/function relationships of calcitonin analogues as agonists, antagonists, or inverse agonists in a constitutively activated receptor cell system.

PubMed

Pozvek, G; Hilton, J M; Quiza, M; Houssami, S; Sexton, P M

1997-04-01

The structure/function relationship of salmon calcitonin (sCT) analogues was investigated in heterologous calcitonin receptor (CTR) expression systems. sCT analogues with progressive amino-terminal truncations intermediate of sCT-(1-32) to sCT-(8-32) were examined for their ability to act as agonists, antagonists, or inverse agonists. Two CTR cell clones, B8-H10 and G12-E12, which express approximately 5 million and 25,000 C1b receptors/cell, respectively, were used for this study. The B8-H10 clone has an approximately 80-fold increase in basal levels of intracellular cAMP due to constitutive activation of the overexpressed receptor. In whole-cell competition binding studies, sCT-(1-32) was more potent than any of its amino-terminally truncated analogues in competition for 125I-sCT binding. In cAMP accumulation studies, sCT-(1-32) and modified analogues sCT-(2-32) and sCT-(3-32) had agonist activities. SDZ-216-710, with an amino-terminal truncation of four amino acids, behaved as a partial agonist/antagonist, whereas amino-terminal truncations of six or seven amino acid residues produced a 16-fold reduction in basal cAMP levels and attenuated the response to the agonist sCT-(1-32) in the constitutively active CTR system. This inverse agonist effect was insensitive to pertussis toxin inhibition. In contrast, the inverse agonist activity of these peptides was not observed in the nonconstitutively active CTR system, in which sCT analogues with amino-terminal truncations of four or more amino acids behaved as neutral competitive antagonists. These results suggest that the inverse agonist activity is mediated by stabilization of the inactive state of the receptor, which does not couple to G protein, and attenuates basal signaling initiated by ligand-independent activation of the effector adenylyl cyclase.

Identification of Determinants Required for Agonistic and Inverse Agonistic Ligand Properties at the ADP Receptor P2Y12

PubMed Central

Schmidt, Philipp; Ritscher, Lars; Dong, Elizabeth N.; Hermsdorf, Thomas; Cöster, Maxi; Wittkopf, Doreen; Meiler, Jens

2013-01-01

The ADP receptor P2Y12 belongs to the superfamily of G protein–coupled receptors (GPCRs), and its activation triggers platelet aggregation. Therefore, potent antagonists, such as clopidogrel, are of high clinical relevance in prophylaxis and treatment of thromboembolic events. P2Y12 displays an elevated basal activity in vitro, and as such, inverse agonists may be therapeutically beneficial compared with antagonists. Only a few inverse agonists of P2Y12 have been described. To expand this limited chemical space and improve understanding of structural determinants of inverse agonist-receptor interaction, this study screened a purine compound library for lead structures using wild-type (WT) human P2Y12 and 28 constitutively active mutants. Results showed that ATP and ATP derivatives are agonists at P2Y12. The potency at P2Y12 was 2-(methylthio)-ADP > 2-(methylthio)-ATP > ADP > ATP. Determinants required for agonistic ligand activity were identified. Molecular docking studies revealed a binding pocket for the ATP derivatives that is bordered by transmembrane helices 3, 5, 6, and 7 in human P2Y12, with Y105, E188, R256, Y259, and K280 playing a particularly important role in ligand interaction. N-Methyl-anthraniloyl modification at the 3′-OH of the 2′-deoxyribose leads to ligands (mant-deoxy-ATP [dATP], mant-deoxy-ADP) with inverse agonist activity. Inverse agonist activity of mant-dATP was found at the WT human P2Y12 and half of the constitutive active P2Y12 mutants. This study showed that, in addition to ADP and ATP, other ATP derivatives are not only ligands of P2Y12 but also agonists. Modification of the ribose within ATP can result in inverse activity of ATP-derived ligands. PMID:23093496

An Investigation of the Differential Effects of Ursane Triterpenoids from Centella asiatica, and Their Semisynthetic Analogues, on GABAA Receptors.

PubMed

Hamid, Kaiser; Ng, Irene; Tallapragada, Vikram J; Váradi, Linda; Hibbs, David E; Hanrahan, Jane; Groundwater, Paul W

2016-09-01

The ursane triterpenoids, asiatic acid 1 and madecassic acid 2, are the major pharmacological constituents of Centella asiatica, commonly known as Gotu Kola, which is used traditionally for the treatment of anxiety and for the improvement of cognition and memory. Using the two-electrode voltage-clamp technique, these triterpenes, and some semisynthetic derivatives, were found to exhibit selective negative modulation of different subtypes of the GABAA receptor expressed in Xenopus laevis oocytes. Despite differing by only one hydroxyl group, asiatic acid 1 was found to be a negative modulator of the GABA-induced current at α1 β2 γ2L, α2 β2 γ2L and α5 β3 γ2L GABAA receptors, while madecassic acid 2 was not. Asiatic acid 1 exhibited the greatest effect at α1 β2 γ2L (IC50 37.05 μm), followed by α5 β3 γ2L (IC50 64.05 μm) then α2 β2 γ2L (IC50 427.2 μm) receptors. Conversion of the carboxylic acid group of asiatic acid 1 to a carboxamide group (2α,3β,23-trihydroxy-urs-12-en-28-amide 5) resulted in enhanced inhibition at both the α1 β2 γ2L (IC50 14.07 μm) and α2 β2 γ2L receptor subtypes (IC50 28.41 μm). The results of this study, and the involvement of α5 -containing GABAA receptors in cognition and memory, suggest that asiatic acid 1 may be a lead compound for the enhancement of cognition and memory. © 2016 John Wiley & Sons A/S.

Improvement in verbal memory following SSRI augmentation of antipsychotic treatment is associated with changes in the expression of mRNA encoding for the GABA-A receptor and BDNF in PMC of schizophrenic patients.

PubMed

Silver, Henry; Mandiuk, Nina; Einoch, Reef; Susser, Ehud; Danovich, Lena; Bilker, Warren; Youdim, Moussa; Weinreb, Orly

2015-05-01

Verbal memory impairment in schizophrenia is associated with abnormalities in gamma-aminobutyric acid (GABA)-ergic and brain-derived neurotrophic factor (BDNF) systems. Recent evidence from animal and clinical studies that adding fluvoxamine to antipsychotics alters the expression of transcripts encoding for the GABA-A receptor and BDNF led us to postulate that fluvoxamine augmentation may improve memory in schizophrenia. To test this, we examined the effect of add-on fluvoxamine on verbal memory and other cognitive functions and related it to the expression of mRNA coding for the GABA-A receptor and BDNF in peripheral mononuclear cells (PMC) of schizophrenic patients. Twenty-nine patients completed a 6-week study in which fluvoxamine (100 mg/day) was added to ongoing antipsychotic treatment. Verbal memory, abstraction working memory, object and face recognition, and psychomotor speed and clinical symptoms were assessed at baseline and after 3 and 6 weeks of treatment. Blood samples were taken at baseline and weeks 1, 3, and 6 and PMC was assayed for the GABA-A beta3 receptor and BDNF mRNA by quantitative real-time reverse transcription-PCR. Associative and logical verbal memory improved significantly and showed a significant correlation with changes in PMC BDNF and GABA-A beta3 receptor mRNA, which increased during treatment. Abstraction and object recognition improved, but this did not correlate with PMC measures. Negative and positive symptoms improved significantly; the latter showed significant correlations with changes in PMC measures. Addition of fluvoxamine to antipsychotics improves verbal memory. It is postulated that the mechanism involves enhanced GABA-A receptor/BDNF-dependent synaptic plasticity in the hippocampus.

GABAA receptors: Various stoichiometrics of subunit arrangement in α1β3 and α1β3ε receptors.

PubMed

Has, Ahmad Tarmizi Che; Chebib, Mary

2018-05-15

GABAA receptors (GABAARs) are members of the Cys-loop ligand-gated ion channel (LGIC) superfamily, which includes nicotinic acetylcholine, glycine, and serotonin (5HT3) receptors [1,2,3,4]. LGICs typically mediate fast synaptic transmission via the movement of ions through channels gated by neurotransmitters, such as acetylcholine for nicotinic receptors and GABA for GABAARs [5]. The term Cys-loop receptors originates from the presence of a conserved disulphide bond (or bridge) which holds together two cysteine amino acids of the loop that forms from the structure of polypeptides in the extracellular domain of the receptor's subunit [6]. GABAARs are pentameric transmembrane protein complexes consisting of five subunits from a variety of polypeptide subunits [7,8]. All of these subunits are pseudo-symmetrically organized in the plane of the membrane, with a Cl--selective channel in the middle of the complex. To date, nineteen GABAAR subunits have been identified and categorized into eight classes, α1-6, β1-3, γ1-3, δ, ε, θ, π and ρ1-3, but their variety is further broadened by the existence of several splice forms for certain subunits (e.g., α6, β2 and γ2) [9,10,11,12]. The subunits within each class have an amino acid sequence homology of 70% or more, whereas those with a sequence homology of 30% or less are grouped into different classes [13,14]. A subunit consists of four transmembrane domains (TM1-TM4), each forming an α-helix; a large extracellular N-terminal domain that incorporates part of the orthosteric agonist/antagonist binding site; a large intracellular loop between the TM3 and TM4; a small intracellular loop between TM1 and TM2; a small extracellular loop between TM2 and TM3; and a C-terminal extracellular domain [15,16]. Each subunit is arranged in such a way as to create principal and complementary interfaces with the other subunits, and in a position such that the TM2 of each subunit forms the wall of the channel pore [17,18,19]. The

Agonist-Directed Desensitization of the β2-Adrenergic Receptor

PubMed Central

Goral, Vasiliy; Jin, Yan; Sun, Haiyan; Ferrie, Ann M.; Wu, Qi; Fang, Ye

2011-01-01

The β2-adrenergic receptor (β2AR) agonists with reduced tachyphylaxis may offer new therapeutic agents with improved tolerance profile. However, receptor desensitization assays are often inferred at the single signaling molecule level, thus ligand-directed desensitization is poorly understood. Here we report a label-free biosensor whole cell assay with microfluidics to determine ligand-directed desensitization of the β2AR. Together with mechanistic deconvolution using small molecule inhibitors, the receptor desensitization and resensitization patterns under the short-term agonist exposure manifested the long-acting agonism of salmeterol, and differentiated the mechanisms of agonist-directed desensitization between a full agonist epinephrine and a partial agonist pindolol. This study reveals the cellular mechanisms of agonist-selective β2AR desensitization at the whole cell level. PMID:21541288

Sports doping: emerging designer and therapeutic β2-agonists.

PubMed

Fragkaki, A G; Georgakopoulos, C; Sterk, S; Nielen, M W F

2013-10-21

Beta2-adrenergic agonists, or β2-agonists, are considered essential bronchodilator drugs in the treatment of bronchial asthma, both as symptom-relievers and, in combination with inhaled corticosteroids, as disease-controllers. The use of β2-agonists is prohibited in sports by the World Anti-Doping Agency (WADA) due to claimed anabolic effects, and also, is prohibited as growth promoters in cattle fattening in the European Union. This paper reviews the last seven-year (2006-2012) literature concerning the development of novel β2-agonists molecules either by modifying the molecule of known β2-agonists or by introducing moieties producing indole-, adamantyl- or phenyl urea derivatives. New emerging β2-agonists molecules for future therapeutic use are also presented, intending to emphasize their potential use for doping purposes or as growth promoters in the near future. © 2013.

Direct pyrogenic input from prostaglandin EP3 receptor-expressing preoptic neurons to the dorsomedial hypothalamus

PubMed Central

Nakamura, Yoshiko; Nakamura, Kazuhiro; Matsumura, Kiyoshi; Kobayashi, Shigeo; Kaneko, Takeshi; Morrison, Shaun F.

2008-01-01

Fever is induced by the neuronal mechanism in the brain. Prostaglandin (PG) E2 acts as a pyrogenic mediator in the preoptic area (POA) probably through the EP3 subtype of PGE receptor expressed on GABAergic neurons, and this PGE2 action triggers neuronal pathways for sympathetic thermogenesis in peripheral effector organs including brown adipose tissue (BAT). To explore pyrogenic efferent pathways from the POA, we here determined projection targets of EP3 receptor-expressing POA neurons with a special focus on rat hypothalamic regions including the dorsomedial hypothalamic nucleus (DMH), which is known as a center for autonomic responses to stress. Among injections of cholera toxin b-subunit (CTb), a retrograde tracer, into hypothalamic regions at the rostrocaudal level of the DMH, injections into the DMH, lateral hypothalamic area (LH), and dorsal hypothalamic area (DH) resulted in EP3 receptor immunolabeling in substantial populations of CTb-labeled neurons in the POA. Bilateral microinjections of muscimol, a GABAA receptor agonist, into the DMH and a ventral region of the DH, but not those into the LH, inhibited thermogenic (BAT sympathetic nerve activity, BAT temperature, core body temperature, and expired CO2) and cardiovascular (arterial pressure and heart rate) responses to an intra-POA PGE2 microinjection. Further immunohistochemical observations revealed close association of POA-derived GABAergic axon swellings with DMH neurons projecting to the medullary raphe regions where sympathetic premotor neurons for febrile and thermoregulatory responses are localized. These results suggest that a direct projection of EP3 receptor-expressing POA neurons to the DMH/DH region mediates febrile responses via a GABAergic mechanism. PMID:16367780

Mixed Kappa/Mu Opioid Receptor Agonists: The 6β-Naltrexamines

PubMed Central

Cami-Kobeci, Gerta; Neal, Adrian P.; Bradbury, Faye A.; Purington, Lauren C.; Aceto, Mario D.; Harris, Louis S.; Lewis, John W.; Traynor, John R.; Husbands, Stephen M.

2011-01-01

Ligands from the naltrexamine series have consistently demonstrated agonist activity at kappa opioid receptors (KOR), with varying activity at the mu opioid receptor (MOR). Various 6β-cinnamoylamino derivatives were made with the aim of generating ligands with a KOR agonist/MOR partial agonist profile, as ligands with this activity may be of interest as treatment agents for cocaine abuse. The ligands all displayed the desired high affinity, non-selective binding in vitro and in the functional assays were high efficacy KOR agonists with some partial agonist activity at MOR. Two of the new ligands (12a, 12b) have been evaluated in vivo, with 12a acting as a KOR agonist, and therefore somewhat similar to the previously evaluated analogues 3–6, while 12b displayed predominant MOR agonist activity. PMID:19253970

Building a bridge between neurobiology and mental illness.

PubMed

Costa, E

1992-10-01

GABA (gamma amino butyric acid) is the most abundant and important inhibitory transmitter in mammalian CNS. It counterbalances the glutamate mediated neuronal excitation. Abnormalities of the interaction of these two transmitters might change the mechanisms of neuronal group selection that according to Edelman [Neural Darwinism. Basic Books, New York] play a role in mediating several brain functions including cognition processes. Indeed imbalances in GABAergic functions were shown to elicit psychoses. They can be obtained by administration of drugs that affect synthesis, metabolism and uptake of GABA and thereby cause a persistent stimulation of GABAA receptors or perhaps by genetic abnormalities in DNA transcription, pre-mRNA splicing, mRNA translation and posttranslation modifications of GABAA receptor subunits. The complexities in the regulation of GABAA receptor subunit structure, synthesis, assembly and the brain location of specific mRNA encoding for these subunits are investigated with in situ mRNA hybridization specific for subunits of GABAA receptors. The role of the variability resulting from the complexities in the regulation of GABAA receptor allosteric modulation by drugs and putative endogenous allosteric modulators of GABA action at GABAA receptors is discussed. This discussion gives relevance to the possibility that genetic abnormalities in the expression of proteins participating in GABAergic function are to be considered as a possible target of the genetic defects operative in psychoses. In line with this thinking, it is suggested that partial allosteric modulators (partial agonists) of GABAA receptors and the phosphothioate or methylphosphonate analogs antisense to specific mRNA oligonucleotides that mediate the expression of genetic information concerning GABAA and glutamate receptor subunits may become valuable tools in psychiatric research. Perhaps in the future these studies might generate new ideas useful in the therapy of genetically

Axonal remodeling for motor recovery after traumatic brain injury requires downregulation of γ-aminobutyric acid signaling

PubMed Central

Lee, S; Ueno, M; Yamashita, T

2011-01-01

Remodeling of the remnant neuronal network after brain injury possibly mediates spontaneous functional recovery; however, the mechanisms inducing axonal remodeling during spontaneous recovery remain unclear. Here, we show that altered γ-aminobutyric acid (GABA) signaling is crucial for axonal remodeling of the contralesional cortex after traumatic brain injury. After injury to the sensorimotor cortex in mice, we found a significant decrease in the expression of GABAAR-α1 subunits in the intact sensorimotor cortex for 2 weeks. Motor functions, assessed by grid walk and cylinder tests, spontaneously improved in 4 weeks after the injury to the sensorimotor cortex. With motor recovery, corticospinal tract (CST) axons from the contralesional cortex sprouted into the denervated side of the cervical spinal cord at 2 and 4 weeks after the injury. To determine the functional implications of the changes in the expression of GABAAR-α1 subunits, we infused muscimol, a GABA R agonist, into the contralesional cortex for a week after the injury. Compared with the vehicle-treated mice, we noted significantly inhibited recovery in the muscimol-treated mice. Further, muscimol infusion greatly suppressed the axonal sprouting into the denervated side of the cervical spinal cord. In conclusion, recovery of motor function and axonal remodeling of the CST following cortical injury requires suppressed GABAAR subunit expression and decreased GABAergic signaling. PMID:21412279

Microinjections into the pedunculopontine tegmentum: effects of the GABAA antagonist, bicuculline, on sleep, PGO waves and behavior.

PubMed

Sanford, L D; Hunt, W K; Ross, R J; Morrison, A R; Pack, A I

1998-07-01

Neurons in the peribrachial region (PB) at the pontine border are implicated in the generation of ponto-geniculo-occipital (PGO) waves, which appear spontaneously during rapid eye movement sleep (REM) and in association with alerting behaviors during waking, as well as in the regulation of REM itself. It has been hypothesized that PGO-related bursting in a subpopulation of these neurons results from low threshold spikes triggered by phasic hyperpolarizations or by excitatory inputs reaching a steadily hyperpolarized neuron. The hyperpolarization necessary for triggering the low threshold spikes may come from local GABA neurons or from GABAergic input into PB. To test the hypothesis that antagonizing GABA would alter PGO wave generation and/or behavioral state, we microinfused, in cats, the GABAA antagonist, bicuculline, locally into PB and monitored behavior, behavioral state and PGO waves recorded in the lateral geniculate bodies. Bicuculline produced no significant alteration in PGO wave activity. In 3 cats, bicuculline produced behaviors ranging from spontaneous orienting and startle (4 cats) to flight behaviors (2 cats) and aggressiveness (2 cats), an effect probably due to diffusion into the central gray region. Thus, the results do not support a GABAA-ergic role in PB in the generation of PGO waves.

Exploring the binding energy profiles of full agonists, partial agonists, and antagonists of the α7 nicotinic acetylcholine receptor.

PubMed

Tabassum, Nargis; Ma, Qianyun; Wu, Guanzhao; Jiang, Tao; Yu, Rilei

2017-09-01

Nicotinic acetylcholine receptors (nAChRs) belong to the Cys-loop receptor family and are important drug targets for the treatment of neurological diseases. However, the precise determinants of the binding efficacies of ligands for these receptors are unclear. Therefore, in this study, the binding energy profiles of various ligands (full agonists, partial agonists, and antagonists) were quantified by docking those ligands with structural ensembles of the α7 nAChR exhibiting different degrees of C-loop closure. This approximate treatment of interactions suggested that full agonists, partial agonists, and antagonists of the α7 nAChR possess distinctive binding energy profiles. Results from docking revealed that ligand binding efficacy may be related to the capacity of the ligand to stabilize conformational states with a closed C loop.

Acute inactivation of the contralesional hemisphere for longer durations improves recovery after cortical injury.

PubMed

Mansoori, Babak K; Jean-Charles, Loyda; Touvykine, Boris; Liu, Aihua; Quessy, Stephan; Dancause, Numa

2014-04-01

A rapidly growing number of studies using inhibition of the contralesional hemisphere after stroke are reporting improvement in motor performance of the paretic hand. These studies have used different treatment onset time, duration and non-invasive methods of inhibition. Whereas these results are encouraging, several questions regarding the mechanisms of inhibition and the most effective treatment parameters are currently unanswered. In the present study, we used a rat model of cortical lesion to study the effects of GABA-mediated inactivation on motor recovery. In particular, we were interested in understanding better the effect of inactivation duration when it is initiated within hours following a cortical lesion. Cortical lesions were induced with endothelin-1 microinjections. The contralesional hemisphere was inactivated with continuous infusion of the GABA-A agonist Muscimol for 3, 7 or 14days in three different groups of animals. In a fourth group, Muscimol was infused at slower rate for 14days to provide additional insights on the relation between the effects of inactivation on the non-paretic forelimb behavior and the recovery of the paretic forelimb. In spontaneously recovered animals, the lesion caused a sustained bias to use the non-paretic forelimb and long-lasting grasping deficits with the paretic forelimb. Contralesional inactivation produced a general decrease of behavioral activity, affected the spontaneous use of the forelimbs and caused a specific reduction of the non-paretic forelimb function. The intensity and the duration of these behavioral effects varied in the different experimental groups. For the paretic forelimb, increasing inactivation duration accelerated the recovery of grasping function. Both groups with 14days of inactivation had similar recovery profiles and performed better than animals that spontaneously recovered. Whereas the plateau performance of the paretic forelimb correlated with the duration of contralesional inactivation

Opposing roles for GABAA and GABAC receptors in short-term memory formation in young chicks.

PubMed

Gibbs, M E; Johnston, G A R

2005-01-01

The inhibitory neurotransmitter GABA has both inhibitory and enhancing effects on short-term memory for a bead discrimination task in the young chick. Low doses of GABA (1-3 pmol/hemisphere) injected into the multimodal association area of the chick forebrain, inhibit strongly reinforced memory, whereas higher doses (30-100 pmol/hemisphere) enhance weakly reinforced memory. The effect of both high and low doses of GABA is clearly on short-term memory in terms of both the time of injection and in the time that the memory loss occurs. We argue on the basis of relative sensitivities to GABA and to selective GABA receptor antagonists that low doses of GABA act at GABAC receptors (EC50 approximately 1 microM) and the higher doses of GABA act via GABAA receptors (EC50 approximately 10 microM). The selective GABAA receptor antagonist bicuculline inhibited strongly reinforced memory in a dose and time dependent manner, whereas the selective GABAC receptor antagonists TPMPA and P4MPA enhanced weakly reinforced in a dose and time dependent manner. Confirmation that different levels of GABA affect different receptor subtypes was demonstrated by the shift in the GABA dose-response curves to the selective antagonists. It is clear that GABA is involved in the control of short-term memory formation and its action, enhancing or inhibiting, depends on the level of GABA released at the time of learning.

The effect of age on the discriminative stimulus effects of ethanol and its GABA(A) receptor mediation in cynomolgus monkeys.

PubMed

Helms, Christa M; Grant, Kathleen A

2011-08-01

Excessive alcohol consumption is less common among aged compared to young adults, with aged adults showing greater sensitivity to many behavioral effects of ethanol. This study compared the discriminative stimulus effects of ethanol in young and middle-aged adult cynomolgus monkeys (Macaca fascicularis) and its γ-aminobutyric acid (GABA)(A) receptor mediation. Two male and two female monkeys trained to discriminate ethanol (1.0 g/kg, i.g.; 60-min pre-treatment interval) from water at 5-6 years of age (Grant et al. in Psychopharmacology 152:181-188, 2000) were re-trained in the current study more than a decade later (19.3 ± 1.0 years of age) for a within-subjects comparison. Also, four experimentally naïve middle-aged (mean ± SEM, 17.0 ± 1.5 years of age) female monkeys were trained to discriminate ethanol for between-subjects comparison with published data from young adult naïve monkeys. Two of the naïve middle-aged monkeys attained criterion performance, with weak stimulus control and few discrimination tests, despite greater blood-ethanol concentration 60 min after 1.0 g/kg ethanol in middle-aged compared to young adult female monkeys (Green et al. in Alcohol Clin Exp Res 23:611-616, 1999). The efficacy of the GABA(A) receptor positive modulators pentobarbital, midazolam, allopregnanolone, pregnanolone, and androsterone to substitute for the discriminative stimulus effects of 1.0 g/kg ethanol was maintained from young adulthood to middle age. The data suggest that 1.0 g/kg ethanol is a weak discriminative stimulus in naive middle-aged monkeys. Nevertheless, the GABA(A) receptor mechanisms mediating the discriminative stimulus effects of ethanol, when learned as a young adult, appear stable across one third of the primate lifespan.

Neurosteroids in hepatic encephalopathy: Novel insights and new therapeutic opportunities.

PubMed

Butterworth, Roger F

2016-06-01

Hepatic encephalopathy (HE) is a serious neuropsychiatric disorder resulting from liver failure. Symptoms of HE include mild cognitive impairment, stupor and coma. Morphological changes to neuroglia (both astrocytes and microglia) occur in HE consisting of cytotoxic brain edema (astrocyte swelling) in acute liver failure and Alzheimer type-2 astrocytosis in cirrhosis. Visual-evoked responses in animals with liver failure and HE manifest striking similarities to those in animals treated with agonists of the GABA-A receptor complex. Neurosteroids are synthesized in brain following activation of translocator protein (TSPO), a mitochondrial neuroglial cholesterol-transporter protein. TSPO sites are activated in both animal models of HE as well as in autopsied brain tissue from HE patients. Activation of TSPO sites results in increased cholesterol transport into the mitochondrion followed by stimulation of a metabolic pathway culminating in the synthesis of allopregnanolone (ALLO) and tetrahydrodeoxycorticosterone (THDOC), neurosteroids with potent positive allosteric modulatory action on the GABA-A receptor complex. Concentrations of ALLO and THDOC in brain tissue from mice with HE resulting from toxic liver injury are sufficient to induce sedation in animals of the same species and significant increases in concentrations of ALLO have been reported in autopsied brain tissue from cirrhotic patients with HE leading to the proposal that "increased GABAergic tone" in HE results from that increased brain concentrations of this neurosteroid. Agents with the potential to decrease neurosteroid synthesis and/or prevent their modulatory actions on the GABA-A receptor complex may provide novel approaches to the management and treatment of HE. Such agents include indomethacin, benzodiazepine receptor inverse agonists and a novel series of compounds known as GABA-A receptor-modulating steroid antagonists (GAMSA). Copyright © 2015 Elsevier Ltd. All rights reserved.

New thrombopoietin receptor agonists for platelet disorders.

PubMed

Homeida, S; Ebdon, C; Batty, P; Jackson, B; Kolade, S; Bateman, C; Peng, Y Y; Stasi, R

2012-04-01

Since thrombopoietin (TPO) was cloned in 1994, TPO receptor (TPO-R) agonists have been developed which have shown significant clinical activity in various conditions characterized by thrombocytopenia. First-generation TPO-R agonists were recombinant forms of human TPO. The clinical development of these molecules was discontinued after one of them, pegylated recombinant human megakaryocyte growth and development factor, was associated with the development of neutralizing autoantibodies cross-reacting with endogenous TPO. Second-generation TPO-R agonists are now available, which present no sequence homology to endogenous TPO. Two of these new agents, romiplostim and eltrombopag, have been granted marketing authorization for use in patients with primary immune thrombocytopenia unresponsive to conventional treatments. Clinical trials with TPO-R agonists are also ongoing in other thrombocytopenias, such as hepatitis C virus-related thrombocytopenia and the myelodysplastic syndromes. Copyright 2012 Prous Science, S.A.U. or its licensors. All rights reserved.

Delta-opioid receptor expression in the ventral tegmental area protects against elevated alcohol consumption.

PubMed

Margolis, Elyssa B; Fields, Howard L; Hjelmstad, Gregory O; Mitchell, Jennifer M

2008-11-26

Alcoholism is a complex and debilitating syndrome affecting approximately 140 million people worldwide. However, not everyone who consumes ethanol develops abuse, raising the possibility that some individuals have a protective mechanism that inhibits elevated alcohol consumption. We tested the hypothesis that the delta-opioid receptor (DOR) plays such a protective role. Here we show that DOR activity in the ventral tegmental area (VTA) robustly decreases ethanol consumption in rats and that these effects depend on baseline ethanol consumption. Intra-VTA microinjection of the DOR agonist DPDPE decreases drinking, particularly in low-drinking animals. Furthermore, VTA microinjection of the DOR selective antagonist TIPP-Psi increases drinking in low, but not high, drinkers and this increase is blocked by comicroinjection of the GABA(A) antagonist bicuculline. Using electrophysiological techniques we found that in VTA brain slices from drinking rats DPDPE presynaptically inhibits GABA(A) receptor mediated IPSCs in low drinkers, but not in high drinkers or naive animals, most likely through activation of DORs on GABA terminals. This DOR-mediated inhibition of IPSCs also correlates inversely with behavioral correlates of anxiety measured in the elevated plus maze. In contrast, presynaptic inhibition of VTA GABA(A) IPSCs by the mu-opioid receptor agonist DAMGO is significantly reduced in both high- and low-drinking rats (<30%) compared with age-matched nondrinking controls (>70%). Together, our findings demonstrate the protective nature of VTA DORs and identify an important new target for therapeutic intervention for alcoholism.

Protein kinase and phosphatase modulation of quail brain GABA(A) and non-NMDA receptors co-expressed in Xenopus oocytes.

PubMed

Moon, C; Fraser, S P; Djamgoz, M B

2000-02-01

The GABA(A) receptor and the non-NMDA subtype of the ionotropic glutamate receptor were co-expressed in Xenopus oocytes by injection of quail brain mRNA. The oocytes were treated with various protein kinase (PK) and protein phosphatase (PP) activators and inhibitors and the effects on receptor functioning were monitored. Two phorbol esters, 4-beta-phorbol 12-myristate-13-acetate (PMA) and 4-beta-phorbol 12,13-dibutyrate (PDBu); the cGMP-dependent PK activators sodium nitroprusside (SNP) and S-nitrosoglutathione (SNOG); and the PP inhibitor okadaic acid (OA) reduced the amplitude of the GABA-induced currents, whilst the PK inhibitor staurosporine potentiated it. In addition, PMA, PDBu, SNP, and OA reduced the desensitization of the GABA-induced response. Identical treatments generally had similar but less pronounced effects on responses generated by kainate (KA) but the desensitization characteristic of the non-NMDA receptor was not affected. None of the treatments had any effect on the reversal potentials of the induced currents. Immunoblots revealed that the oocytes express endogenous PKG and guanylate cyclase. The results are discussed in terms of the molecular structures of GABA(A) and non-NMDA receptors and the potential functional consequences of phosphorylation/dephosphorylation.

Sensitivity of GBM cells to cAMP agonist-mediated apoptosis correlates with CD44 expression and agonist resistance with MAPK signaling.

PubMed

Daniel, Paul M; Filiz, Gulay; Mantamadiotis, Theo

2016-12-01

In some cell types, activation of the second messenger cAMP leads to increased expression of proapoptotic Bim and subsequent cell death. We demonstrate that suppression of the cAMP pathway is a common event across many cancers and that pharmacological activation of cAMP in glioblastoma (GBM) cells leads to enhanced BIM expression and apoptosis in specific GBM cell types. We identified the MAPK signaling axis as the determinant of cAMP agonist sensitivity in GBM cells, with high MAPK activity corresponding to cAMP resistance and low activity corresponding to sensitization to cAMP-induced apoptosis. Sensitive cells were efficiently killed by cAMP agonists alone, while targeting both the cAMP and MAPK pathways in resistant GBM cells resulted in efficient apoptosis. We also show that CD44 is differentially expressed in cAMP agonist-sensitive and -resistant cells. We thus propose that CD44 may be a useful biomarker for distinguishing tumors that may be sensitive to cAMP agonists alone or cAMP agonists in combination with other pathway inhibitors. This suggests that using existing chemotherapeutic compounds in combination with existing FDA-approved cAMP agonists may fast track trials toward improved therapies for difficult-to-treat cancers, such as GBM.

Metabotropic glutamate receptor 5 upregulation in children with autism is associated with underexpression of both Fragile X mental retardation protein and GABAA receptor beta 3 in adults with autism

PubMed Central

Fatemi, S. Hossein; Folsom, Timothy D.; Kneeland, Rachel E.; Liesch, Stephanie B.

2011-01-01

Recent work has demonstrated the impact of dysfunction of the GABAergic signaling system in brain and the resultant behavioral pathologies in subjects with autism. In animal models, altered expression of Fragile X mental retardation protein (FMRP) has been linked to downregulation of GABA receptors. Interestingly, the autistic phenotype is also observed in individuals with Fragile X syndrome. This study was undertaken to test previous theories relating abnormalities in levels of FMRP to GABAA receptor underexpression. We observed a significant reduction in levels of FMRP in the vermis of adults with autism. Additionally, we found that levels of metabotropic glutamate receptor 5 (mGluR5) protein were significantly increased in vermis of children with autism vs. age and postmortem interval (PMI) matched controls. There was also a significant decrease in level of GABAA receptor beta 3 (GABRβ3) protein in vermis of adult subjects with autism. Finally, we found significant increases in glial fibrillary acidic protein (GFAP) in vermis of both children and adults with autism when compared with controls. Taken together, our results provide further evidence that altered FMRP expression and increased mGluR5 protein production potentially leads to altered expression of GABAA receptors. PMID:21901840

Magnolol, a major bioactive constituent of the bark of Magnolia officinalis, induces sleep via the benzodiazepine site of GABA(A) receptor in mice.

PubMed

Chen, Chang-Rui; Zhou, Xu-Zhao; Luo, Yan-Jia; Huang, Zhi-Li; Urade, Yoshihiro; Qu, Wei-Min

2012-11-01

Magnolol (6,6',7,12-tetramethoxy-2,2'-dimethyl-1-beta-berbaman, C(18)H(18)O(2)), an active ingredient of the bark of Magnolia officinalis, has been reported to exert potent anti-epileptic effects via the GABA(A) receptor. The receptor also mediates sleep in humans and animals. The aim of this study was to determine whether magnolol could modulate sleep behaviors by recording EEG and electromyogram in mice. The results showed that magnolol administered i.p. at a dose of 5 or 25 mg/kg could significantly shorten the sleep latency, increase the amount of non-rapid eye movement (non-REM, NREM) and rapid eye movement (REM) sleep for 3 h after administration with an increase in the number of NREM and REM sleep episodes. Magnolol at doses of 5 and 25 mg/kg increased the number of bouts of wakefulness but decreased their duration. On the other hand, magnolol increased the number of state transitions from wakefulness to NREM sleep and subsequently from NREM sleep to wakefulness. Immunohistochemical study showed that magnolol increased c-Fos expression in the neurons of ventrolateral preoptic area, a sleep center in the anterior hypothalamus, and decreased c-Fos expression in the arousal tuberomammillary nucleus, which was located in the caudolateral hypothalamus. The sleep-promoting effects and changes in c-Fos induced by magnolol were reversed by flumazenil, an antagonist at the benzodiazepine site of the GABA(A) receptor. These results indicate that magnolol increased NREM and REM sleep via the GABA(A) receptor. Copyright © 2012 Elsevier Ltd. All rights reserved.

Role of amino acids in salivation and the localization of their receptors in the rat salivary gland.

PubMed

Shida, T; Kondo, E; Ueda, Y; Takai, N; Yoshida, Y; Araki, T; Kiyama, H; Tohyama, M

1995-11-01

The distribution of gamma-aminobutyric acid (GABA) receptor subunits such as GABAAR-gamma 1 and GABAAR-gamma 2, and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) type receptor subunits such as GluR-1, GluR-2/3 and GluR-4, and N-methyl-D-aspartic acid (NMDA) type subunits such as NR1 were investigated by immunocytochemistry. Furthermore, the roles of these amino acids, GABA and glutamate, on salivation were analyzed in the rat submandibular and sublingual glands. Some similarities were observed in the distribution patterns of GABAA type receptors and AMPA receptors. In the submandibular ganglion cells, collecting ducts and striated ducts, these subunits were expressed strongly; however, there were some differences in their expression patterns between the submandibular and sublingual gland acinar cells. Since these receptor subunits were expressed in the acinar cell bodies of the submandibular gland, they were not expressed in the acinar cells but were expressed in the myoepithelial cells in the sublingual gland. On the other hand, no NR1 expression was observed. To examine the roles of GABA and glutamate in salivation, the submandibular and sublingual glands were perfused partially with Ringer's solution via a facial artery to avoid systemic influence, and substrates were infused into the perfusion solution. No salivary secretion was evoked by GABA or glutamate infusion in the absence of electrical stimulation (2-3 V, 5 ms, 20 Hz). Salivary flow evoked by electrical stimulation of the chorda-lingual nerve caused significant inhibition by GABA (10(-6), 10(-5), 10(-4) and 10(-3) M) and the GABAAR agonist muscimol 10(-3) and 10(-6) M) (n = 6, P < 0.05). Such GABA-induced inhibition was antagonized by the GABAAR antagonists bicuculline (BCC; 10(-6) and 10(-3) M) and picrotoxin (PTX; 10(-6) and 10(-3) M). On the other hand, salivary flow evoked by electrical stimulation (8-10 V, 5 ms, 20 Hz) of the superior cervical ganglion (SCG) was not affected by

Noradrenaline Triggers GABAA Inhibition of Bed Nucleus of the Stria Terminalis Neurons Projecting to the Ventral Tegmental Area

PubMed Central

Dumont, Éric C.; Williams, John T.

2014-01-01

The lateral part of the ventral bed nucleus of the stria terminalis (vlBNST) is a critical site for the antiaversive effects of noradrenergic drugs during opioid withdrawal. The objective of the present study is to identify the cellular action(s) of noradrenaline in the vlBNST after withdrawal from a 5 d treatment with morphine. The vlBNST is a heterogeneous cell group with multiple efferent projections. Therefore, neurons projecting to the midbrain were identified by retrograde transport of fluorescent microspheres injected in the ventral tegmental area (VTA). Whole-cell voltage clamp recordings of these neurons and of those sharing physiological properties were done in brain slices. Noradrenaline activated α1-adrenergic receptors to increase GABAA-IPSC frequency. Noradrenaline produced a similar increase in GABAA-IPSCs during acute opioid withdrawal, but this increase resulted from activation of β-adrenergic receptors, adenylyl cyclase, and protein kinase A, as well as α1-adrenergic receptors. Given that neurons in the vlBNST send an excitatory projection to the VTA, noradrenaline may reduce excitatory drive to mesolimbic dopamine cells. This mechanism might contribute to the withdrawal-induced inhibition of dopamine neurons and explain how noradrenergic drugs microinjected into the vlBNST reduce aversive aspects of opioid withdrawal. PMID:15385602

Gamma-aminobutyric acid (GABA) stimulates pancreatic cancer growth through overexpressing GABAA receptor pi subunit.

PubMed

Takehara, Akio; Hosokawa, Masayo; Eguchi, Hidetoshi; Ohigashi, Hiroaki; Ishikawa, Osamu; Nakamura, Yusuke; Nakagawa, Hidewaki

2007-10-15

Gamma-aminobutyric acid (GABA) functions primarily as an inhibitory neurotransmitter in the mature central nervous system, and GABA/GABA receptors are also present in nonneural tissues, including cancer, but their precise function in nonneuronal or cancerous cells has thus far been poorly defined. Through the genome-wide cDNA microarray analysis of pancreatic ductal adenocarcinoma (PDAC) cells as well as subsequent reverse transcription-PCR and Northern blot analyses, we identified the overexpression of GABA receptor pi subunit (GABRP) in PDAC cells. We also found the expression of this peripheral type GABAA receptor subunit in few adult human organs. Knockdown of endogenous GABRP expression in PDAC cells by small interfering RNA attenuated PDAC cell growth, suggesting its essential role in PDAC cell viability. Notably, the addition of GABA into the cell culture medium promoted the proliferation of GABRP-expressing PDAC cells, but not GABRP-negative cells, and GABAA receptor antagonists inhibited this growth-promoting effect by GABA. The HEK293 cells constitutively expressing exogenous GABRP revealed the growth-promoting effect of GABA treatment. Furthermore, GABA treatment in GABRP-positive cells increased intracellular Ca2+ levels and activated the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/Erk) cascade. Clinical PDAC tissues contained a higher level of GABA than normal pancreas tissues due to the up-regulation of glutamate decarboxylase 1 expression, suggesting their autocrine/paracrine growth-promoting effect in PDACs. These findings imply that GABA and GABRP could play important roles in PDAC development and progression, and that this pathway can be a promising molecular target for the development of new therapeutic strategies for PDAC.

Modification of opiate agonist binding by pertussis toxin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Abood, M.E.; Lee, N.M.; Loh, H.H.

1986-03-05

Opiate agonist binding is decreased by GTP, suggesting the possible involvement of GTP binding proteins in regulation of opiate receptor binding. This possibility was addressed by asking whether pertussis toxin treatment, which results in ADP-ribosylation and modification of G proteins, would alter opiate agonist binding. The striatum was chosen for the initial brain area to be studied, since regulation of opiate action in this area had been shown to be modified by pertussis toxin. Treatment of striatal membranes with pertussis toxin results in up to a 55% decrease in /sup 3/(H)-DADLE binding as compared with membranes treated identically without toxin.more » This corresponds to a near complete ADP-ribosylation of both G proteins in the striatal membrane. The decrease in agonist binding appears to be due to an altered affinity of the receptor for agonist as opposed to a decrease in the number of sites. This effect of pertussis toxin on opiate agonist binding demonstrates the actual involvement of G proteins in regulation of opiate receptor binding.« less

Structural determinants for antagonist pharmacology that distinguish the rho1 GABAC receptor from GABAA receptors.

PubMed

Zhang, Jianliang; Xue, Fenqin; Chang, Yongchang

2008-10-01

GABA receptor (GABAR) types C (GABACR) and A (GABAAR) are both GABA-gated chloride channels that are distinguished by their distinct competitive antagonist properties. The structural mechanism underlying these distinct properties is not well understood. In this study, using previously identified binding residues as a guide, we made individual or combined mutations of nine binding residues in the rho1 GABACR subunit to their counterparts in the alpha1beta2gamma2 GABAAR or reverse mutations in alpha1 or beta2 subunits. The mutants were expressed in Xenopus laevis oocytes and tested for sensitivities of GABA-induced currents to the GABAA and GABAC receptor antagonists. The results revealed that bicuculline insensitivity of the rho1 GABACR was mainly determined by Tyr106, Phe138 and Phe240 residues. Gabazine insensitivity of the rho1 GABACR was highly dependent on Tyr102, Tyr106, and Phe138. The sensitivity of the rho1 GABACR to 3-aminopropyl-phosphonic acid and its analog 3-aminopropyl-(methyl)phosphinic acid mainly depended on residues Tyr102, Val140, FYS240-242, and Phe138. Thus, the residues Tyr102, Tyr106, Phe138, and Phe240 in the rho1 GABACR are major determinants for its antagonist properties distinct from those in the GABAAR. In addition, Val140 in the GABACR contributes to 3-APA binding. In conclusion, we have identified the key structural elements underlying distinct antagonist properties for the GABACR. The mechanistic insights were further extended and discussed in the context of antagonists docking to the homology models of GABAA or GABAC receptors.

Antagonism of the Ethanol-Like Discriminative Stimulus Effects of Ethanol, Pentobarbital, and Midazolam in Cynomolgus Monkeys Reveals Involvement of Specific GABAA Receptor SubtypesS⃞

PubMed Central

Rogers, Laura S. M.; Grant, Kathleen A.

2009-01-01

The γ-aminobutyric acid (GABA)A receptors mediating the discriminative stimulus effects of ethanol were studied by comparing the potency of ethyl-8-azido-5,6-dihydro-5-methyl-6-oxo-4H-imidazol(1,5-a)benzodiazepine-3-carboxylate (Ro15-4513) and ethyl 8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazol(1,5-a)-benzodiazepine-3-carboxylate (flumazenil, Ro15-1788) to antagonize ethanol, pentobarbital (PB), and midazolam substitution for ethanol. Ro15-4513 has high affinity for receptors containing α4/6 and α5 subunits and lower affinity for α1, α2, and α3 subunits. Flumazenil is nonselective for GABAA receptors containing α1, α2, α3, and α5 subunits and has low affinity for α4/6-containing receptors. Male (n = 9) and female (n = 8) cynomolgus monkeys (Macaca fascicularis) were trained to discriminate ethanol (1.0 or 2.0 g/kg i.g., 30-min pretreatment) from water. Ethanol, PB, and midazolam dose-dependently substituted for ethanol (80% ethanol-appropriate responding). Ro15-4513 (0.003–0.56 mg/kg i.m., 5-min pretreatment) shifted the ethanol, PB, and midazolam dose-response functions rightward in a vast majority of monkeys tested (15/15, 16/17, and 11/12, respectively). In contrast, flumazenil (0.30–10.0 mg/kg i.m., 5-min pretreatment) shifted the ethanol, PB, and midazolam dose-response functions rightward in 9 of 16, 12 of 16, and 7 of 9 monkeys tested, respectively. In the monkeys showing antagonism with both Ro15-4513 and flumazenil, ethanol and PB substitution were antagonized more potently by Ro15-4513 than by flumazenil, whereas midazolam substitution was antagonized with similar potency. There were no sex or training dose differences, with the exception that flumazenil failed to antagonize ethanol substitution in males trained to discriminate 2.0 g/kg ethanol. GABAA receptors with high affinity for Ro15-4513 (i.e., containing α4/6 and α5 subunits) may be particularly important mediators of the multiple discriminative stimulus effects of ethanol

The PPARgamma agonist FMOC-L-leucine protects both mature and immature brain.

PubMed

Maurois, Pierre; Rocchi, Stéphane; Pages, Nicole; Bac, Pierre; Stables, James P; Gressens, Pierre; Vamecq, Joseph

2008-01-01

(N-[9-fluorenylmethoxycarbonyl]-)-L-leucine (FMOC-L-leucine) and rosiglitazone, two ligands of peroxisome proliferator-activated receptor gamma (PPARgamma), were evaluated in mature (adult mice) and immature (pups) brain injury models. In adult magnesium-deficient mice, a model responsive to both neuroprotective and anti-seizure compounds, FMOC-L-leucine, but not rosiglitazone, protected against audiogenic seizures. The protection afforded by FMOC-L-leucine was alleviated by the PPARgamma antagonist GW9662 (1-2 mg/kg) and was induced in 50% animals by 4.8+/-1.2 mg/kg. At this dose, FMOC-L-leucine modified audiogenic seizure phase durations in convulsing mice differently than prototype antiepileptic drugs did. FMOC-L-leucine (up to 100 mg/kg) was inactive in the 6 Hz seizure test, an adult animal model largely responsive to anti-seizure drugs. In a model of neonatal brain injury, FMOC-L-leucine (4 microg/kg) was neuroprotective against cerebral ibotenate toxicity. It reduced significantly the size of lesions in grey but not in white matter, while rosiglitazone (10 microg/kg) was inactive. Taken as a whole, the present data support neuroprotective potentialities of FMOC-L-leucine towards both mature and immature brain. The PPAR-based protection of immature brain is more important as it is known that classic adult brain protectants (GABA(A) activators, N-methyl-D-aspartate and sodium channel blockers) may be toxic for immature brain. The PPARgamma agonist FMOC-L-leucine is likely to be devoid of these classic protective mechanisms because of its inactivity in the 6 Hz seizure test, its activity in the audiogenic test being explained by neuroprotective rather than intrinsic anti-seizure mechanisms. Targeting PPARs might be thus a promising way to protect immature brain.

Reciprocity of agonistic support in ravens.

PubMed

Fraser, Orlaith N; Bugnyar, Thomas

2012-01-01

Cooperative behaviour through reciprocation or interchange of valuable services in primates has received considerable attention, especially regarding the timeframe of reciprocation and its ensuing cognitive implications. Much less, however, is known about reciprocity in other animals, particularly birds. We investigated patterns of agonistic support (defined as a third party intervening in an ongoing conflict to attack one of the conflict participants, thus supporting the other) in a group of 13 captive ravens, Corvus corax. We found support for long-term, but not short-term, reciprocation of agonistic support. Ravens were more likely to support individuals who preened them, kin and dominant group members. These results suggest that ravens do not reciprocate on a calculated tit-for-tat basis, but aid individuals from whom reciprocated support would be most useful and those with whom they share a good relationship. Additionally, dyadic levels of agonistic support and consolation (postconflict affiliation from a bystander to the victim) correlated strongly with each other, but we found no evidence to suggest that receiving agonistic support influences the victim's likelihood of receiving support (consolation) after the conflict ends. Our findings are consistent with an emotionally mediated form of reciprocity in ravens and provide additional support for convergent cognitive evolution in birds and mammals.

Reciprocity of agonistic support in ravens

PubMed Central

Fraser, Orlaith N.; Bugnyar, Thomas

2012-01-01

Cooperative behaviour through reciprocation or interchange of valuable services in primates has received considerable attention, especially regarding the timeframe of reciprocation and its ensuing cognitive implications. Much less, however, is known about reciprocity in other animals, particularly birds. We investigated patterns of agonistic support (defined as a third party intervening in an ongoing conflict to attack one of the conflict participants, thus supporting the other) in a group of 13 captive ravens, Corvus corax. We found support for long-term, but not short-term, reciprocation of agonistic support. Ravens were more likely to support individuals who preened them, kin and dominant group members. These results suggest that ravens do not reciprocate on a calculated tit-for-tat basis, but aid individuals from whom reciprocated support would be most useful and those with whom they share a good relationship. Additionally, dyadic levels of agonistic support and consolation (postconflict affiliation from a bystander to the victim) correlated strongly with each other, but we found no evidence to suggest that receiving agonistic support influences the victim’s likelihood of receiving support (consolation) after the conflict ends. Our findings are consistent with an emotionally mediated form of reciprocity in ravens and provide additional support for convergent cognitive evolution in birds and mammals. PMID:22298910

Identification and cloning of a gamma 3 subunit splice variant of the human GABA(A) receptor.

PubMed

Poulsen, C F; Christjansen, K N; Hastrup, S; Hartvig, L

2000-05-31

cDNA sequences encoding two forms of the GABA(A) gamma 3 receptor subunit were cloned from human hippocampus. The nucleotide sequences differ by the absence (gamma 3S) or presence (gamma 3L) of 18 bp located in the presumed intracellular loop between transmembrane region (TM) III and IV. The extra 18 bp in the gamma 3L subunit generates a consensus site for phosphorylation by protein kinase C (PKC). Analysis of human genomic DNA encoding the gamma 3 subunit reveals that the 18 bp insert is contiguous with the upstream proximal exon.

The central GABAergic system and control of food intake under different experimental conditions.

PubMed

Olgiati, V R; Netti, C; Guidobono, F; Pecile, A

1980-01-01

Intracerebroventricular injections of gamma-aminobutyric acid (GABA) and of the GABA-transaminase inhibitor, ethanolamine-O-sulphate (EOS), decreased the food intake of freely-fed (GABA and EOS) and food-deprived rats (EOS). The effect, still evident 24 h after treatment, was not decreased by the GABA receptor-blocker bicuculline. In contrast, intracerebroventricular injections of the GABA receptor-agonist, muscimol, caused an increase in food intake of freely-fed rats that was antagonized by bicuculline. The eating of animals receiving only bicuculline was stimulated in free-feeding and depressed in food-deprived conditions. These opposite results suggest that muscimol binds preferentially to some GABA receptors, probably those within the satiety-controlling areas (i.e. ventromedial hypothalamus), and that bicuculline influences mainly those postsynaptic neurons where GABAergic inputs prevail. These observations and the data from EOS- and GABA-treated rats provide evidence for involvement of GABA neurons in the regulation of feeding behaviour. The balance of the different effects produced in each of these areas by this modulation appears to be a decrease in feeding behaviour.

Mechanisms of inverse agonist action at D2 dopamine receptors

PubMed Central

Roberts, David J; Strange, Philip G

2005-01-01

Mechanisms of inverse agonist action at the D2(short) dopamine receptor have been examined. Discrimination of G-protein-coupled and -uncoupled forms of the receptor by inverse agonists was examined in competition ligand-binding studies versus the agonist [3H]NPA at a concentration labelling both G-protein-coupled and -uncoupled receptors. Competition of inverse agonists versus [3H]NPA gave data that were fitted best by a two-binding site model in the absence of GTP but by a one-binding site model in the presence of GTP. Ki values were derived from the competition data for binding of the inverse agonists to G-protein-uncoupled and -coupled receptors. Kcoupled and Kuncoupled were statistically different for the set of compounds tested (ANOVA) but the individual values were different in a post hoc test only for (+)-butaclamol. These observations were supported by simulations of these competition experiments according to the extended ternary complex model. Inverse agonist efficacy of the ligands was assessed from their ability to reduce agonist-independent [35S]GTPγS binding to varying degrees in concentration–response curves. Inverse agonism by (+)-butaclamol and spiperone occurred at higher potency when GDP was added to assays, whereas the potency of (−)-sulpiride was unaffected. These data show that some inverse agonists ((+)-butaclamol, spiperone) achieve inverse agonism by stabilising the uncoupled form of the receptor at the expense of the coupled form. For other compounds tested, we were unable to define the mechanism. PMID:15735658

4,5-Substituted 3-Isoxazolols with Insecticidal Activity Act as Competitive Antagonists of Housefly GABA Receptors.

PubMed

Liu, Genyan; Ozoe, Fumiyo; Furuta, Kenjiro; Ozoe, Yoshihisa

2015-07-22

The insect GABA receptor (GABAR), which is composed of five RDL subunits, represents an important target for insecticides. A series of 4,5-disubstituted 3-isoxazolols, including muscimol analogues, were synthesized and examined for their activities against four splice variants (ac, ad, bc, and bd) of housefly GABARs expressed in Xenopus oocytes. Muscimol was a more potent agonist than GABA in all four splice variants, whereas synthesized analogues did not exhibit agonism but rather antagonism in housefly GABARs. The introduction of bicyclic aromatic groups at the 4-position of muscimol and the simultaneous replacement of the aminomethyl group with a carbamoyl group at the 5-position to afford six 4-aryl-5-carbamoyl-3-isoxazolols resulted in compounds that exhibited significantly enhanced antagonism with IC50 values in the low micromolar range in the ac variant. The inhibition of GABA-induced currents by 100 μM analogues was approximately 1.5-4-fold greater in the ac and bc variants than in the ad and bd variants. 4-(3-Biphenylyl)-5-carbamoyl-3-isoxazolol displayed competitive antagonism, with IC50 values of 30, 34, 107, and 96 μM in the ac, bc, ad, and bd variants, respectively, and exhibited moderate insecticidal activity against houseflies, with an LD50 value of 5.6 nmol/fly. These findings suggest that these 3-isoxazolol analogues are novel lead compounds for the design and development of insecticides that target the orthosteric site of housefly GABARs.

A neuronal mechanism of propofol-induced central respiratory depression in newborn rats.

PubMed

Kashiwagi, Masanori; Okada, Yasumasa; Kuwana, Shun-Ichi; Sakuraba, Shigeki; Ochiai, Ryoichi; Takeda, Junzo

2004-07-01

The neural mechanisms of propofol-induced central respiratory depression remain poorly understood. In the present study, we studied these mechanisms and the involvement of gamma-aminobutyric acid (GABA)A receptors in propofol-induced central respiratory depression. The brainstem and the cervical spinal cord of 1- to 4-day-old rats were isolated, and preparations were maintained in vitro with oxygenated artificial cerebrospinal fluid. Rhythmic inspiratory burst activity was recorded from the C4 spinal ventral root. The activity of respiratory neurons in the ventrolateral medulla was recorded using a perforated patch-clamp technique. We found that bath-applied propofol decreased C4 inspiratory burst rate, which could be reversed by the administration of a GABAA antagonist, bicuculline. Propofol caused resting membrane potentials to hyperpolarize and suppressed the firing of action potentials in preinspiratory and expiratory neurons. In contrast, propofol had little effect on resting membrane potentials and action potential firing in inspiratory neurons. Our findings suggest that the depressive effects of propofol are, at least in part, mediated by the agonistic action of propofol on GABAA receptors. It is likely that the GABAA receptor-mediated hyperpolarization of preinspiratory neurons serves as the neuronal basis of propofol-induced respiratory depression in the newborn rat.

Negative modulation of α5 GABAA receptors in rats may partially prevent memory impairment induced by MK-801, but not amphetamine- or MK-801-elicited hyperlocomotion

PubMed Central

Stamenić, Tamara Timić; Joksimović, Srdjan; Biawat, Poonam; Stanković, Tamara; Marković, Bojan; Cook, James M; Savić, Miroslav M

2016-01-01

Reportedly, negative modulation of α5 GABAA receptors may improve cognition in normal and pharmacologically-impaired animals, and such modulation has been proposed as an avenue for treatment of cognitive symptoms in schizophrenia. This study assessed the actions of PWZ-029, administered at doses (2, 5 and 10 mg/kg) at which it reached micromolar concentrations in brain tissue with estimated free concentrations adequate for selective modulation of α5 GABAA receptors, in three cognitive tasks in male Wistar rats acutely treated with the noncompetitive N-methyl-D-aspartate – receptor antagonist, MK-801 (0.1 mg/kg), as well in tests of locomotor activity potentiated by MK-801 (0.2 mg/kg) or amphetamine (0.5 mg/kg). In a hormetic-like manner, only 5 mg/kg PWZ-029 reversed MK-801-induced deficits in novel object recognition test (visual recognition memory), whereas in the Morris water maze, the 2 mg/kg dose of PWZ-029 exerted partial beneficial effects on spatial learning impairment. PWZ-029 did not affect recognition memory deficits in social novelty discrimination procedure. Motor hyperactivity induced with MK-801 or amphetamine was not preventable by PWZ-029. Our results show that certain MK-801-induced memory deficits can be ameliorated by negative modulation of α5 GABAA receptors, and point to the need for further elucidation of their translational relevance to cognitive deterioration in schizophrenia. PMID:26105958

Negative modulation of α₅ GABAA receptors in rats may partially prevent memory impairment induced by MK-801, but not amphetamine- or MK-801-elicited hyperlocomotion.

PubMed

Timić Stamenić, Tamara; Joksimović, Srdjan; Biawat, Poonam; Stanković, Tamara; Marković, Bojan; Cook, James M; Savić, Miroslav M

2015-09-01

Reportedly, negative modulation of α5 GABAA receptors may improve cognition in normal and pharmacologically-impaired animals, and such modulation has been proposed as an avenue for treatment of cognitive symptoms in schizophrenia. This study assessed the actions of PWZ-029, administered at doses (2, 5, and 10 mg/kg) at which it reached micromolar concentrations in brain tissue with estimated free concentrations adequate for selective modulation of α5 GABAA receptors, in three cognitive tasks in male Wistar rats acutely treated with the noncompetitive N-methyl-d-aspartate receptor antagonist, MK-801 (0.1 mg/kg), as well in tests of locomotor activity potentiated by MK-801 (0.2 mg/kg) or amphetamine (0.5 mg/kg). In a hormetic-like manner, only 5 mg/kg PWZ-029 reversed MK-801-induced deficits in novel object recognition test (visual recognition memory), whereas in the Morris water maze, the 2 mg/kg dose of PWZ-029 exerted partial beneficial effects on spatial learning impairment. PWZ-029 did not affect recognition memory deficits in social novelty discrimination procedure. Motor hyperactivity induced with MK-801 or amphetamine was not preventable by PWZ-029. Our results show that certain MK-801-induced memory deficits can be ameliorated by negative modulation of α5 GABAA receptors, and point to the need for further elucidation of their translational relevance to cognitive deterioration in schizophrenia. © The Author(s) 2015.

GABAergic anxiolytic drug in water increases migration behaviour in salmon

NASA Astrophysics Data System (ADS)

Hellström, Gustav; Klaminder, Jonatan; Finn, Fia; Persson, Lo; Alanärä, Anders; Jonsson, Micael; Fick, Jerker; Brodin, Tomas

2016-12-01

Migration is an important life-history event in a wide range of taxa, yet many migrations are influenced by anthropogenic change. Although migration dynamics are extensively studied, the potential effects of environmental contaminants on migratory physiology are poorly understood. In this study we show that an anxiolytic drug in water can promote downward migratory behaviour of Atlantic salmon (Salmo salar) in both laboratory setting and in a natural river tributary. Exposing salmon smolt to a dilute concentration of a GABAA receptor agonist (oxazepam) increased migration intensity compared with untreated smolt. These results implicate that salmon migration may be affected by human-induced changes in water chemical properties, such as acidification and pharmaceutical residues in wastewater effluent, via alterations in the GABAA receptor function.

Human brain somatostatin release from isolated cortical nerve endings and its modulation through GABAB receptors.

PubMed Central

Bonanno, G.; Gemignani, A.; Schmid, G.; Severi, P.; Cavazzani, P.; Raiteri, M.

1996-01-01

1. The release of somatostatin-like immunoreactivity (SRIF-LI) in the human brain was studied in synaptosomal preparations from fresh neocortical specimens obtained from patients undergoing neurosurgery to remove deeply sited tumours. 2. The basal outflow of SRIF-LI from superfused synaptosomes was increased about 3 fold during exposure to a depolarizing medium containing 15 mM KCl. The K(+)-evoked overflow of SRIF-LI was almost totally dependent on the presence of Ca2+ in the superfusion medium. 3. The GABAB receptor agonist, (-)-baclofen (0.3 - 100 microM), inhibited the overflow of SRIF-LI in a concentration-dependent manner (EC50 = 1.84 +/- 0.20 microM; maximal effect: about 50%). The novel GABAB receptor ligand, 3-aminopropyl(difluoromethyl)phosphinic acid (CGP 47656) mimicked (-)-baclofen in inhibiting the SRIF-LI overflow (EC50 = 3.06 +/- 0.52 microM; maximal effect: about 50%), whereas the GABAA receptor agonist, muscimol, was ineffective up to 100 microM. 4. The inhibition by 10 microM (-)-baclofen of the K(+)-evoked SRIF-LI overflow was concentration-dependently prevented by two selective GABAB receptor antagonists, 3-amino-propyl (diethoxymethyl)-phosphinic acid (CGP 35348) (IC50 = 24.40 +/- 2.52 microM) and [3-[[(3,4-dichlorophenyl) methyl]amino]propyl] (diethoxymethyl) phosphinic acid (CGP 52432) (IC50 = 0.06 +/- 0.005 microM). 5. The inhibition of SRIF-LI overflow caused by 10 microM CGP 47656 was abolished by 1 microM CGP 52432. 6. When human synaptosomes were labelled with [3H]-GABA and depolarized in superfusion with 15 mM KCl, the inhibition by 10 microM (-)-baclofen of the depolarization-evoked [3H]-GABA overflow was largely prevented by 10 microM CGP 47656 which therefore behaved as an autoreceptor antagonist. 7. In conclusion: (a) the characteristics of SRIF-LI release from synaptosomal preparations of human neocortex are compatible with a neuronal origin; (b) the nerve terminals releasing the neuropeptide possess inhibitory receptors of the

Neurokinin-1 receptor activation in Botzinger complex evokes bradypnoea.

PubMed

Fong, Angelina Y; Potts, Jeffrey T

2006-09-15

In the present study, we examined the role of the neurokinin-1 receptor (NK1R) in the modulation of respiratory rhythm in a functionally identified bradypnoeic region of the ventral respiratory group (VRG) in the in situ arterially perfused juvenile rat preparation. In electrophysiologically and functionally identified bradypnoeic sites corresponding to the Bötzinger complex (BötC), microinjection of the selective NK1R agonist [Sar(9)-Met(O(2))(11)]-substance P (SSP) produced a significant reduction in phrenic frequency mediated exclusively by an increase in expiratory duration (T(E)). The reduction was characterized by a significant increase in postinspiratory (post-I) duration with no effect on either late-expiratory duration (E2) or inspiratory duration (T(I)). In contrast, in a functionally identified tachypnoeic region, corresponding to the preBötzinger complex (Pre-BötC), control microinjection of SSP elicited tachypnoea. Pretreatment with the NK1R antagonist CP99994 in the BötC significantly attenuated the bradypnoeic response to SSP injection and blunted the increase in T(E) duration. This effect of SSP mimicked the extension of T(E) produced by activation of the Hering-Breuer reflex. Therefore, we hypothesized that activation of NK1Rs in the BötC is requisite for the expiratory-lengthening effect of the Hering-Breuer reflex. Unilateral electrical stimulation of the cervical vagus nerve produced bradypnoea by exclusively extending T(E). Ipsilateral blockade of NK1Rs by CP99994 following blockade of the contralateral BötC by the GABA(A) receptor agonist muscimol significantly reduced the extension of T(E) produced by vagal stimulation. Results from the present study demonstrate that selective activation of NK1Rs in a functionally identified bradypnoeic region of the VRG can depress respiratory frequency by selectively lengthening post-I duration and provide evidence that endogenous activation of NK1Rs in the BötC appears to be involved in the

Ghrelin and motilin receptor agonists: time to introduce bias into drug design.

PubMed

Sanger, G J

2014-02-01

Ghrelin and motilin receptor agonists increase gastric motility and are attractive drug targets. However, 14 years after the receptors were described (18-24 years since ligands became available) the inactivity of the ghrelin agonist TZP-102 in patients with gastroparesis joins the list of unsuccessful motilin agonists. Fundamental questions must be asked. Pustovit et al., have now shown that the ghrelin agonist ulimorelin evokes prolonged increases in rat colorectal propulsion yet responses to other ghrelin agonists fade. Similarly, different motilin agonists induce short- or long-lasting effects in a cell-dependent manner. Together, these and other data create the hypothesis that the receptors can be induced to preferentially signal ('biased agonism') via particular pathways to evoke different responses with therapeutic advantages/disadvantages. Biased agonism has been demonstrated for ghrelin. Are motilin agonists which cause long-lasting facilitation of human stomach cholinergic function (compared with motilin) biased agonists (e.g., camicinal, under development for patients with gastric hypo-motility)? For ghrelin, additional complications exist because the therapeutic aims/mechanisms of action are uncertain, making it difficult to select the best (biased) agonist. Will ghrelin agonists be useful treatments of nausea and/or as suggested by Pustovit et al., chronic constipation? How does ghrelin increase gastric motility? As gastroparesis symptoms poorly correlate with delayed gastric emptying (yet gastro-prokinetic drugs can provide relief: e.g., low-dose erythromycin), would low doses of ghrelin and motilin agonists relieve symptoms simply by restoring neuromuscular rhythm? These questions on design and functions need addressing if ghrelin and motilin agonists are to reach patients as drugs. © 2014 John Wiley & Sons Ltd.

Reverse Induced Fit-Driven MAS-Downstream Transduction: Looking for Metabotropic Agonists.

PubMed

Pernomian, Larissa; Gomes, Mayara S; de Paula da Silva, Carlos H Tomich; Rosa, Joaquin M C

2017-01-01

Protective effects of MAS activation have spurred clinical interests in developing MAS agonists. However, current bases that drive this process preclude that physiological concentrations of peptide MAS agonists induce an atypical signaling that does not reach the metabotropic efficacy of constitutive activation. Canonical activation of MAS-coupled G proteins is only achieved by supraphysiological concentrations of peptide MAS agonists or physiological concentrations of chemically modified analogues. These pleiotropic differences are because of two overlapped binding domains: one non-metabotropic site that recognizes peptide agonists and one metabotropic domain that recognizes modified analogues. It is feasible that supraphysiological concentrations of peptide MAS agonists undergo to chemical modifications required for binding to metabotropic domain. Receptor oligomerization enhances pharmacological parameters coupled to metabotropic signaling. The formation of receptor-signalosome complex makes the transduction of agonists more adaptive. Considering the recent identification of MAS-signalosome, we aimed to postulate the reverse induced fit hypothesis in which MAS-signalosome would trigger chemical modifications required for agonists bind to MAS metabotropic domain. Here we cover rational perspectives for developing novel metabotropic MAS agonists in the view of the reverse induced-fit hypothesis. Predicting a 3D model of MAS metabotropic domain may guide the screening of chemical modifications required for metabotropic efficacy. Pharmacophore-based virtual screening would select potential metabotropic MAS agonists from virtual libraries from human proteome. Rational perspectives that consider reverse induced fit hypothesis during MAS activation for developing metabotropic MAS agonists represents the best approach in providing MAS ligands with constitutive efficacy at physiological concentrations. Copyright© Bentham Science Publishers; For any queries, please email

Subjective Effects of Thiopental in Young Adults with and without a Family History of Alcoholism

PubMed Central

Petrakis, Ismene L; Kerfoot, Karin; Pittman, Brian; Perrino, Albert; Koretski, Julia; Newcomb, Jenelle; Limoncelli, Diana; Acampora, Gregory; Ralevski, Elizabeth

2013-01-01

Background The development of alcohol use disorders is genetically influenced, and may be mediated through differences in the subjective response to alcohol. There is some evidence to suggest that response differences to alcohol could be conveyed by heritable differences in GABAA receptors. The purpose of this study was to investigate whether individuals with a family history positive (FHP) for alcohol dependence would experience alterations in response to the GABAA receptor agonist thiopental, in comparison to family history negative (FHN) subjects. Methods 73 subjects (24 FHP and 49 FHN) between the ages of 21 and 30 years were administered sub-anesthetic doses of the GABAA receptor agonist thiopental and placebo on two separate test days. Various alcohol-related measures were administered, including those examining subjective effects, coordination, and cognition. Results Sub-anesthetic doses of thiopental produced alcohol-like subjective effects, as well as alcohol-like impaired coordination and cognition in healthy subjects. While there were no significant main effects in subjective, coordination, or cognitive effects between FHP and FHN individuals, analysis of peak effects suggested FHP had blunted sedative, but not stimulant effects compared to FHN. Conclusion Thiopental produced alcohol-like effects and perceived similarities to alcohol in healthy individuals. Subtle differences in sedative effects are consistent with reports of blunted FHP response to the negative but not stimulant effects of alcohol. Future studies are needed to better understand how this insight informs our understanding of the heritable risk for alcoholism and the treatment of alcohol use disorders. PMID:24273687

The stressed female brain: neuronal activity in the prelimbic but not infralimbic region of the medial prefrontal cortex suppresses learning after acute stress.

PubMed

Maeng, Lisa Y; Shors, Tracey J

2013-01-01

Women are nearly twice as likely as men to suffer from anxiety and post-traumatic stress disorder (PTSD), indicating that many females are especially vulnerable to stressful life experience. A profound sex difference in the response to stress is also observed in laboratory animals. Acute exposure to an uncontrollable stressful event disrupts associative learning during classical eyeblink conditioning in female rats but enhances this same type of learning process in males. These sex differences in response to stress are dependent on neuronal activity in similar but also different brain regions. Neuronal activity in the basolateral nucleus of the amygdala (BLA) is necessary in both males and females. However, neuronal activity in the medial prefrontal cortex (mPFC) during the stressor is necessary to modify learning in females but not in males. The mPFC is often divided into its prelimbic (PL) and infralimbic (IL) subregions, which differ both in structure and function. Through its connections to the BLA, we hypothesized that neuronal activity within the PL, but not IL, during the stressor is necessary to suppress learning in females. To test this hypothesis, either the PL or IL of adult female rats was bilaterally inactivated with GABAA agonist muscimol during acute inescapable swim stress. About 24 h later, all subjects were trained with classical eyeblink conditioning. Though stressed, females without neuronal activity in the PL learned well. In contrast, females with IL inactivation during the stressor did not learn well, behaving similarly to stressed vehicle-treated females. These data suggest that exposure to a stressful event critically engages the PL, but not IL, to disrupt associative learning in females. Together with previous studies, these data indicate that the PL communicates with the BLA to suppress learning after a stressful experience in females. This circuit may be similarly engaged in women who become cognitively impaired after stressful life

The Basolateral Amygdala Can Mediate the Effects of Fear Memory on Sleep Independently of Fear Behavior and the Peripheral Stress Response

PubMed Central

Wellman, Laurie L.; Fitzpatrick, Mairen E.; Hallum, Olga Y.; Sutton, Amy M.; Williams, Brook L.; Sanford, Larry D.

2016-01-01

Fear conditioning associated with inescapable shock training (ST) and fearful context re-exposure (CR) alone can produce significant behavioral fear, a stress response and alterations in subsequent REM sleep. These alterations may vary among animals and are mediated by the basolateral nucleus of the amygdala (BLA). Here, we used the GABAA agonist, muscimol (Mus), to inactivate BLA prior to CR and examined the effects on sleep, freezing and stress-induced hyperthermia (SIH). Wistar rats (n=28) were implanted with electrodes for recording sleep, data loggers for recording core body temperature, and with cannulae aimed bilaterally into BLA. After recovery, the animals were habituated to the injection procedure and baseline sleep was recorded. On experimental day 1, rats received ST (20 footshocks, 0.8mA, 0.5s duration, 60s interstimulus interval). On experimental day 7, the rats received microinjections (0.5ul) into BLA of either Mus (1.0uM; n = 13) or vehicle (Veh; n = 15) prior to CR (CR1). On experimental day 21, the animals experienced a second CR (CR2) without Mus. For analysis, the rats were separated into 4 groups: (Veh-vulnerable (Veh-Vul; n=8), Veh-resilient (Veh-Res; n=7), Mus-vulnerable (Mus-Vul; n=7), and Mus-resilient (Mus-Res; n=6)) based on whether or not REM was decreased, compared to baseline, during the first 4 h following ST. Pre-CR1 inactivation of BLA did not alter freezing or SIH, but did block the reduction in REM in the Mus-Vul group compared to the Veh-Vul group. These data indicate that BLA is an important region for mediating the effects of fearful memories on sleep. PMID:27818268

A Sensitive Membrane-Targeted Biosensor for Monitoring Changes in Intracellular Chloride in Neuronal Processes

PubMed Central

Watts, Spencer D.; Suchland, Katherine L.; Amara, Susan G.; Ingram, Susan L.

2012-01-01

Background Regulation of chloride gradients is a major mechanism by which excitability is regulated in neurons. Disruption of these gradients is implicated in various diseases, including cystic fibrosis, neuropathic pain and epilepsy. Relatively few studies have addressed chloride regulation in neuronal processes because probes capable of detecting changes in small compartments over a physiological range are limited. Methodology/Principal Findings In this study, a palmitoylation sequence was added to a variant of the yellow fluorescent protein previously described as a sensitive chloride indicator (YFPQS) to target the protein to the plasma membrane (mbYFPQS) of cultured midbrain neurons. The reporter partitions to the cytoplasmic face of the cellular membranes, including the plasma membrane throughout the neurons and fluorescence is stable over 30–40 min of repeated excitation showing less than 10% decrease in mbYFPQS fluorescence compared to baseline. The mbYFPQS has similar chloride sensitivity (k50 =  41 mM) but has a shifted pKa compared to the unpalmitoylated YFPQS variant (cytYFPQS) that remains in the cytoplasm when expressed in midbrain neurons. Changes in mbYFPQS fluorescence were induced by the GABAA agonist muscimol and were similar in the soma and processes of the midbrain neurons. Amphetamine also increased mbYFPQS fluorescence in a subpopulation of cultured midbrain neurons that was reversed by the selective dopamine transporter (DAT) inhibitor, GBR12909, indicating that mbYFPQS is sensitive enough to detect endogenous DAT activity in midbrain dopamine (DA) neurons. Conclusions/Significance The mbYFPQS biosensor is a sensitive tool to study modulation of intracellular chloride levels in neuronal processes and is particularly advantageous for simultaneous whole-cell patch clamp and live-cell imaging experiments. PMID:22506078

Trial Watch: Toll-like receptor agonists for cancer therapy.

PubMed

Vacchelli, Erika; Eggermont, Alexander; Sautès-Fridman, Catherine; Galon, Jérôme; Zitvogel, Laurence; Kroemer, Guido; Galluzzi, Lorenzo

2013-08-01

Toll-like receptors (TLRs) have long been known for their ability to initiate innate immune responses upon exposure to conserved microbial components such as lipopolysaccharide (LPS) and double-stranded RNA. More recently, this family of pattern recognition receptors has been attributed a critical role in the elicitation of anticancer immune responses, raising interest in the development of immunochemotherapeutic regimens based on natural or synthetic TLR agonists. In spite of such an intense wave of preclinical and clinical investigation, only three TLR agonists are currently licensed by FDA for use in cancer patients: bacillus Calmette-Guérin (BCG), an attenuated strain of Mycobacterium bovis that operates as a mixed TLR2/TLR4 agonist; monophosphoryl lipid A (MPL), a derivative of Salmonella minnesota that functions as a potent agonist of TLR4; and imiquimod, a synthetic imidazoquinoline that activates TLR7. One year ago, in the August and September issues of OncoImmunology , we described the main biological features of TLRs and discussed the progress of clinical studies evaluating the safety and therapeutic potential of TLR agonists in cancer patients. Here, we summarize the latest developments in this exciting area of research, focusing on preclinical studies that have been published during the last 13 mo and clinical trials launched in the same period to investigate the antineoplastic activity of TLR agonists.

GABA(A) receptor blockade in dorsomedial and ventromedial nuclei of the hypothalamus evokes panic-like elaborated defensive behaviour followed by innate fear-induced antinociception.

PubMed

Freitas, Renato Leonardo; Uribe-Mariño, Andrés; Castiblanco-Urbina, Maria Angélica; Elias-Filho, Daoud Hibraim; Coimbra, Norberto Cysne

2009-12-11

Dysfunction in the hypothalamic GABAergic system has been implicated in panic syndrome in humans. Furthermore, several studies have implicated the hypothalamus in the elaboration of pain modulation. Panic-prone states are able to be experimentally induced in laboratory animals to study this phenomenon. The aim of the present work was to investigate the involvement of medial hypothalamic nuclei in the organization of panic-like behaviour and the innate fear-induced oscillations of nociceptive thresholds. The blockade of GABA(A) receptors in the neuronal substrates of the ventromedial or dorsomedial hypothalamus was followed by elaborated defensive panic-like reactions. Moreover, innate fear-induced antinociception was consistently elicited after the escape behaviour. The escape responses organized by the dorsomedial and ventromedial hypothalamic nuclei were characteristically more elaborated, and a remarkable exploratory behaviour was recorded during GABA(A) receptor blockade in the medial hypothalamus. The motor characteristic of the elaborated defensive escape behaviour and the patterns of defensive alertness and defensive immobility induced by microinjection of the bicuculline either into the dorsomedial or into the ventromedial hypothalamus were very similar. This was followed by the same pattern of innate fear-induced antinociceptive response that lasted approximately 40 min after the elaborated defensive escape reaction in both cases. These findings suggest that dysfunction of the GABA-mediated neuronal system in the medial hypothalamus causes panic-like responses in laboratory animals, and that the elaborated escape behaviour organized in both dorsomedial and ventromedial hypothalamic nuclei are followed by significant innate-fear-induced antinociception. Our findings indicate that the GABA(A) receptor of dorsomedial and ventromedial hypothalamic nuclei are critically involved in the modulation of panic-like behaviour.

Effect of progesterone on the expression of GABA(A) receptor subunits in the prefrontal cortex of rats: implications of sex differences and brain hemisphere.

PubMed

Andrade, Susie; Arbo, Bruno D; Batista, Bruna A M; Neves, Alice M; Branchini, Gisele; Brum, Ilma S; Barros, Helena M T; Gomez, Rosane; Ribeiro, Maria Flavia M

2012-12-01

Progesterone is a neuroactive hormone with non-genomic effects on GABA(A) receptors (GABA(A)R). Changes in the expression of GABA(A)R subunits are related to depressive-like behaviors in rats. Moreover, sex differences and depressive behaviors have been associated with prefrontal brain asymmetry in rodents and humans. Thus, our objective was to investigate the effect of progesterone on the GABA(A)R α1 and γ2 subunits mRNA expression in the right and left prefrontal cortex of diestrus female and male rats exposed to the forced swimming test (FST). Male and female rats (n = 8/group) were randomly selected to receive a daily dose of progesterone (0·4 mg·kg⁻¹) or vehicle, during two complete female estrous cycles (8-10 days). On the experiment day, male rats or diestrus female rats were euthanized 30 min after the FST. Our results showed that progesterone significantly increased the α1 subunit mRNA in both hemispheres of male and female rats. Moreover, there was an inverse correlation between depressive-like behaviors and GABA(A)R α1 subunit mRNA expression in the right hemisphere in female rats. Progesterone decreased the GABA(A)R γ2 mRNA expression only in the left hemisphere of male rats. Therefore, we conclude that the GABA(A) system displays an asymmetric distribution according to sex and that progesterone, at lower doses, presents an antidepressant effect after increasing the GABA(A) R α1 subunit expression in the right prefrontal cortex of female rats. Copyright © 2012 John Wiley & Sons, Ltd.

GABAA receptor-mediated currents in interneurons and pyramidal cells of rat visual cortex

PubMed Central

Xiang, Zixiu; Huguenard, John R; Prince, David A

1998-01-01

We compared γ-aminobutyric acid (GABA)-mediated responses of identified pyramidal cells and fast spiking interneurons in layer V of visual cortical slices from young rats (P11-14). The frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was similar in pyramidal cells and interneurons (1.7 vs. 1.9 Hz). For events with 10-90 % rise times less than 0.9 ms, no significant differences were found in mean amplitude (61 vs. 65 pA), mean rise time (0.58 vs. 0.61 ms), or the first time constant of decay (τ1, 6.4 vs. 6.5 ms) between pyramidal cells and interneurons. The second decay time constant (τ2) was significantly longer in interneurons than in pyramidal cells (49 vs. 22 ms). The difference in sIPSC decay kinetics between two cell types also existed in adult rats (P36-42), suggesting the kinetic difference is not due to differential development of GABAA receptors in these cell types. The decay kinetics of monosynaptic evoked IPSCs were also longer in interneurons. As in the case of sIPSCs, the difference was accounted for by the second decay time constant. τ1 and τ2 were, respectively, 13 and 64 ms for interneurons and 12 and 47 ms for pyramidal cells. Cell-attached patch recordings revealed that the mean open time for single Cl− channels in response to 2 μM GABA was significantly longer in interneurons than pyramidal cells (5.0 vs. 2.8 ms). The chord conductance of these channels in interneurons (12 pS) was significantly smaller than in pyramidal cells (15 pS). Single channel currents reversed polarity when the pipette potential was approximately -10 mV for both cell types. These results show that there is a functional diversity of GABAA receptors in electrophysiologically and morphologically identified cortical pyramidal cells and interneurons. This diversity might derive from the different molecular composition of the receptors in these two cell types. PMID:9503333

Cancer immunotherapy: activating innate and adaptive immunity through CD40 agonists

PubMed Central

Beatty, Gregory L.; Li, Yan; Long, Kristen B.

2017-01-01

INTRODUCTION CD40 is a promising therapeutic target for cancer immunotherapy. In patients with advanced solid malignancies, CD40 agonists have demonstrated some anti-tumor activity and a manageable toxicity profile. A 2nd generation of CD40 agonists has now been designed with optimized Fc receptor (FcR) binding based on preclinical evidence suggesting a critical role for FcR engagement in defining the potency of CD40 agonists in vivo. AREAS COVERED We provide a comprehensive review using PubMed and Google Patent databases on the current clinical status of CD40 agonists, strategies for applying CD40 agonists in cancer therapy, and the preclinical data that supports and is guiding the future development of CD40 agonists. EXPERT COMMENTARY There is a wealth of preclinical data that provide rationale on several distinct approaches for using CD40 agonists in cancer immunotherapy. This data illustrates the need to strategically combine CD40 agonists with other clinically active treatment regimens in order to realize the full potential of activating CD40 in vivo. Thus, critical to the success of this class of immune-oncology drugs, which have the potential to restore both innate and adaptive immunosurveillance, will be the identification of biomarkers for monitoring and predicting responses as well as informing mechanisms of treatment resistance. PMID:27927088

Identification of M-CSF agonists and antagonists

DOEpatents

Pandit, Jayvardhan [Mystic, CT; Jancarik, Jarmila [Walnut Creek, CA; Kim, Sung-Hou [Moraga, CA; Koths, Kirston [El Cerrito, CA; Halenbeck, Robert [San Rafael, CA; Fear, Anna Lisa [Oakland, CA; Taylor, Eric [Oakland, CA; Yamamoto, Ralph [Martinez, CA; Bohm, Andrew [Armonk, NY

2000-02-15

The present invention is directed to methods for crystallizing macrophage colony stimulating factor. The present invention is also directed to methods for designing and producing M-CSF agonists and antagonists using information derived from the crystallographic structure of M-CSF. The invention is also directed to methods for screening M-CSF agonists and antagonists. In addition, the present invention is directed to an isolated, purified, soluble and functional M-CSF receptor.

The novel antiepileptic drug imepitoin compares favourably to other GABA-mimetic drugs in a seizure threshold model in mice and dogs.

PubMed

Löscher, Wolfgang; Hoffmann, Katrin; Twele, Friederike; Potschka, Heidrun; Töllner, Kathrin

2013-11-01

Recently, the imidazolinone derivative imepitoin has been approved for treatment of canine epilepsy. Imepitoin acts as a low-affinity partial agonist at the benzodiazepine (BZD) site of the GABAA receptor and is the first compound with such mechanism that has been developed as an antiepileptic drug (AED). This mechanism offers several advantages compared to full agonists, including less severe adverse effects and a lack of tolerance and dependence liability, which has been demonstrated in rodents, dogs, and nonhuman primates. In clinical trials in epileptic dogs, imepitoin was shown to be an effective and safe AED. Recently, seizures in dogs have been proposed as a translational platform for human therapeutic trials on new epilepsy treatments. In the present study, we compared the anticonvulsant efficacy of imepitoin, phenobarbital and the high-affinity partial BZD agonist abecarnil in the timed i.v. pentylenetetrazole (PTZ) seizure threshold test in dogs and, for comparison, in mice. Furthermore, adverse effects of treatments were compared in both species. All drugs dose-dependently increased the PTZ threshold in both species, but anticonvulsant efficacy was higher in dogs than mice. At the doses selected for this study, imepitoin was slightly less potent than phenobarbital in increasing seizure threshold, but markedly more tolerable in both species. Effective doses of imepitoin in the PTZ seizure model were in the same range as those suppressing spontaneous recurrent seizures in epileptic dogs. The study demonstrates that low-affinity partial agonists at the benzodiazepine site of the GABAA receptor, such as imepitoin, offer advantages as a new category of AEDs. Copyright © 2013 Elsevier Ltd. All rights reserved.

Macroscopic kinetics of pentameric ligand gated ion channels: comparisons between two prokaryotic channels and one eukaryotic channel.

PubMed

Laha, Kurt T; Ghosh, Borna; Czajkowski, Cynthia

2013-01-01

Electrochemical signaling in the brain depends on pentameric ligand-gated ion channels (pLGICs). Recently, crystal structures of prokaryotic pLGIC homologues from Erwinia chrysanthemi (ELIC) and Gloeobacter violaceus (GLIC) in presumed closed and open channel states have been solved, which provide insight into the structural mechanisms underlying channel activation. Although structural studies involving both ELIC and GLIC have become numerous, thorough functional characterizations of these channels are still needed to establish a reliable foundation for comparing kinetic properties. Here, we examined the kinetics of ELIC and GLIC current activation, desensitization, and deactivation and compared them to the GABAA receptor, a prototypic eukaryotic pLGIC. Outside-out patch-clamp recordings were performed with HEK-293T cells expressing ELIC, GLIC, or α1β2γ2L GABAA receptors, and ultra-fast ligand application was used. In response to saturating agonist concentrations, we found both ELIC and GLIC current activation were two to three orders of magnitude slower than GABAA receptor current activation. The prokaryotic channels also had slower current desensitization on a timescale of seconds. ELIC and GLIC current deactivation following 25 s pulses of agonist (cysteamine and pH 4.0 buffer, respectively) were relatively fast with time constants of 24.9 ± 5.1 ms and 1.2 ± 0.2 ms, respectively. Surprisingly, ELIC currents evoked by GABA activated very slowly with a time constant of 1.3 ± 0.3 s and deactivated even slower with a time constant of 4.6 ± 1.2 s. We conclude that the prokaryotic pLGICs undergo similar agonist-mediated gating transitions to open and desensitized states as eukaryotic pLGICs, supporting their use as experimental models. Their uncharacteristic slow activation, slow desensitization and rapid deactivation time courses are likely due to differences in specific structural elements, whose future identification may help uncover mechanisms underlying p

Synthetic RORγt Agonists Enhance Protective Immunity

PubMed Central

Chang, Mi Ra; Dharmarajan, Venkatasubramanian; Doebelin, Christelle; Garcia-Ordonez, Ruben D.; Novick, Scott J.; Kuruvilla, Dana S.; Kamenecka, Theodore M.; Griffin, Patrick R.

2016-01-01

The T cell specific RORγ isoform RORγt has been shown to be the key lineage-defining transcription factor to initiate the differentiation program of TH17 and Tc17 cells, cells that have demonstrated anti-tumor efficacy. RORγt controls gene networks that enhance immunity including increased IL17 production and decreased immune suppression. Both synthetic and putative endogenous agonists of RORγt have been shown to increase the basal activity of RORγt enhancing TH17 cell proliferation. Here we show that activation of RORγt using synthetic agonists drives proliferation of TH17 cells while decreasing levels of the immune checkpoint protein PD-1, a mechanism that should enhance anti-tumor immunity while blunting tumor associated adaptive immune resistance. Interestingly, putative endogenous agonists drive proliferation of TH17 cells but do not repress PD-1. These findings suggest that synthetic agonists of RORγt should activate TC17/TH17 cells (with concomitant reduction in the Tregs population), repress PD-1, and produce IL17 in situ (a factor associated with good prognosis in cancer). Enhanced immunity and blockage of immune checkpoints has transformed cancer treatment, thus such a molecule would provide a unique approach for the treatment of cancer. PMID:26785144

Magnesium ions and opioid agonists in vincristine-induced neuropathy.

PubMed

Bujalska, Magdalena; Makulska-Nowak, Helena; Gumułka, Stanisław W

2009-01-01

Neuropathic pain is difficult to treat. Classic analgesics (i.e., opioid receptor agonists) usually possess low activity. Therefore other agents such as antidepressants, anticonvulsants, and corticosteroids are used. It is commonly known that NMDA antagonists increase analgesic activity of opioids. Unfortunately, clinical use of NMDA antagonists is limited because of the relatively frequent occurrence of adverse effects e.g., memory impairment, psychomimetic effects, ataxia and motor in-coordination. Magnesium ions (Mg(2+)) are NMDA receptor blockers in physiological conditions. Therefore, in this study the effect of opioid receptor agonists and the influence of Mg(2+) on the action of opioid agonists in vincristine-induced hyperalgesia were examined. Opioid agonists such as morphine (5 mg/kg, ip), and fentanyl (0.0625 mg/kg, ip), as well as the partial agonist buprenorphine (0.075 mg/kg, ip) administered alone on 5 consecutives days did not modify the hyperalgesia in vincristine rats. In contrast, pretreatment with a low dose of magnesium sulfate (30 mg/kg, ip) resulted in a progressive increase of the analgesic action of all three investigated opioids. After discontinuation of drug administration, the effect persisted for several days.

Effects of AMPA receptor antagonist, NBQX, and extrasynaptic GABAA agonist, THIP, on social behavior of adolescent and adult rats.

PubMed

Dannenhoffer, Carol A; Varlinskaya, Elena I; Spear, Linda Patia

2018-05-22

Adolescence is characterized by high significance of social interactions, along with a propensity to exhibit social facilitating effects of ethanol while being less sensitive than adults to the inhibition of social behavior that emerges at higher doses of ethanol. Among the neural characteristics of adolescence are generally enhanced levels of glutamatergic (especially NMDA receptor) activity relative to adults, whereas the GABA system is still developmentally immature. Activation of NMDA receptors likely plays a role in modulation of social behavior in adolescent animals as well as in socially facilitating and suppressing effects of ethanol. For instance, adolescent and adult rats differ in their sensitivities to the effects of NMDA antagonists and ethanol on social behavior, with adolescents but not adults demonstrating social facilitation at lower doses of both drugs and adults being more sensitive to the socially suppressing effects evident at higher doses of each. The roles of AMPA and extrasynaptic GABA A receptors in modulation of social behavior during adolescence and in adulthood are still unknown. The present study was designed to assess whether pharmacological blockade of AMPA receptors and/or activation of extrasynaptic GABA A receptors results in age-dependent alterations of social behavior. Adolescent and adult male and female Sprague-Dawley rats were injected with an assigned dose of either a selective AMPA antagonist, NBQX (Experiment 1) or extrasynaptic GABA A agonist, THIP (Experiment 2) and placed into a modified social interaction chamber for a 30-min habituation period prior to a 10-min social interaction test with a novel age- and sex-matched partner. Behaviors such as social investigation, contact behavior and play behavior were scored from video recordings of the interaction tests. In Experiment 1, NBQX produced similar social inhibition at higher doses in both age groups. In Experiment 2, THIP induced inhibition in adolescents, but not

TLR9 agonist protects mice from radiation-induced gastrointestinal syndrome.

PubMed

Saha, Subhrajit; Bhanja, Payel; Liu, Laibin; Alfieri, Alan A; Yu, Dong; Kandimalla, Ekambar R; Agrawal, Sudhir; Guha, Chandan

2012-01-01

Radiation-induced gastrointestinal syndrome (RIGS) is due to the clonogenic loss of crypt cells and villi depopulation, resulting in disruption of mucosal barrier, bacterial invasion, inflammation and sepsis. Intestinal macrophages could recognize invading bacterial DNA via TLR9 receptors and transmit regenerative signals to the neighboring crypt. We therefore investigated whether systemic administration of designer TLR9 agonist could ameliorate RIGS by activating TLR9. Male C57Bl6 mice were distributed in four experimental cohorts, whole body irradiation (WBI) (8.4-10.4 Gy), TLR9 agonist (1 mg/kg s.c.), 1 h pre- or post-WBI and TLR9 agonist+WBI+iMyd88 (pretreatment with inhibitory peptide against Myd88). Animals were observed for survival and intestine was harvested for histological analysis. BALB/c mice with CT26 colon tumors in abdominal wall were irradiated with 14 Gy single dose of whole abdominal irradiation (AIR) for tumor growth study. Mice receiving pre-WBI TLR9 agonist demonstrated improvement of survival after 10.4 Gy (p<0.03), 9.4 Gy (p<0.008) and 8.4 Gy (p<0.002) of WBI, compared to untreated or iMyd88-treated controls. Post-WBI TLR9 agonist mitigates up to 8.4 Gy WBI (p<0.01). Histological analysis and xylose absorption test demonstrated significant structural and functional restitution of the intestine in WBI+TLR9 agonist cohorts. Although, AIR reduced tumor growth, all animals died within 12 days from RIGS. TLR9 agonist improved the survival of mice beyond 28 days post-AIR (p<0.008) with significant reduction of tumor growth (p<0.0001). TLR9 agonist treatment could serve both as a prophylactic or mitigating agent against acute radiation syndrome and also as an adjuvant therapy to increase the therapeutic ratio of abdominal Radiation Therapy for Gastro Intestinal malignancies.

The effect of propofol on CA1 pyramidal cell excitability and GABAA-mediated inhibition in the rat hippocampal slice.

PubMed

Albertson, T E; Walby, W F; Stark, L G; Joy, R M

1996-05-24

An in vitro paired-pulse orthodromic stimulation technique was used to examine the effects of propofol on excitatory afferent terminals, CA1 pyramidal cells and recurrent collateral evoked inhibition in the rat hippocampal slice. Hippocampal slices 400 microns thick were perfused with oxygenated artificial cerebrospinal fluid, and electrodes were placed in the CA1 region to record extracellular field population spike (PS) or excitatory postsynaptic potential (EPSP) responses to stimulation of Schaffer collateral/commissural fibers. Gamma-aminobutyric acid (GABA)-mediated recurrent inhibition was measured using a paired-pulse technique. The major effect of propofol (7-28 microM) was a dose and time dependent increase in the intensity and duration of GABA-mediated inhibition. This propofol effect could be rapidly and completely reversed by exposure to known GABAA antagonists, including picrotoxin, bicuculline and pentylenetetrazol. It was also reversed by the chloride channel antagonist, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). It was not antagonized by central (flumazenil) or peripheral (PK11195) benzodiazepine antagonists. Reversal of endogenous inhibition was also noted with the antagonists picrotoxin and pentylenetetrazol. Input/output curves constructed using stimulus propofol caused only a small enhancement of EPSPs at higher stimulus intensities but had no effect on PS amplitudes. These studies are consistent with propofol having a GABAA-chloride channel mechanism causing its effect on recurrent collateral evoked inhibition in the rat hippocampal slice.

The Homeostatic Interaction Between Anodal Transcranial Direct Current Stimulation and Motor Learning in Humans is Related to GABAA Activity.

PubMed

Amadi, Ugwechi; Allman, Claire; Johansen-Berg, Heidi; Stagg, Charlotte J

2015-01-01

The relative timing of plasticity-induction protocols is known to be crucial. For example, anodal transcranial direct current stimulation (tDCS), which increases cortical excitability and typically enhances plasticity, can impair performance if it is applied before a motor learning task. Such timing-dependent effects have been ascribed to homeostatic plasticity, but the specific synaptic site of this interaction remains unknown. We wished to investigate the synaptic substrate, and in particular the role of inhibitory signaling, underpinning the behavioral effects of anodal tDCS in homeostatic interactions between anodal tDCS and motor learning. We used transcranial magnetic stimulation (TMS) to investigate cortical excitability and inhibitory signaling following tDCS and motor learning. Each subject participated in four experimental sessions and data were analyzed using repeated measures ANOVAs and post-hoc t-tests as appropriate. As predicted, we found that anodal tDCS prior to the motor task decreased learning rates. This worsening of learning after tDCS was accompanied by a correlated increase in GABAA activity, as measured by TMS-assessed short interval intra-cortical inhibition (SICI). This provides the first direct demonstration in humans that inhibitory synapses are the likely site for the interaction between anodal tDCS and motor learning, and further, that homeostatic plasticity at GABAA synapses has behavioral relevance in humans. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Convulsions induced by centrally administered NMDA in mice: effects of NMDA antagonists, benzodiazepines, minor tranquilizers and anticonvulsants.

PubMed Central

Moreau, J. L.; Pieri, L.; Prud'hon, B.

1989-01-01

1. Convulsions were induced reproducibly by intracerebroventricular injection of N-methyl-D-aspartic acid (NMDA) to conscious mice. 2. Competitive (carboxypiperazine-propylphosphonic acid, CPP; 2-amino-7-phosphonoheptanoic acid, AP7) and non-competitive (MK801; phencyclidine, PCP; thienylcyclohexylpiperidine, TCP; dextrorphan; dextromethorphan) NMDA antagonists prevented NMDA-induced convulsions. 3. Benzodiazepine receptor agonists and partial agonists (triazolam, diazepam, clonazepam, Ro 16-6028), classical anticonvulsants (diphenylhydantoin, phenobarbitone, sodium valproate) and meprobamate were also found to prevent NMDA-induced convulsions. 4. Flumazenil (a benzodiazepine receptor antagonist) and the GABA agonists THIP and muscimol (up to subtoxic doses) were without effect. 5. Flumazenil reversed the anticonvulsant action of diazepam, but not that of MK801. 6. Results obtained in this model differ somewhat from those described in a seizure model with systemic administration of NMDA. An explanation for this discrepancy is offered. 7. This model is a simple test for assessing the in vivo activity of NMDA antagonists and also expands the battery of chemically-induced seizure models for characterizing anticonvulsants not acting at NMDA receptors. PMID:2574061

Anxiolytic-Like Effects and Increase in Locomotor Activity Induced by Infusions of NMDA into the Ventral Hippocampus in Rat: Interaction with GABAergic System.

PubMed

Bina, Payvand; Rezvanfard, Mehrnaz; Ahmadi, Shamseddin; Zarrindast, Mohammad Reza

2014-10-01

In this study, we investigated the role of N-Methyl-D-Aspartate (NMDA) receptors in the ventral hippocampus (VH) and their possible interactions with GABAA system on anxiety-like behaviors. We used an elevated-plus maze test (EPM) to assess anxiety-like behaviors and locomotor activity in male Wistar rats. The results showed that intra-VH infusions of different doses of NMDA (0.25 and 0.5 μg/rat) increased locomotor activity, and also induced anxiolytic-like behaviors, as revealed by a tendency to increase percentage of open arm time (%OAT), and a significant increase in percentage of open arm entries (%OAE). The results also showed that intra-VH infusions of muscimol (0.5 and 1 μg/rat) or bicuculline (0.5 and 1 μg/rat) did not significantly affect anxiety-like behaviors, but bicuculline at dose of 1 μg/rat increased locomotor activity. Intra-VH co-infusions of muscimol (0.5 μg/rat) along with low doses of NMDA (0.0625 and 0.125 μg/rat) showed a tendency to increase %OAT, %OAE and locomotor activity; however, no interaction was observed between the drugs. Interestingly, intra-VH co-infusions of bicuculline (0.5 μg/rat) along with effective doses of NMDA (0.25 and 0.5 μg/rat) decreased %OAT, %OAE and locomotor activity, and a significant interaction between two drugs was observed. It can be concluded that GABAergic system may mediate the anxiolytic-like effects and increase in locomotor activity induced by NMDA in the VH.

GABAA receptor γ2 subunit knockdown mice have enhanced anxiety-like behavior but unaltered hypnotic response to benzodiazepines

PubMed Central

Chandra, Dev; Korpi, Esa R; Miralles, Celia P; De Blas, Angel L; Homanics, Gregg E

2005-01-01

Background Gamma-aminobutyric acid type A receptors (GABAA-Rs) are the major inhibitory receptors in the mammalian brain and are modulated by a number of sedative/hypnotic drugs including benzodiazepines and anesthetics. The significance of specific GABAA-Rs subunits with respect to behavior and in vivo drug responses is incompletely understood. The γ2 subunit is highly expressed throughout the brain. Global γ2 knockout mice are insensitive to the hypnotic effects of diazepam and die perinatally. Heterozygous γ2 global knockout mice are viable and have increased anxiety-like behaviors. To further investigate the role of the γ2 subunit in behavior and whole animal drug action, we used gene targeting to create a novel mouse line with attenuated γ2 expression, i.e., γ2 knockdown mice. Results Knockdown mice were created by inserting a neomycin resistance cassette into intron 8 of the γ2 gene. Knockdown mice, on average, showed a 65% reduction of γ2 subunit mRNA compared to controls; however γ2 gene expression was highly variable in these mice, ranging from 10–95% of normal. Immunohistochemical studies demonstrated that γ2 protein levels were also variably reduced. Pharmacological studies using autoradiography on frozen brain sections demonstrated that binding of the benzodiazepine site ligand Ro15-4513 was decreased in mutant mice compared to controls. Behaviorally, knockdown mice displayed enhanced anxiety-like behaviors on the elevated plus maze and forced novelty exploration tests. Surprisingly, mutant mice had an unaltered response to hypnotic doses of the benzodiazepine site ligands diazepam, midazolam and zolpidem as well as ethanol and pentobarbital. Lastly, we demonstrated that the γ2 knockdown mouse line can be used to create γ2 global knockout mice by crossing to a general deleter cre-expressing mouse line. Conclusion We conclude that: 1) insertion of a neomycin resistance gene into intron 8 of the γ2 gene variably reduced the amount of γ2

Evidence that GABA ρ subunits contribute to functional ionotropic GABA receptors in mouse cerebellar Purkinje cells

PubMed Central

Harvey, Victoria L; Duguid, Ian C; Krasel, Cornelius; Stephens, Gary J

2006-01-01

Ionotropic γ-amino butyric acid (GABA) receptors composed of heterogeneous molecular subunits are major mediators of inhibitory responses in the adult CNS. Here, we describe a novel ionotropic GABA receptor in mouse cerebellar Purkinje cells (PCs) using agents reported to have increased affinity for ρ subunit-containing GABAC over other GABA receptors. Exogenous application of the GABAC-preferring agonist cis-4-aminocrotonic acid (CACA) evoked whole-cell currents in PCs, whilst equimolar concentrations of GABA evoked larger currents. CACA-evoked currents had a greater sensitivity to the selective GABAC antagonist (1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid (TPMPA) than GABA-evoked currents. Focal application of agonists produced a differential response profile; CACA-evoked currents displayed a much more pronounced attenuation with increasing distance from the PC soma, displayed a slower time-to-peak and exhibited less desensitization than GABA-evoked currents. However, CACA-evoked currents were also completely blocked by bicuculline, a selective agent for GABAA receptors. Thus, we describe a population of ionotropic GABA receptors with a mixed GABAA/GABAC pharmacology. TPMPA reduced inhibitory synaptic transmission at interneurone–Purkinje cell (IN–PC) synapses, causing clear reductions in miniature inhibitory postsynaptic current (mIPSC) amplitude and frequency. Combined application of NO-711 (a selective GABA transporter subtype 1 (GAT-1) antagonist) and SNAP-5114 (a GAT-(2)/3/4 antagonist) induced a tonic GABA conductance in PCs; however, TPMPA had no effect on this current. Immunohistochemical studies suggest that ρ subunits are expressed predominantly in PC soma and proximal dendritic compartments with a lower level of expression in more distal dendrites; this selective immunoreactivity contrasted with a more uniform distribution of GABAA α1 subunits in PCs. Finally, co-immunoprecipitation studies suggest that ρ subunits can form complexes

TOXICITY OF AHR AGONISTS TO FISH EARLY LIFE STAGES

EPA Science Inventory

Fish early life stages are exceptionally sensitive to the lethal toxicity of chemicals that act as arylhydrocarbon receptor (AhR) agonists. Toxicity characterizations based on 2,3,7,8-tetrachlorodibenzo-p-dioxin, generally the most potent AhR agonist, support the toxicity equiva...

Electrophysiological Perspectives on the Therapeutic Use of Nicotinic Acetylcholine Receptor Partial AgonistsS⃞

PubMed Central

Trocmé-Thibierge, Caryn; Guendisch, Daniela; Al Rubaiy, Shehd Abdullah Abbas; Bloom, Stephen A.

2011-01-01

Partial agonist therapies rely variously on two hypotheses: the partial agonists have their effects through chronic low-level receptor activation or the partial agonists work by decreasing the effects of endogenous or exogenous full agonists. The relative significance of these activities probably depends on whether acute or chronic effects are considered. We studied nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus laevis oocytes to test a model for the acute interactions between acetylcholine (ACh) and weak partial agonists. Data were best-fit to a basic competition model that included an additional factor for noncompetitive inhibition. Partial agonist effects were compared with the nAChR antagonist bupropion in prolonged bath application experiments that were designed to mimic prolonged drug exposure typical of therapeutic drug delivery. A primary effect of prolonged application of nicotine was to decrease the response of all nAChR subtypes to acute applications of ACh. In addition, nicotine, cytisine, and varenicline produced detectable steady-state activation of α4β2* [(α4)2(β2)3, (α4)3(β2)2, and (α4)2(β2)2α5)] receptor subtypes that was not seen with other test compounds. Partial agonists produced no detectable steady-state activation of α7 nAChR, but seemed to show small potentiation of ACh-evoked responses; however, “run-up” of α7 ACh responses was also sometimes observed under control conditions. Potential off-target effects of the partial agonists therefore included the modulation of α7 responses by α4β2 partial agonists and decreases in α4β2* responses by α7-selective agonists. These data indicate the dual effects expected for α4β2* partial agonists and provide models and insights for utility of partial agonists in therapeutic development. PMID:21285282

Agonist-Specific Recruitment of Arrestin Isoforms Differentially Modify Delta Opioid Receptor Function

PubMed Central

Perroy, Julie; Walwyn, Wendy M.; Smith, Monique L.; Vicente-Sanchez, Ana; Segura, Laura; Bana, Alia; Kieffer, Brigitte L.; Evans, Christopher J.

2016-01-01

Ligand-specific recruitment of arrestins facilitates functional selectivity of G-protein-coupled receptor signaling. Here, we describe agonist-selective recruitment of different arrestin isoforms to the delta opioid receptor in mice. A high-internalizing delta opioid receptor agonist (SNC80) preferentially recruited arrestin 2 and, in arrestin 2 knock-outs (KOs), we observed a significant increase in the potency of SNC80 to inhibit mechanical hyperalgesia and decreased acute tolerance. In contrast, the low-internalizing delta agonists (ARM390, JNJ20788560) preferentially recruited arrestin 3 with unaltered behavioral effects in arrestin 2 KOs. Surprisingly, arrestin 3 KO revealed an acute tolerance to these low-internalizing agonists, an effect never observed in wild-type animals. Furthermore, we examined delta opioid receptor–Ca2+ channel coupling in dorsal root ganglia desensitized by ARM390 and the rate of resensitization was correspondingly decreased in arrestin 3 KOs. Live-cell imaging in HEK293 cells revealed that delta opioid receptors are in pre-engaged complexes with arrestin 3 at the cell membrane and that ARM390 strengthens this membrane interaction. The disruption of these complexes in arrestin 3 KOs likely accounts for the altered responses to low-internalizing agonists. Together, our results show agonist-selective recruitment of arrestin isoforms and reveal a novel endogenous role of arrestin 3 as a facilitator of resensitization and an inhibitor of tolerance mechanisms. SIGNIFICANCE STATEMENT Agonists that bind to the same receptor can produce highly distinct signaling events and arrestins are a major mediator of this ligand bias. Here, we demonstrate that delta opioid receptor agonists differentially recruit arrestin isoforms. We found that the high-internalizing agonist SNC80 preferentially recruits arrestin 2 and knock-out (KO) of this protein results in increased efficacy of SNC80. In contrast, low-internalizing agonists (ARM390 and JNJ20788560

Differential effects of AMPK agonists on cell growth and metabolism

PubMed Central

Vincent, Emma E.; Coelho, Paula P.; Blagih, Julianna; Griss, Takla; Viollet, Benoit; Jones, Russell G.

2016-01-01

As a sensor of cellular energy status, the AMP-activated protein kinase (AMPK) is believed to act in opposition to the metabolic phenotypes favored by proliferating tumor cells. Consequently, compounds known to activate AMPK have been proposed as cancer therapeutics. However, the extent to which the anti-neoplastic properties of these agonists are mediated by AMPK is unclear. Here we examined the AMPK-dependence of six commonly used AMPK agonists (metformin, phenformin, AICAR, 2DG, salicylate and A-769662) and their influence on cellular processes often deregulated in tumor cells. We demonstrate that the majority of these agonists display AMPK-independent effects on cell proliferation and metabolism with only the synthetic activator, A-769662, exerting AMPK-dependent effects on these processes. We find that A-769662 promotes an AMPK-dependent increase in mitochondrial spare respiratory capacity (SRC). Finally, contrary to the view of AMPK activity being tumor suppressive, we find A-769662 confers a selective proliferative advantage to tumor cells growing under nutrient deprivation. Our results indicate that many of the anti-growth properties of these agonists cannot be attributed to AMPK activity in cells, and thus any observed effects using these agonists should be confirmed using AMPK-deficient cells. Ultimately, our data urge caution, not only regarding the type of AMPK agonist proposed for cancer treatment, but also the context in which they are used. PMID:25241895

Regulation of GABAA receptors by fragile X mental retardation protein

PubMed Central

Liu, Baosong; Li, Lijun; Chen, Juan; Wang, Zefen; Li, Zhiqiang; Wan, Qi

2013-01-01

Fragile X syndrome (FXS) is caused by the loss of fragile X mental retardation protein (FMRP). The deficiency of GABAA receptors (GABAARs) is implicated in FXS. However, the underlying mechanisms remain unclear. To investigate the effect of FMRP on GABAARs, we transfected FMRP cDNAs in rat cortical neurons. We measured the protein expression of GABAARs and phosphatase PTEN, and recorded GABAAR-mediated whole-cell currents in the transfected neurons. We show that the transfection of FMRP cDNAs causes increased protein expression of GABAARs in cortical neurons, but GABAAR-mediated whole-cell currents are not potentiated by FMRP transfection. These results suggest the possibility that intracellular signaling antagonizing GABAAR activity may play a role in inhibiting GABAAR function in FMRP-transfected neurons. We further show that FMRP transfection results in an enhanced protein expression of PTEN, which contributes to the inhibition of GABAAR function in FMRP-transfected neurons. These results indicate that GABAARs are regulated by FMRP through both an up-regulation of GABAAR expression and a PTEN enhancement-induced inhibition of GABAAR function, suggesting that an abnormal regulation of GABAAR and PTEN by the loss of FMRP underlies the pathogenesis of FXS. PMID:24044036

How theories evolved concerning the mechanism of action of barbiturates.

PubMed

Löscher, Wolfgang; Rogawski, Michael A

2012-12-01

The barbiturate phenobarbital has been in use in the treatment of epilepsy for 100 years. It has long been recognized that barbiturates act by prolonging and potentiating the action of γ-aminobutyric acid (GABA) on GABA(A) receptors and at higher concentrations directly activating the receptors. A large body of data supports the concept that GABA(A) receptors are the primary central nervous system target for barbiturates, including the finding that transgenic mice with a point mutation in the β3 GABA(A) -receptor subunit exhibit diminished sensitivity to the sedative and immobilizing actions of the anesthetic barbiturate pentobarbital. Although phenobarbital is only modestly less potent as a GABA(A) -receptor modulator than pentobarbital, phenobarbital is minimally sedating at effective anticonvulsant doses. Possible explanations for the reduced sedative effect of phenobarbital include more regionally restricted action; partial agonist activity; reduced propensity to directly activate GABA(A) receptors (possibly including extrasynaptic receptors containing δ subunits); and reduced activity at other ion channel targets, including voltage-gated calcium channels. In recent years, substantial progress has been made in defining the structural features of GABA(A) receptors responsible for gating and allosteric modulation by drugs. Although the precise sites of action of barbiturates have not yet been defined, the second and third transmembrane domains of the β subunit appear to be critical; binding may involve a pocket formed by β-subunit methionine 286 as well as α-subunit methionine 236. In addition to effects on GABA(A) receptors, barbiturates block AMPA/kainate receptors, and they inhibit glutamate release through an effect on P/Q-type high-voltage activated calcium channels. The combination of these various actions likely accounts for their diverse clinical activities. Despite the remarkable progress of the last century, there is still much to learn about the

Influence of age, body temperature, GABAA receptor inhibition and caffeine on the Hering-Breuer inflation reflex in unanesthetized rat pups

PubMed Central

Arnal, Ashley V.; Gore, Julie L.; Rudkin, Alison; Bartlett, Donald; Leiter, J.C.

2013-01-01

We measured the duration of apnea induced by sustained end-inspiratory lung inflation (the Hering Breuer Reflex; HBR) in unanesthetized infant rat pups aged 4 days (P4) to P20 at body temperatures of 32°C and 36°C. The expiratory prolongation elicited by the HBR lasted longer in the younger pups and lasted longer at the higher body temperature. Blockade of adenosine receptors by caffeine following injection into the cisterna magna (ICM) significantly blunted the thermal prolongation of the HBR. Blockade of gama-amino-butyric acid A (GABAA) receptors by pre-treatment with ICM bicuculline had no effect on the HBR duration at either body temperature. To test the hypothesis that developmental maturation of GABAergic inhibition of breathing was modifying the response to bicuculline, we pretreated rat pups with systemically administered bumetanide to lower the intracellular chloride concentration, and repeated the bicuculline studies. Bicuculline still did not alter the HBR at either temperature after bumetanide treatment. We administered PSB-36, a selective adenosine A1 receptor antagonist, and this drug treatment did not modify the HBR. We conclude that caffeine blunts the thermal prolongation of the HBR, probably by blocking adenosine A2a receptors. The thermally-sensitive adenosinergic prolongation of the HBR in these intact animals does not seem to depend on GABAA receptors PMID:23318703

Influence of age, body temperature, GABAA receptor inhibition and caffeine on the Hering-Breuer inflation reflex in unanesthetized rat pups.

PubMed

Arnal, Ashley V; Gore, Julie L; Rudkin, Alison; Bartlett, Donald; Leiter, J C

2013-03-01

We measured the duration of apnea induced by sustained end-inspiratory lung inflation (the Hering Breuer Reflex, HBR) in unanesthetized infant rat pups aged 4 days (P4) to P20 at body temperatures of 32°C and 36°C. The expiratory prolongation elicited by the HBR lasted longer in the younger pups and lasted longer at the higher body temperature. Blockade of adenosine receptors by caffeine following injection into the cisterna magna (ICM) significantly blunted the thermal prolongation of the HBR. Blockade of gama-amino-butyric acid A (GABAA) receptors by pre-treatment with ICM bicuculline had no effect on the HBR duration at either body temperature. To test the hypothesis that developmental maturation of GABAergic inhibition of breathing was modifying the response to bicuculline, we pretreated rat pups with systemically administered bumetanide to lower the intracellular chloride concentration, and repeated the bicuculline studies. Bicuculline still did not alter the HBR at either temperature after bumetanide treatment. We administered PSB-36, a selective adenosine A1 receptor antagonist, and this drug treatment did not modify the HBR. We conclude that caffeine blunts the thermal prolongation of the HBR, probably by blocking adenosine A2a receptors. The thermally sensitive adenosinergic prolongation of the HBR in these intact animals does not seem to depend on GABAA receptors. Copyright © 2013 Elsevier B.V. All rights reserved.

Lack of cocaine-like discriminative-stimulus effects of σ-receptor agonists in rats.

PubMed

Hiranita, Takato; Soto, Paul L; Tanda, Gianluigi; Katz, Jonathan L

2011-09-01

Previous studies demonstrated the effectiveness of selective σ-receptor (σR) agonists [1,3-di-o-tolylguanidine (DTG), PRE-084] as reinforcers in rats trained to self-administer cocaine. Similar to cocaine, these drugs increased nucleus accumbens shell dopamine levels, and effects of DTG, but not PRE-084, on dopamine seemed to be mediated by σRs. In addition, σR antagonists blocked self-administration of σR agonists, but were inactive against reinforcing and neurochemical effects of cocaine. Thus, pharmacologically distinct mechanisms likely underlie the reinforcing and neurochemical effects of σR agonists and cocaine. This study further examined the cocaine-like effects of σR agonists in rats trained to discriminate injections of cocaine from saline to assess the similarity of their subjective effects. Standard dopamine-uptake inhibitors (WIN 35,428, methylphenidate), but neither σR agonist (PRE-084, DTG), produced full cocaine-like discriminative-stimulus effects. The lack of effects of σR agonists was obtained regardless of route of administration (intraperitoneal, subcutaneous, or intravenous) or pretreatment time (5 or 30 min before sessions). The present results demonstrate differences in the discriminative-stimulus effects of cocaine and selective σR agonists, indicating that an overlap of subjective effects is not necessary for σR agonist self-administration. The previously found differences in neurochemical effects of cocaine and σR agonists may contribute to their different subjective effects.

Lack of Cocaine-Like Discriminative-Stimulus Effects of σ Receptor Agonists in Rats

PubMed Central

Hiranita, Takato; Soto, Paul L.; Tanda, Gianluigi; Katz, Jonathan L.

2013-01-01

Previous studies demonstrated effectiveness of selective sigma-receptor (σR) agonists (DTG, PRE-084) as reinforcers in rats trained to self-administer cocaine. Like cocaine, these drugs increased nucleus accumbens shell dopamine levels, and effects of DTG, but not PRE-084, on dopamine appeared to be mediated by σRs. Additionally, σR antagonists blocked self-administration of σR agonists, but were inactive against reinforcing and neurochemical effects of cocaine. Thus pharmacologically distinct mechanisms likely underlie the reinforcing and neurochemical effects of σR agonists and cocaine. The present study further examined the cocaine-like effects of σR agonists in rats trained to discriminate injections of cocaine from saline to assess the similarity of their subjective effects. Standard dopamine-uptake inhibitors (WIN 35,428, methylphenidate), but neither σR agonist (PRE-084, DTG) produced full cocaine-like discriminative-stimulus effects. The lack of effects of σR agonists was obtained regardless of route of administration (i.p., s.c. or i.v.) or pretreatment time (5- or 30-min before sessions). The present results demonstrate differences in the discriminative-stimulus effects of cocaine and selective σR agonists, indicating that an overlap of subjective effects is not necessary for σR agonist self-administration. The previously found differences in neurochemical effects of cocaine and σR agonists may contribute to their different subjective effects. PMID:21808192

Pharmacological characterization of the new histamine H4 receptor agonist VUF 8430

PubMed Central

Lim, Herman D; Adami, Maristella; Guaita, Elena; Werfel, Thomas; Smits, Rogier A; de Esch, Iwan JP; Bakker, Remko A; Gutzmer, Ralf; Coruzzi, Gabriella; Leurs, Rob

2009-01-01

Background and purpose: We compare the pharmacological profiles of a new histamine H4 receptor agonist 2-(2-guanidinoethyl)isothiourea (VUF 8430) with that of a previously described H4 receptor agonist, 4-methylhistamine. Experimental approach: Radioligand binding and functional assays were performed using histamine H4 receptors expressed in mammalian cell lines. Compounds were also evaluated ex vivo in monocyte-derived dendritic cells endogenously expressing H4 receptors and in vivo in anaesthetized rats for gastric acid secretion activity. Key results: Both VUF 8430 and 4-methylhistamine were full agonists at human H4 receptors with lower affinity at rat and mouse H4 receptors. Both compounds induced chemotaxis of monocyte-derived dendritic cells. VUF 8430 also showed reasonable affinity and was a full agonist at the H3 receptor. Agmatine is a metabolite of arginine, structurally related to VUF 8430, and was a H4 receptor agonist with micromolar affinity. At histamine H3 receptors, agmatine was a full agonist, whereas 4-methylhistamine was an agonist only at high concentrations. Both VUF 8430 and agmatine were inactive at H1 and H2 receptors, whereas 4-methylhistamine is as active as histamine at H2 receptors. In vivo, VUF 8430 only caused a weak secretion of gastric acid mediated by H2 receptors, whereas 4-methylhistamine, dimaprit, histamine and amthamine, at equimolar doses, induced 2.5- to 6-fold higher output than VUF 8430. Conclusions and implications: Our results suggest complementary use of 4-methylhistamine and VUF 8430 as H4 receptor agonists. Along with H4 receptor antagonists, both agonists can serve as useful pharmacological tools in studies of histamine H4 receptors. PMID:19413569

Kavalactones and dihydrokavain modulate GABAergic activity in a rat gastric-brainstem preparation.

PubMed

Yuan, Chun-Su; Dey, Lucy; Wang, Anbao; Mehendale, Sangeeta; Xie, Jing-Tian; Aung, Han H; Ang-Lee, Michael K

2002-12-01

Using an in vitro neonatal rat gastric-brainstem preparation, the activity of majority neurons recorded in the nucleus tractus solitarius (NTS) of the brainstem were significantly inhibited by GABA A receptor agonist, muscimol (30 microM), and this inhibition was reversed by selective GABA A receptor antagonist, bicuculline (10 microM). Application of kavalactones (300 microg/ml) and dihydrokavain (300 microM) into the brainstem compartment of the preparation also significantly reduced the discharge rate of these NTS neurons (39 % and 32 %, respectively, compared to the control level), and this reduction was partially reversed by bicuculline (10 microM). Kavalactones or dihydrokavain induced inhibitory effects were not reduced after co-application of saclofen (10 microM; a selective GABA B receptor antagonist) or naloxone (100 nM; an opioid receptor antagonist). Pretreatment with kavalactones (300 microg/ml) or dihydrokavain (300 microM) significantly decreased the NTS inhibitory effects induced by muscimol (30 microM), approximately from 51 % to 36 %. Our results demonstrated modulation of brainstem GABAergic mechanism by kavalactones and dihydrokavain, and suggested that these compounds may play an important role in regulation of GABAergic neurotransmission.

Differential effects of AMPK agonists on cell growth and metabolism.

PubMed

Vincent, E E; Coelho, P P; Blagih, J; Griss, T; Viollet, B; Jones, R G

2015-07-01

As a sensor of cellular energy status, the AMP-activated protein kinase (AMPK) is believed to act in opposition to the metabolic phenotypes favored by proliferating tumor cells. Consequently, compounds known to activate AMPK have been proposed as cancer therapeutics. However, the extent to which the anti-neoplastic properties of these agonists are mediated by AMPK is unclear. Here we examined the AMPK dependence of six commonly used AMPK agonists (metformin, phenformin, 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), 2-deoxy-D-glucose (2DG), salicylate and A-769662) and their influence on cellular processes often deregulated in tumor cells. We demonstrate that the majority of these agonists display AMPK-independent effects on cell proliferation and metabolism with only the synthetic activator, A-769662, exerting AMPK-dependent effects on these processes. We find that A-769662 promotes an AMPK-dependent increase in mitochondrial spare respiratory capacity. Finally, contrary to the view of AMPK activity being tumor suppressive, we find that A-769662 confers a selective proliferative advantage to tumor cells growing under nutrient deprivation. Our results indicate that many of the antigrowth properties of these agonists cannot be attributed to AMPK activity in cells, and thus any observed effects using these agonists should be confirmed using AMPK-deficient cells. Ultimately, our data urge caution not only regarding the type of AMPK agonist proposed for cancer treatment but also the context in which they are used.

Sex-Dependent Anti-Stress Effect of an α5 Subunit Containing GABAA Receptor Positive Allosteric Modulator

PubMed Central

Piantadosi, Sean C.; French, Beverly J.; Poe, Michael M.; Timić, Tamara; Marković, Bojan D.; Pabba, Mohan; Seney, Marianne L.; Oh, Hyunjung; Orser, Beverley A.; Savić, Miroslav M.; Cook, James M.; Sibille, Etienne

2016-01-01

Rationale: Current first-line treatments for stress-related disorders such as major depressive disorder (MDD) act on monoaminergic systems and take weeks to achieve a therapeutic effect with poor response and low remission rates. Recent research has implicated the GABAergic system in the pathophysiology of depression, including deficits in interneurons targeting the dendritic compartment of cortical pyramidal cells. Objectives: The present study evaluates whether SH-053-2’F-R-CH3 (denoted “α5-PAM”), a positive allosteric modulator selective for α5-subunit containing GABAA receptors found predominantly on cortical pyramidal cell dendrites, has anti-stress effects. Methods: Female and male C57BL6/J mice were exposed to unpredictable chronic mild stress (UCMS) and treated with α5-PAM acutely (30 min prior to assessing behavior) or chronically before being assessed behaviorally. Results: Acute and chronic α5-PAM treatments produce a pattern of decreased stress-induced behaviors (denoted as “behavioral emotionality”) across various tests in female, but not in male mice. Behavioral Z-scores calculated across a panel of tests designed to best model the range and heterogeneity of human symptomatology confirmed that acute and chronic α5-PAM treatments consistently produce significant decreases in behavioral emotionality in several independent cohorts of females. The behavioral responses to α5-PAM could not be completely accounted for by differences in drug brain disposition between female and male mice. In mice exposed to UCMS, expression of the Gabra5 gene was increased in the frontal cortex after acute treatment and in the hippocampus after chronic treatment with α5-PAM in females only, and these expression changes correlated with behavioral emotionality. Conclusion: We showed that acute and chronic positive modulation of α5 subunit-containing GABAA receptors elicit anti-stress effects in a sex-dependent manner, suggesting novel therapeutic modalities

Differential agonist sensitivity of glycine receptor α2 subunit splice variants

PubMed Central

Miller, Paul S; Harvey, Robert J; Smart, Trevor G

2004-01-01

The glycine receptor (GlyR) α2A and α2B splice variants differ by a dual, adjacent amino acid substitution from α2AV58,T59 to α2BI58,A59 in the N-terminal extracellular domain. Comparing the effects of the GlyR agonists, glycine, β-alanine and taurine, on the GlyR α2 isoforms, revealed a significant increase in potency for all three agonists at the α2B variant. The sensitivities of the splice variants to the competitive antagonist, strychnine, and to the biphasic modulator Zn2+, were comparable. In contrast, the allosteric inhibitor picrotoxin was more potent on GlyR α2A compared to GlyR α2B receptors. Coexpression of α2A or α2B subunits with the GlyR β subunit revealed that the higher agonist potencies observed with the α2B homomer were retained for the α2Bβ heteromer. The identical sensitivity to strychnine combined with a reduction in the maximum current induced by the partial agonist taurine at the GlyR α2A homomer, suggested that the changed sensitivity to agonists is in accordance with a modulation of agonist efficacy rather than agonist affinity. An effect on agonist efficacy was also supported by using a structural model of the GlyR, localising the region of splice variation to the proposed docking region between GlyR loop 2 and the TM2-3 loop, an area associated with channel activation. The existence of a spasmodic mouse phenotype linked to a GlyR α1A52S mutation, the equivalent position to the source of the α2 splice variation, raises the possibility that the GlyR α2 splice variants may be responsible for distinct roles in neuronal function. PMID:15302677

Antinociceptive effects of fisetin against diabetic neuropathic pain in mice: Engagement of antioxidant mechanisms and spinal GABAA receptors.

PubMed

Zhao, Xin; Li, Xin-Lin; Liu, Xin; Wang, Chuang; Zhou, Dong-Sheng; Ma, Qing; Zhou, Wen-Hua; Hu, Zhen-Yu

2015-12-01

Peripheral painful neuropathy is one of the most common complications in diabetes and necessitates improved treatment. Fisetin, a naturally occurring flavonoid, has been reported to exert antidepressant-like effect in previous studies. As antidepressant drugs are employed clinically to treat neuropathic pain, this work aimed to investigate whether fisetin possess beneficial effect on diabetic neuropathic pain and explore the mechanism(s). We subjected mice to diabetes by a single intraperitoneal (i.p.) injection of streptozotocin (200mg/kg), and von Frey test or Hargreaves test was used to assess mechanical allodynia or thermal hyperalgesia, respectively. Chronic treatment of diabetic mice with fisetin not only ameliorated the established symptoms of thermal hyperalgesia and mechanical allodynia, but also arrested the development of neuropathic pain when given at low doses. Although chronic fisetin administration did not impact on the symptom of hyperglycemia in diabetic mice, it reduced exacerbated oxidative stress in tissues of spinal cord, dorsal root ganglion (DRG) and sciatic verve. Furthermore, the analgesic actions of fisetin were abolished by repetitive co-treatment with the reactive oxygen species (ROS) donor tert-butyl hydroperoxide (t-BOOH), but potentiated by the ROS scavenger phenyl-N-tert-butylnitrone (PBN). Finally, acute blockade of spinal GABAA receptors by bicuculline totally counteracted such fisetin analgesia. These findings indicate that chronic fisetin treatment can delay or correct neuropathic hyperalgesia and allodynia in mice with type 1 diabetes. Mechanistically, the present fisetin analgesia may be associated with its antioxidant activity, and spinal GABAA receptors are likely rendered as downstream targets. Copyright © 2015 Elsevier Ltd. All rights reserved.

Attenuating GABAA Receptor Signaling in Dopamine Neurons Selectively Enhances Reward Learning and Alters Risk Preference in Mice

PubMed Central

Parker, Jones G.; Wanat, Matthew J.; Soden, Marta E.; Ahmad, Kinza; Zweifel, Larry S.; Bamford, Nigel S.; Palmiter, Richard D.

2011-01-01

Phasic dopamine transmission encodes the value of reward-predictive stimuli and influences both learning and decision-making. Altered dopamine signaling is associated with psychiatric conditions characterized by risky choices such as pathological gambling. These observations highlight the importance of understanding how dopamine neuron activity is modulated. While excitatory drive onto dopamine neurons is critical for generating phasic dopamine responses, emerging evidence suggests that inhibitory signaling also modulates these responses. To address the functional importance of inhibitory signaling in dopamine neurons, we generated mice lacking the β3 subunit of the GABAA receptor specifically in dopamine neurons (β3-KO mice) and examined their behavior in tasks that assessed appetitive learning, aversive learning, and risk preference. Dopamine neurons in midbrain slices from β3-KO mice exhibited attenuated GABA-evoked inhibitory post-synaptic currents. Furthermore, electrical stimulation of excitatory afferents to dopamine neurons elicited more dopamine release in the nucleus accumbens of β3-KO mice as measured by fast-scan cyclic voltammetry. β3-KO mice were more active than controls when given morphine, which correlated with potential compensatory upregulation of GABAergic tone onto dopamine neurons. β3-KO mice learned faster in two food-reinforced learning paradigms, but extinguished their learned behavior normally. Enhanced learning was specific for appetitive tasks, as aversive learning was unaffected in β3-KO mice. Finally, we found that β3-KO mice had enhanced risk preference in a probabilistic selection task that required mice to choose between a small certain reward and a larger uncertain reward. Collectively, these findings identify a selective role for GABAA signaling in dopamine neurons in appetitive learning and decision-making. PMID:22114279

Relative Neurotoxicity of Ivermectin and Moxidectin in Mdr1ab (−/−) Mice and Effects on Mammalian GABA(A) Channel Activity

PubMed Central

Ménez, Cécile; Sutra, Jean-François; Prichard, Roger; Lespine, Anne

2012-01-01

The anthelmintics ivermectin (IVM) and moxidectin (MOX) display differences in toxicity in several host species. Entrance into the brain is restricted by the P-glycoprotein (P-gp) efflux transporter, while toxicity is mediated through the brain GABA(A) receptors. This study compared the toxicity of IVM and MOX in vivo and their interaction with GABA(A) receptors in vitro. Drug toxicity was assessed in Mdr1ab(−/−) mice P-gp-deficient after subcutaneous administration of increasing doses (0.11–2.0 and 0.23–12.9 µmol/kg for IVM and MOX in P-gp-deficient mice and half lethal doses (LD50) in wild-type mice). Survival was evaluated over 14-days. In Mdr1ab(−/−) mice, LD50 was 0.46 and 2.3 µmol/kg for IVM and MOX, respectively, demonstrating that MOX was less toxic than IVM. In P-gp-deficient mice, MOX had a lower brain-to-plasma concentration ratio and entered into the brain more slowly than IVM. The brain sublethal drug concentrations determined after administration of doses close to LD50 were, in Mdr1ab(−/−) and wild-type mice, respectively, 270 and 210 pmol/g for IVM and 830 and 740–1380 pmol/g for MOX, indicating that higher brain concentrations are required for MOX toxicity than IVM. In rat α1β2γ2 GABA channels expressed in Xenopus oocytes, IVM and MOX were both allosteric activators of the GABA-induced response. The Hill coefficient was 1.52±0.45 for IVM and 0.34±0.56 for MOX (p<0.001), while the maximum potentiation caused by IVM and MOX relative to GABA alone was 413.7±66.1 and 257.4±40.6%, respectively (p<0.05), showing that IVM causes a greater potentiation of GABA action on this receptor. Differences in the accumulation of IVM and MOX in the brain and in the interaction of IVM and MOX with GABA(A) receptors account for differences in neurotoxicity seen in intact and Mdr1-deficient animals. These differences in neurotoxicity of IVM and MOX are important in considering their use in humans. PMID:23133688

Glutamate and GABA in lateral hypothalamic mechanisms controlling food intake.

PubMed