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

Α2 GABAA receptor sub-units in the ventral hippocampus and α5 GABAA receptor sub-units in the dorsal hippocampus mediate anxiety and fear memory.

PubMed

McEown, K; Treit, D

2013-11-12

Temporary neuronal inactivation of the ventral hippocampus with the GABAA agonist muscimol suppresses unconditioned fear behavior (anxiety) but inactivation of the dorsal hippocampus does not. On the other hand, inactivating the dorsal hippocampus disrupts fear memory, while inactivating the ventral hippocampus does not. Here we investigate the roles of hippocampal GABAA receptor sub-units in mediating these anxiolytic and amnesic effects of GABAA receptor agonists. We microinfused TPA023 (α2 agonist) or TB-21007 (inverse α5 agonist) into the dorsal or ventral hippocampus prior to testing rats in two animal models of anxiety: the elevated plus-maze and shock-probe burying test. Twenty-four hours later rats were re-tested in the shock-probe chamber with a non-electrified probe to assess their memory of the initial shock-probe experience (i.e., fear memory). We found that TPA023 was anxiolytic in the plus-maze and shock-probe burying tests when microinfused into the ventral hippocampus. However, TPA023 did not affect anxiety-related behavior when infused into the dorsal hippocampus. Conversely, we found that the α5 sub-unit inverse agonist TB-21007 impaired rats' memory of the initial shock-probe experience when infused into the dorsal hippocampus, but not when infused into the ventral hippocampus. This double dissociation suggests that α2 GABAA receptor sub-units in the ventral hippocampus mediate unconditioned fear or anxiety, while α5 GABAA receptor sub-units in the dorsal hippocampus mediate conditioned fear memory. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

Extrasynaptic GABAA receptors in rat pontine reticular formation increase wakefulness.

PubMed

Vanini, Giancarlo; Baghdoyan, Helen A

2013-03-01

Gamma-aminobutyric acid (GABA) causes phasic inhibition via synaptic GABAA receptors and tonic inhibition via extrasynaptic GABAA receptors. GABA levels in the extracellular space regulate arousal state and cognition by volume transmission via extrasynaptic GABAA receptors. GABAergic transmission in the pontine reticular formation promotes wakefulness. No previous studies have determined whether an agonist at extrasynaptic GABAA receptors administered into the pontine reticular formation alters sleep and wakefulness. Therefore, this study used gaboxadol (THIP; agonist at extrasynaptic GABAA receptors that contain a δ subunit) to test the hypothesis that extrasynaptic GABAA receptors within the pontine reticular formation modulate sleep and wakefulness. Within/between subjects. University of Michigan. Adult male Crl:CD*(SD) (Sprague-Dawley) rats (n = 10). Microinjection of gaboxadol, the nonsubtype selective GABAA receptor agonist muscimol (positive control), and saline (negative control) into the rostral pontine reticular formation. Gaboxadol significantly increased wakefulness and decreased both nonrapid eye movement sleep and rapid eye movement sleep in a concentration-dependent manner. Relative to saline, gaboxadol did not alter electroencephalogram power. Microinjection of muscimol into the pontine reticular formation of the same rats that received gaboxadol increased wakefulness and decreased sleep. Tonic inhibition via extrasynaptic GABAA receptors that contain a δ subunit may be one mechanism by which the extracellular pool of endogenous GABA in the rostral pontine reticular formation promotes wakefulness. Vanini G; Baghdoyan HA. Extrasynaptic GABAA receptors in rat pontine reticular formation increase wakefulness. SLEEP 2013;36(3):337-343.

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.

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.

Extrasynaptic GABAA Receptors in Rat Pontine Reticular Formation Increase Wakefulness

PubMed Central

Vanini, Giancarlo; Baghdoyan, Helen A.

2013-01-01

Study Objectives: Gamma-aminobutyric acid (GABA) causes phasic inhibition via synaptic GABAA receptors and tonic inhibition via extrasynaptic GABAA receptors. GABA levels in the extracellular space regulate arousal state and cognition by volume transmission via extrasynaptic GABAA receptors. GABAergic transmission in the pontine reticular formation promotes wakefulness. No previous studies have determined whether an agonist at extrasynaptic GABAA receptors administered into the pontine reticular formation alters sleep and wakefulness. Therefore, this study used gaboxadol (THIP; agonist at extrasynaptic GABAA receptors that contain a δ subunit) to test the hypothesis that extrasynaptic GABAA receptors within the pontine reticular formation modulate sleep and wakefulness. Design: Within/between subjects. Setting: University of Michigan. Patients or Participants: Adult male Crl:CD*(SD) (Sprague-Dawley) rats (n = 10). Interventions: Microinjection of gaboxadol, the nonsubtype selective GABAA receptor agonist muscimol (positive control), and saline (negative control) into the rostral pontine reticular formation. Measurements and Results: Gaboxadol significantly increased wakefulness and decreased both nonrapid eye movement sleep and rapid eye movement sleep in a concentration-dependent manner. Relative to saline, gaboxadol did not alter electroencephalogram power. Microinjection of muscimol into the pontine reticular formation of the same rats that received gaboxadol increased wakefulness and decreased sleep. Conclusion: Tonic inhibition via extrasynaptic GABAA receptors that contain a δ subunit may be one mechanism by which the extracellular pool of endogenous GABA in the rostral pontine reticular formation promotes wakefulness. Citation: Vanini G; Baghdoyan HA. Extrasynaptic GABAA receptors in rat pontine reticular formation increase wakefulness. SLEEP 2013;36(3):337-343. PMID:23450652

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.

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-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

Synaptic pruning in the female hippocampus is triggered at puberty by extrasynaptic GABAA receptors on dendritic spines

PubMed Central

Afroz, Sonia; Parato, Julie; Shen, Hui; Smith, Sheryl Sue

2016-01-01

Adolescent synaptic pruning is thought to enable optimal cognition because it is disrupted in certain neuropathologies, yet the initiator of this process is unknown. One factor not yet considered is the α4βδ GABAA receptor (GABAR), an extrasynaptic inhibitory receptor which first emerges on dendritic spines at puberty in female mice. Here we show that α4βδ GABARs trigger adolescent pruning. Spine density of CA1 hippocampal pyramidal cells decreased by half post-pubertally in female wild-type but not α4 KO mice. This effect was associated with decreased expression of kalirin-7 (Kal7), a spine protein which controls actin cytoskeleton remodeling. Kal7 decreased at puberty as a result of reduced NMDAR activation due to α4βδ-mediated inhibition. In the absence of this inhibition, Kal7 expression was unchanged at puberty. In the unpruned condition, spatial re-learning was impaired. These data suggest that pubertal pruning requires α4βδ GABARs. In their absence, pruning is prevented and cognition is not optimal. DOI: http://dx.doi.org/10.7554/eLife.15106.001 PMID:27136678

Control of Inhibition by the Direct Action of Cannabinoids on GABAA Receptors.

PubMed

Golovko, Tatiana; Min, Rogier; Lozovaya, Natalia; Falconer, Caroline; Yatsenko, Natalia; Tsintsadze, Timur; Tsintsadze, Vera; Ledent, Catherine; Harvey, Robert J; Belelli, Delia; Lambert, Jeremy J; Rozov, Andrei; Burnashev, Nail

2015-09-01

Cannabinoids are known to regulate inhibitory synaptic transmission via activation of presynaptic G protein-coupled cannabinoid CB1 receptors (CB1Rs). Additionally, recent studies suggest that cannabinoids can also directly interact with recombinant GABAA receptors (GABAARs), potentiating currents activated by micromolar concentrations of γ-aminobutyric acid (GABA). However, the impact of this direct interaction on GABAergic inhibition in central nervous system is unknown. Here we report that currents mediated by recombinant GABAARs activated by high (synaptic) concentrations of GABA as well as GABAergic inhibitory postsynaptic currents (IPSCs) at neocortical fast spiking (FS) interneuron to pyramidal neuron synapses are suppressed by exogenous and endogenous cannabinoids in a CB1R-independent manner. This IPSC suppression may account for disruption of inhibitory control of pyramidal neurons by FS interneurons. At FS interneuron to pyramidal neuron synapses, endocannabinoids induce synaptic low-pass filtering of GABAAR-mediated currents evoked by high-frequency stimulation. The CB1R-independent suppression of inhibition is synapse specific. It does not occur in CB1R containing hippocampal cholecystokinin-positive interneuron to pyramidal neuron synapses. Furthermore, in contrast to synaptic receptors, the activity of extrasynaptic GABAARs in neocortical pyramidal neurons is enhanced by cannabinoids in a CB1R-independent manner. Thus, cannabinoids directly interact differentially with synaptic and extrasynaptic GABAARs, providing a potent novel context-dependent mechanism for regulation of inhibition. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Ethanol exposure during the third trimester equivalent does not affect GABAA or AMPA receptor-mediated spontaneous synaptic transmission in rat CA3 pyramidal neurons.

PubMed

Baculis, Brian Charles; Valenzuela, Carlos Fernando

2015-12-02

Ethanol exposure during the rodent equivalent to the 3(rd) trimester of human pregnancy (i.e., first 1-2 weeks of neonatal life) has been shown to produce structural and functional alterations in the CA3 hippocampal sub-region, which is involved in associative memory. Synaptic plasticity mechanisms dependent on retrograde release of brain-derived neurotrophic factor (BDNF) driven by activation of L-type voltage-gated Ca(2+) channels (L-VGCCs) are thought to play a role in stabilization of both GABAergic and glutamatergic synapses in CA3 pyramidal neurons. We previously showed that ethanol exposure during the first week of life blocks BDNF/L-VGCC-dependent long-term potentiation of GABAA receptor-mediated synaptic transmission in these neurons. Here, we tested whether this effect is associated with lasting alterations in GABAergic and glutamatergic transmission. Rats were exposed to air or ethanol for 3 h/day between postnatal days three and five in vapor inhalation chambers, a paradigm that produces peak serum ethanol levels near 0.3 g/dl. Whole-cell patch-clamp electrophysiological recordings of spontaneous inhibitory and excitatory postsynaptic currents (sIPSCs and sEPSCs, respectively) were obtained from CA3 pyramidal neurons in coronal brain slices prepared at postnatal days 13-17. Ethanol exposure did not significantly affect the frequency, amplitude, rise-time and half-width of either sIPSCs or sEPSCs. We show that an ethanol exposure paradigm known to inhibit synaptic plasticity mechanisms that may participate in the stabilization of GABAergic and glutamatergic synapses in CA3 pyramidal neurons does not produce lasting functional alterations in these synapses, suggesting that compensatory mechanisms restored the balance of excitatory and inhibitory synaptic transmission.

Regulated lysosomal trafficking as a mechanism for regulating GABAA receptor abundance at synapses in Caenorhabditis elegans.

PubMed

Davis, Kathleen M; Sturt, Brianne L; Friedmann, Andrew J; Richmond, Janet E; Bessereau, Jean-Louis; Grant, Barth D; Bamber, Bruce A

2010-08-01

GABA(A) receptor plasticity is important for both normal brain function and disease progression. We are studying GABA(A) receptor plasticity in Caenorhabditis elegans using a genetic approach. Acute exposure of worms to the GABA(A) agonist muscimol hyperpolarizes postsynaptic cells, causing paralysis. Worms adapt after several hours, but show uncoordinated locomotion consistent with decreased GABA signaling. Using patch-clamp and immunofluorescence approaches, we show that GABA(A) receptors are selectively removed from synapses during adaptation. Subunit mRNA levels were unchanged, suggesting a post-transcriptional mechanism. Mutants with defective lysosome function (cup-5) show elevated GABA(A) receptor levels at synapses prior to muscimol exposure. During adaptation, these receptors are removed more slowly, and accumulate in intracellular organelles positive for the late endosome marker GFP-RAB-7. These findings suggest that chronic agonist exposure increases endocytosis and lysosomal trafficking of GABA(A) receptors, leading to reduced levels of synaptic GABA(A) receptors and reduced postsynaptic GABA sensitivity.

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.

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.

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

Bidirectional control of spike timing by GABA(A) receptor-mediated inhibition during theta oscillation in CA1 pyramidal neurons.

PubMed

Kwag, Jeehyun; Paulsen, Ole

2009-08-26

Precisely controlled spike times relative to theta-frequency network oscillations play an important role in hippocampal memory processing. Here we study how inhibitory synaptic input during theta oscillation contributes to the control of spike timing. Using whole-cell patch-clamp recordings from CA1 pyramidal cells in vitro with dynamic clamp to simulate theta-frequency oscillation (5 Hz), we show that gamma-aminobutyric acid-A (GABA(A)) receptor-mediated inhibitory postsynaptic potentials (IPSPs) can not only delay but also advance the postsynaptic spike depending on the timing of the inhibition relative to the oscillation. Spike time advancement with IPSP was abolished by the h-channel blocker ZD7288 (10 microM), suggesting that IPSPs can interact with intrinsic membrane conductances to yield bidirectional control of spike timing.

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.

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

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.

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

Novel Fast Adapting Interneurons Mediate Cholinergic-Induced Fast GABAA IPSCs In Striatal Spiny Neurons

PubMed Central

Faust, Thomas W.; Assous, Maxime; Shah, Fulva; Tepper, James M.; Koós, Tibor

2015-01-01

Previous work suggests that neostriatal cholinergic interneurons control the activity of several classes of GABAergic interneurons through fast nicotinic receptor mediated synaptic inputs. Although indirect evidence has suggested the existence of several classes of interneurons controlled by this mechanism only one such cell type, the neuropeptide-Y expressing neurogliaform neuron, has been identified to date. Here we tested the hypothesis that in addition to the neurogliaform neurons that elicit slow GABAergic inhibitory responses, another interneuron type exists in the striatum that receives strong nicotinic cholinergic input and elicits conventional fast GABAergic synaptic responses in projection neurons. We obtained in vitro slice recordings from double transgenic mice in which Channelrhodopsin-2 was natively expressed in cholinergic neurons and a population of serotonin receptor-3a-Cre expressing GABAergic interneurons were visualized with tdTomato. We show that among the targeted GABAergic interneurons a novel type of interneuron, termed the fast-adapting interneuron, can be identified that is distinct from previously known interneurons based on immunocytochemical and electrophysiological criteria. We show using optogenetic activation of cholinergic inputs that fast-adapting interneurons receive a powerful supra-threshold nicotinic cholinergic input in vitro. Moreover, fast adapting neurons are densely connected to projection neurons and elicit fast, GABAA receptor mediated inhibitory postsynaptic responses. The nicotinic receptor mediated activation of fast-adapting interneurons may constitute an important mechanism through which cholinergic interneurons control the activity of projection neurons and perhaps the plasticity of their synaptic inputs when animals encounter reinforcing or otherwise salient stimuli. PMID:25865337

GABAA receptor dependent synaptic inhibition rapidly tunes KCC2 activity via the Cl--sensitive WNK1 kinase.

PubMed

Heubl, Martin; Zhang, Jinwei; Pressey, Jessica C; Al Awabdh, Sana; Renner, Marianne; Gomez-Castro, Ferran; Moutkine, Imane; Eugène, Emmanuel; Russeau, Marion; Kahle, Kristopher T; Poncer, Jean Christophe; Lévi, Sabine

2017-11-24

The K + -Cl - co-transporter KCC2 (SLC12A5) tunes the efficacy of GABA A receptor-mediated transmission by regulating the intraneuronal chloride concentration [Cl - ] i . KCC2 undergoes activity-dependent regulation in both physiological and pathological conditions. The regulation of KCC2 by synaptic excitation is well documented; however, whether the transporter is regulated by synaptic inhibition is unknown. Here we report a mechanism of KCC2 regulation by GABA A receptor (GABA A R)-mediated transmission in mature hippocampal neurons. Enhancing GABA A R-mediated inhibition confines KCC2 to the plasma membrane, while antagonizing inhibition reduces KCC2 surface expression by increasing the lateral diffusion and endocytosis of the transporter. This mechanism utilizes Cl - as an intracellular secondary messenger and is dependent on phosphorylation of KCC2 at threonines 906 and 1007 by the Cl - -sensing kinase WNK1. We propose this mechanism contributes to the homeostasis of synaptic inhibition by rapidly adjusting neuronal [Cl - ] i to GABA A R activity.

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

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

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

DISC1 Protein Regulates γ-Aminobutyric Acid, Type A (GABAA) Receptor Trafficking and Inhibitory Synaptic Transmission in Cortical Neurons.

PubMed

Wei, Jing; Graziane, Nicholas M; Gu, Zhenglin; Yan, Zhen

2015-11-13

Association studies have suggested that Disrupted-in-Schizophrenia 1 (DISC1) confers a genetic risk at the level of endophenotypes that underlies many major mental disorders. Despite the progress in understanding the significance of DISC1 at neural development, the mechanisms underlying DISC1 regulation of synaptic functions remain elusive. Because alterations in the cortical GABA system have been strongly linked to the pathophysiology of schizophrenia, one potential target of DISC1 that is critically involved in the regulation of cognition and emotion is the GABAA receptor (GABAAR). We found that cellular knockdown of DISC1 significantly reduced GABAAR-mediated synaptic and whole-cell current, whereas overexpression of wild-type DISC1, but not the C-terminal-truncated DISC1 (a schizophrenia-related mutant), significantly increased GABAAR currents in pyramidal neurons of the prefrontal cortex. These effects were accompanied by DISC1-induced changes in surface GABAAR expression. Moreover, the regulation of GABAARs by DISC1 knockdown or overexpression depends on the microtubule motor protein kinesin 1 (KIF5). Our results suggest that DISC1 exerts an important effect on GABAergic inhibitory transmission by regulating KIF5/microtubule-based GABAAR trafficking in the cortex. The knowledge gained from this study would shed light on how DISC1 and the GABA system are linked mechanistically and how their interactions are critical for maintaining a normal mental state. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

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

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.

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

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.

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.

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

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

Mitochondrial reactive oxygen species regulate the strength of inhibitory GABA-mediated synaptic transmission

NASA Astrophysics Data System (ADS)

Accardi, Michael V.; Daniels, Bryan A.; Brown, Patricia M. G. E.; Fritschy, Jean-Marc; Tyagarajan, Shiva K.; Bowie, Derek

2014-01-01

Neuronal communication imposes a heavy metabolic burden in maintaining ionic gradients essential for action potential firing and synaptic signalling. Although cellular metabolism is known to regulate excitatory neurotransmission, it is still unclear whether the brain’s energy supply affects inhibitory signalling. Here we show that mitochondrial-derived reactive oxygen species (mROS) regulate the strength of postsynaptic GABAA receptors at inhibitory synapses of cerebellar stellate cells. Inhibition is strengthened through a mechanism that selectively recruits α3-containing GABAA receptors into synapses with no discernible effect on resident α1-containing receptors. Since mROS promotes the emergence of postsynaptic events with unique kinetic properties, we conclude that newly recruited α3-containing GABAA receptors are activated by neurotransmitter released onto discrete postsynaptic sites. Although traditionally associated with oxidative stress in neurodegenerative disease, our data identify mROS as a putative homeostatic signalling molecule coupling cellular metabolism to the strength of inhibitory transmission.

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

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

Chronic Exposure to Anabolic Androgenic Steroids Alters Neuronal Function in the Mammalian Forebrain via Androgen Receptor- and Estrogen Receptor-Mediated Mechanisms

PubMed Central

Penatti, Carlos A A; Porter, Donna M; Henderson, Leslie P

2009-01-01

Anabolic androgenic steroids (AAS) can promote detrimental effects on social behaviors for which γ-aminobutyric acid type A (GABAA) receptor-mediated circuits in the forebrain play a critical role. While all AAS bind to androgen receptors (AR), they may also be aromatized to estrogens and thus potentially impart effects via estrogen receptors (ER). Chronic exposure of wild type male mice to a combination of chemically distinct AAS increased action potential (AP) frequency, selective GABAA receptor subunit mRNAs, and GABAergic synaptic current decay in the medial preoptic area (mPOA). Experiments performed with pharmacological agents and in AR-deficient Tfm mutant mice suggest that the AAS-dependent enhancement of GABAergic transmission in wild type mice is AR-mediated. In AR-deficient mice, the AAS elicited dramatically different effects, decreasing AP frequency, sIPSC amplitude and frequency and the expression of selective GABAA receptor subunit mRNAs. Surprisingly, in the absence of AR signaling, the data indicate that the AAS do not act as ER agonists, but rather suggest a novel in vivo action in which the AAS inhibit aromatase and impair endogenous ER signaling. These results show that the AAS have the capacity to alter neuronal function in the forebrain via multiple steroid signaling mechanisms and suggest that effects of these steroids in the brain will depend not only on the balance of AR- vs. ER-mediated regulation for different target genes, but also on the ability of these drugs to alter steroid metabolism and thus the endogenous steroid milieu. PMID:19812324

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

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

Anesthetic agent-specific effects on synaptic inhibition.

PubMed

MacIver, M Bruce

2014-09-01

Anesthetics enhance γ-aminobutyric acid (GABA)-mediated inhibition in the central nervous system. Different agents have been shown to act on tonic versus synaptic GABA receptors to different degrees, but it remains unknown whether different forms of synaptic inhibition are also differentially engaged. With this in mind, we tested the hypothesis that different types of GABA-mediated synapses exhibit different anesthetic sensitivities. The present study compared effects produced by isoflurane, halothane, pentobarbital, thiopental, and propofol on paired-pulse GABAA receptor-mediated synaptic inhibition. Effects on glutamate-mediated facilitation were also studied. Synaptic responses were measured in rat hippocampal brain slices. Orthodromic paired-pulse stimulation was used to assess anesthetic effects on either glutamate-mediated excitatory inputs or GABA-mediated inhibitory inputs to CA1 neurons. Antidromic stimulation was used to assess anesthetic effects on CA1 background excitability. Agents were studied at equieffective concentrations for population spike depression to compare their relative degree of effect on synaptic inhibition. Differing degrees of anesthetic effect on paired-pulse facilitation at excitatory glutamate synapses were evident, and blocking GABA inhibition revealed a previously unseen presynaptic action for pentobarbital. Although all 5 anesthetics depressed synaptically evoked excitation of CA1 neurons, the involvement of enhanced GABA-mediated inhibition differed considerably among agents. Single-pulse inhibition was enhanced by propofol, thiopental, and pentobarbital, but only marginally by halothane and isoflurane. In contrast, isoflurane enhanced paired-pulse inhibition strongly, as did thiopental, but propofol, pentobarbital, and halothane were less effective. These observations support the idea that different GABA synapses use receptors with differing subunit compositions and that anesthetics exhibit differing degrees of selectivity for

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

Context-Dependent Modulation of αβγ and αβγ GABAA Receptors by Penicillin: Implications for Phasic and Tonic Inhibition

PubMed Central

Feng, Hua-Jun; Botzolakis, Emmanuel J.; Macdonald, Robert L.

2009-01-01

Summary Penicillin, an open-channel blocker of GABAA receptors, was recently reported to inhibit phasic, but not tonic, currents in hippocampal neurons. To distinguish between isoform-specific and context-dependent modulation as possible explanations for this selectivity, the effects of penicillin were evaluated on recombinant GABAA receptors expressed in HEK293T cells. When co-applied with saturating GABA, penicillin decreased peak amplitude, induced rebound, and prolonged deactivation of currents evoked from both synaptic and extrasynaptic receptor isoforms. However, penicillin had isoform-specific effects on the extent of desensitization, reflecting its ability to differentially modulate peak (non-equilibrium) and residual (near-equilibrium) currents. This suggested that the context of activation could determine the apparent sensitivity of a given receptor isoform to penicillin. To test this hypothesis, we explored the ability of penicillin to modulate synaptic and extrasynaptic isoforms that were activated under more physiologically relevant conditions. Interestingly, while currents evoked from synaptic isoforms under phasic conditions (transient activation by a saturating concentration of GABA) were substantially inhibited by penicillin, currents evoked from extrasynaptic isoforms under tonic conditions (prolonged application by a sub-saturating concentration of GABA) were minimally affected. We therefore concluded that the reported inability of penicillin to modulate tonic currents could not simply be attributed to insensitivity of extrasynaptic receptors, but rather, reflected an inability to modulate these receptors in their native context of activation. PMID:18775733

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.

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

Anesthetic Agent-Specific Effects on Synaptic Inhibition

PubMed Central

MacIver, M. Bruce

2014-01-01

Background Anesthetics enhance gamma-aminobutyric acid (GABA)-mediated inhibition in the central nervous system. Different agents have been shown to act on tonic versus synaptic GABA receptors to different degrees, but it remains unknown whether different forms of synaptic inhibition are also differentially engaged. With this in mind, we tested the hypothesis that different types of GABA-mediated synapses exhibit different anesthetic sensitivities. The present study compared effects produced by isoflurane, halothane, pentobarbital, thiopental and propofol on paired pulse GABAA receptor-mediated synaptic inhibition. Effects on glutamate-mediated facilitation were also studied. Methods Synaptic responses were measured in rat hippocampal brain slices. Orthodromic paired pulse stimulation was used to assess anesthetic effects on either glutamate-mediated excitatory inputs or GABA-mediated inhibitory inputs to CA1 neurons. Antidromic stimulation was used to assess anesthetic effects on CA1 background excitability. Agents were studied at equi-effective concentrations for population spike depression to compare their relative degree of effect on synaptic inhibition. Results Differing degrees of anesthetic effect on paired pulse facilitation at excitatory glutamate synapses were evident, and blocking GABA inhibition revealed a previously unseen presynaptic action for pentobarbital. Although all five anesthetics depressed synaptically evoked excitation of CA1 neurons, the involvement of enhanced GABA-mediated inhibition differed considerably among agents. Single pulse inhibition was enhanced by propofol, thiopental and pentobarbital, but only marginally by halothane and isoflurane. In contrast, isoflurane enhanced paired pulse inhibition strongly, as did thiopental, but propofol, pentobarbital and halothane were less effective. Conclusions These observations support the idea that different GABA synapses use receptors with differing subunit compositions, and that anesthetics

Postnatal aniracetam treatment improves prenatal ethanol induced attenuation of AMPA receptor-mediated synaptic transmission.

PubMed

Wijayawardhane, Nayana; Shonesy, Brian C; Vaglenova, Julia; Vaithianathan, Thirumalini; Carpenter, Mark; Breese, Charles R; Dityatev, Alexander; Suppiramaniam, Vishnu

2007-06-01

Aniracetam is a nootropic compound and an allosteric modulator of AMPA receptors (AMPARs) which mediate synaptic mechanisms of learning and memory. Here we analyzed impairments in AMPAR-mediated synaptic transmission caused by moderate prenatal ethanol exposure and investigated the effects of postnatal aniracetam treatment on these abnormalities. Pregnant Sprague-Dawley rats were gavaged with ethanol or isocaloric sucrose throughout pregnancy, and subsequently the offspring were treated with aniracetam on postnatal days (PND) 18 to 27. Hippocampal slices prepared from these pups on PND 28 to 34 were used for the whole-cell patch-clamp recordings of AMPAR-mediated spontaneous and miniature excitatory postsynaptic currents in CA1 pyramidal cells. Our results indicate that moderate ethanol exposure during pregnancy results in impaired hippocampal AMPAR-mediated neurotransmission, and critically timed aniracetam treatment can abrogate this deficiency. These results highlight the possibility that aniracetam treatment can restore synaptic transmission and ameliorate cognitive deficits associated with the fetal alcohol syndrome.

Control of synaptic function by endocannabinoid-mediated retrograde signaling.

PubMed

Kano, Masanobu

2014-01-01

Since the first reports in 2001, great advances have been made towards the understanding of endocannabinoid-mediated synaptic modulation. Electrophysiological studies have revealed that one of the two major endocannabinoids, 2-arachidonoylglycerol (2-AG), is produced from membrane lipids upon postsynaptic Ca(2+) elevation and/or activation of Gq/11-coupled receptors, and released from postsynaptic neurons. The released 2-AG then acts retrogradely onto presynaptic cannabinoid CB1 receptors and induces suppression of neurotransmitter release either transiently or persistently. These forms of 2-AG-mediated retrograde synaptic modulation are functional throughout the brain. The other major endocannabinoid, anandamide, mediates a certain form of endocannabinoid-mediated long-term depression (LTD). Anandamide also functions as an agonist for transient receptor potential vanilloid receptor type 1 (TRPV1) and mediates endocannabinoid-independent and TRPV1-dependent forms of LTD. It has also been demonstrated that the endocannabinoid system itself is plastic, which can be either up- or down-regulated by experimental or environmental conditions. In this review, I will make an overview of the mechanisms underlying endocannabinoid-mediated synaptic modulation.

Control of synaptic function by endocannabinoid-mediated retrograde signaling

PubMed Central

KANO, Masanobu

2014-01-01

Since the first reports in 2001, great advances have been made towards the understanding of endocannabinoid-mediated synaptic modulation. Electrophysiological studies have revealed that one of the two major endocannabinoids, 2-arachidonoylglycerol (2-AG), is produced from membrane lipids upon postsynaptic Ca2+ elevation and/or activation of Gq/11-coupled receptors, and released from postsynaptic neurons. The released 2-AG then acts retrogradely onto presynaptic cannabinoid CB1 receptors and induces suppression of neurotransmitter release either transiently or persistently. These forms of 2-AG-mediated retrograde synaptic modulation are functional throughout the brain. The other major endocannabinoid, anandamide, mediates a certain form of endocannabinoid-mediated long-term depression (LTD). Anandamide also functions as an agonist for transient receptor potential vanilloid receptor type 1 (TRPV1) and mediates endocannabinoid-independent and TRPV1-dependent forms of LTD. It has also been demonstrated that the endocannabinoid system itself is plastic, which can be either up- or down-regulated by experimental or environmental conditions. In this review, I will make an overview of the mechanisms underlying endocannabinoid-mediated synaptic modulation. PMID:25169670

Involvement of GABAA receptor in Bufo arenarum oocyte maturation.

PubMed

Toranzo, G Sánchez; Zelarayán, L; Bonilla, F; Oterino, J; Bühler, M I

2008-05-01

Amphibian oocytes meiotic arrest is released under the stimulus of progesterone; this hormone interacts with the oocyte surface and starts a cascade of events leading to the activation of a cytoplasmic maturation promoting factor (MPF) that induces germinal vesicle breakdown (GVBD), chromosome condensation and extrusion of the first polar body. The aim of this work was to determine whether the activation of a GABAA receptor is able to induce GVBD in fully grown denuded oocytes of Bufo arenarum and to analyse its possible participation in progesterone-induced maturation. We also evaluated the role of purines and phospholipids in the maturation process induced by a GABAA receptor agonist such as muscimol. Our results indicated that the activation of the GABAA receptor by muscimol induces maturation in a dose- and time-dependent manner and that this activation is a genuine maturation that enables oocytes to form pronuclei. Assays with a receptor antagonist, picrotoxine, showed that the maturation induced by muscimol was inhibited. Treatment with picrotoxine, however, shows that the participation of GABAA receptor in progesterone-induced maturation is not significant. In addition, our results indicate that high intracellular levels of purines obtained by the use of db-AMPc and theophylline or the inhibition of the phosphatidylinositol 4,5-bisphosphate (PIP2 hydrolysis by neomycin and PIP2 turn over by LiCl, respectively, inhibited the maturation induced by muscimol. Treatment with H-7 indicated, however, that PKC activation is not necessary for GVBD induced by the GABAA receptor agonist. Results suggest that the transduction pathway used by the GABAA receptor to induce maturation is different from those used by progesterone.

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

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.

Modulation of NMDA and AMPA-Mediated Synaptic Transmission by CB1 Receptors in Frontal Cortical Pyramidal Cells

PubMed Central

Li, Qiang; Yan, Haidun; Wilson, Wilkie A.; Swartzwelder, H. Scott

2010-01-01

Although the endogenous cannabinoid system modulates a variety of physiological and pharmacological processes, the specific role of cannabinoid CB1 receptors in the modulation of glutamatergic neurotransmission and neural plasticity is not well understood. Using whole-cell patch clamp recording techniques, evoked or spontaneous excitatory postsynaptic currents (eEPSCs or sEPSCs) were recorded from visualized, layer II/III pyramidal cells in frontal cortical slices from rat brain. Bath application of the CB1 receptor agonist, WIN 55212-2 (WIN), reduced the amplitude of NMDA receptor-mediated EPSCs in a concentration-dependent manner. When co-applied with the specific CB1 antagonists, AM251 or AM281, WIN did not suppress NMDA receptor mediated EPSCs. WIN also reduced the amplitude of evoked AMPA receptor-mediated EPSCs, an effect that was also reversed by AM251. Both the frequency and amplitude of spontaneous AMPA receptor-mediated EPSCs were significantly reduced by WIN. In contrast, WIN reduced the frequency, but not the amplitude of miniature EPSCs, suggesting that the suppression of glutmatergic activity by CB1 receptors in the frontal neocortex is mediated by a pre-synaptic mechanism. Taken together, these data indicate a critical role for endocannabinoid signaling in the regulation of excitatory synaptic transmission in frontal neocortex, and suggest a possible neuronal mechanism whereby THC regulates cortical function. PMID:20420813

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

Fragrant Dioxane Derivatives Identify β1-Subunit-containing GABAA Receptors*

PubMed Central

Sergeeva, Olga A.; Kletke, Olaf; Kragler, Andrea; Poppek, Anja; Fleischer, Wiebke; Schubring, Stephan R.; Görg, Boris; Haas, Helmut L.; Zhu, Xin-Ran; Lübbert, Hermann; Gisselmann, Günter; Hatt, Hanns

2010-01-01

Nineteen GABAA receptor (GABAAR) subunits are known in mammals with only a restricted number of functionally identified native combinations. The physiological role of β1-subunit-containing GABAARs is unknown. Here we report the discovery of a new structural class of GABAAR positive modulators with unique β1-subunit selectivity: fragrant dioxane derivatives (FDD). At heterologously expressed α1βxγ2L (x-for 1,2,3) GABAAR FDD were 6 times more potent at β1- versus β2- and β3-containing receptors. Serine at position 265 was essential for the high sensitivity of the β1-subunit to FDD and the β1N286W mutation nearly abolished modulation; vice versa the mutation β3N265S shifted FDD sensitivity toward the β1-type. In posterior hypothalamic neurons controlling wakefulness GABA-mediated whole-cell responses and GABAergic synaptic currents were highly sensitive to FDD, in contrast to β1-negative cerebellar Purkinje neurons. Immunostaining for the β1-subunit and the potency of FDD to modulate GABA responses in cultured hypothalamic neurons was drastically diminished by β1-siRNA treatment. In conclusion, with the help of FDDs we reveal a functional expression of β1-containing GABAARs in the hypothalamus, offering a new tool for studies on the functional diversity of native GABAARs. PMID:20511229

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

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.

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

The role of gamma-aminobutyric acid/glycinergic synaptic transmission in mediating bilirubin-induced hyperexcitation in developing auditory neurons.

PubMed

Yin, Xin-Lu; Liang, Min; Shi, Hai-Bo; Wang, Lu-Yang; Li, Chun-Yan; Yin, Shan-Kai

2016-01-05

Hyperbilirubinemia is a common clinical phenomenon observed in human newborns. A high level of bilirubin can result in severe jaundice and bilirubin encephalopathy. However, the cellular mechanisms underlying bilirubin excitotoxicity are unclear. Our previous studies showed the action of gamma-aminobutyric acid (GABA)/glycine switches from excitatory to inhibitory during development in the ventral cochlear nucleus (VCN), one of the most sensitive auditory nuclei to bilirubin toxicity. In the present study, we investigated the roles of GABAA/glycine receptors in the induction of bilirubin hyperexcitation in early developing neurons. Using the patch clamp technique, GABAA/glycine receptor-mediated spontaneous inhibitory synaptic currents (sIPSCs) were recorded from bushy and stellate cells in acute brainstem slices from young mice (postnatal day 2-6). Bilirubin significantly increased the frequency of sIPSCs, and this effect was prevented by pretreatments of slices with either fast or slow Ca(2+) chelators BAPTA-AM and EGTA-AM suggesting that bilirubin can increase the release of GABA/glycine via Ca(2+)-dependent mechanisms. Using cell-attached recording configuration, we found that antagonists of GABAA and glycine receptors strongly attenuated spontaneous spiking firings in P2-6 neurons but produced opposite effect in P15-19 neurons. Furthermore, these antagonists reversed bilirubin-evoked hyperexcitability in P2-6 neurons, indicating that excitatory action of GABA/glycinergic transmission specifically contribute to bilirubin-induced hyperexcitability in the early stage of development. Our results suggest that bilirubin-induced enhancement of presynaptic release GABA/Glycine via Ca(2+)-dependent mechanisms may play a critical role in mediating neuronal hyperexcitation associated with jaundice, implicating potential new strategies for predicting, preventing, and treating bilirubin neurotoxicity. Copyright © 2015. Published by Elsevier Ireland 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

Activation of endogenous GABAA channels on airway smooth muscle potentiates isoproterenol-mediated relaxation.

PubMed

Gallos, George; Gleason, Neil R; Zhang, Yi; Pak, Sang-Woo; Sonett, J R; Yang, Jay; Emala, Charles W

2008-12-01

Reactive airway disease predisposes patients to episodes of acute smooth muscle mediated bronchoconstriction. We have for the first time recently demonstrated the expression and function of endogenous ionotropic GABA(A) channels on airway smooth muscle cells. We questioned whether endogenous GABA(A) channels on airway smooth muscle could augment beta-agonist-mediated relaxation. Guinea pig tracheal rings or human bronchial airway smooth muscles were equilibrated in organ baths with continuous digital tension recordings. After pretreatment with or without the selective GABA(A) antagonist gabazine (100 muM), airway muscle was contracted with acetylcholine or beta-ala neurokinin A, followed by relaxation induced by cumulatively increasing concentrations of isoproterenol (1 nM to 1 muM) in the absence or presence of the selective GABA(A) agonist muscimol (10-100 muM). In separate experiments, guinea pig tracheal rings were pretreated with the large conductance K(Ca) channel blocker iberiotoxin (100 nM) after an EC(50) contraction with acetylcholine but before cumulatively increasing concentrations of isoproterenol (1 nM to 1 uM) in the absence or presence of muscimol (100 uM). GABA(A) activation potentiated the relaxant effects of isoproterenol after an acetylcholine or tachykinin-induced contraction in guinea pig tracheal rings or an acetylcholine-induced contraction in human endobronchial smooth muscle. This muscimol-induced potentiation of relaxation was abolished by gabazine pretreatment but persisted after blockade of the maxi K(Ca) channel. Selective activation of endogenous GABA(A) receptors significantly augments beta-agonist-mediated relaxation of guinea pig and human airway smooth muscle, which may have important therapeutic implications for patients in severe bronchospasm.

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

Fragrant dioxane derivatives identify beta1-subunit-containing GABAA receptors.

PubMed

Sergeeva, Olga A; Kletke, Olaf; Kragler, Andrea; Poppek, Anja; Fleischer, Wiebke; Schubring, Stephan R; Görg, Boris; Haas, Helmut L; Zhu, Xin-Ran; Lübbert, Hermann; Gisselmann, Günter; Hatt, Hanns

2010-07-30

Nineteen GABA(A) receptor (GABA(A)R) subunits are known in mammals with only a restricted number of functionally identified native combinations. The physiological role of beta1-subunit-containing GABA(A)Rs is unknown. Here we report the discovery of a new structural class of GABA(A)R positive modulators with unique beta1-subunit selectivity: fragrant dioxane derivatives (FDD). At heterologously expressed alpha1betaxgamma2L (x-for 1,2,3) GABA(A)R FDD were 6 times more potent at beta1- versus beta2- and beta3-containing receptors. Serine at position 265 was essential for the high sensitivity of the beta1-subunit to FDD and the beta1N286W mutation nearly abolished modulation; vice versa the mutation beta3N265S shifted FDD sensitivity toward the beta1-type. In posterior hypothalamic neurons controlling wakefulness GABA-mediated whole-cell responses and GABAergic synaptic currents were highly sensitive to FDD, in contrast to beta1-negative cerebellar Purkinje neurons. Immunostaining for the beta1-subunit and the potency of FDD to modulate GABA responses in cultured hypothalamic neurons was drastically diminished by beta1-siRNA treatment. In conclusion, with the help of FDDs we reveal a functional expression of beta1-containing GABA(A)Rs in the hypothalamus, offering a new tool for studies on the functional diversity of native GABA(A)Rs.

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

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

An autism-associated point mutation in the neuroligin cytoplasmic tail selectively impairs AMPA receptor-mediated synaptic transmission in hippocampus.

PubMed

Etherton, Mark R; Tabuchi, Katsuhiko; Sharma, Manu; Ko, Jaewon; Südhof, Thomas C

2011-06-03

Neuroligins are evolutionarily conserved postsynaptic cell-adhesion molecules that function, at least in part, by forming trans-synaptic complexes with presynaptic neurexins. Different neuroligin isoforms perform diverse functions and exhibit distinct intracellular localizations, but contain similar cytoplasmic sequences whose role remains largely unknown. Here, we analysed the effect of a single amino-acid substitution (R704C) that targets a conserved arginine residue in the cytoplasmic sequence of all neuroligins, and that was associated with autism in neuroligin-4. We introduced the R704C mutation into mouse neuroligin-3 by homologous recombination, and examined its effect on synapses in vitro and in vivo. Electrophysiological and morphological studies revealed that the neuroligin-3 R704C mutation did not significantly alter synapse formation, but dramatically impaired synapse function. Specifically, the R704C mutation caused a major and selective decrease in AMPA receptor-mediated synaptic transmission in pyramidal neurons of the hippocampus, without similarly changing NMDA or GABA receptor-mediated synaptic transmission, and without detectably altering presynaptic neurotransmitter release. Our results suggest that the cytoplasmic tail of neuroligin-3 has a central role in synaptic transmission by modulating the recruitment of AMPA receptors to postsynaptic sites at excitatory synapses.

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

PSD-95 regulates synaptic kainate receptors at mouse hippocampal mossy fiber-CA3 synapses.

PubMed

Suzuki, Etsuko; Kamiya, Haruyuki

2016-06-01

Kainate-type glutamate receptors (KARs) are the third class of ionotropic glutamate receptors whose activation leads to the unique roles in regulating synaptic transmission and circuit functions. In contrast to AMPA receptors (AMPARs), little is known about the mechanism of synaptic localization of KARs. PSD-95, a major scaffold protein of the postsynaptic density, is a candidate molecule that regulates the synaptic KARs. Although PSD-95 was shown to bind directly to KARs subunits, it has not been tested whether PSD-95 regulates synaptic KARs in intact synapses. Using PSD-95 knockout mice, we directly investigated the role of PSD-95 in the KARs-mediated components of synaptic transmission at hippocampal mossy fiber-CA3 synapse, one of the synapses with the highest density of KARs. Mossy fiber EPSCs consist of AMPA receptor (AMPAR)-mediated fast component and KAR-mediated slower component, and the ratio was significantly reduced in PSD-95 knockout mice. The size of KARs-mediated field EPSP reduced in comparison with the size of the fiber volley. Analysis of KARs-mediated miniature EPSCs also suggested reduced synaptic KARs. All the evidence supports critical roles of PSD-95 in regulating synaptic KARs. Copyright © 2015 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

Synaptic excitation mediated by AMPA receptors in rat cerebellar slices is selectively enhanced by aniracetam and cyclothiazide.

PubMed

Boxall, A R; Garthwaite, J

1995-05-01

AMPA receptors mediate fast, glutamatergic synaptic transmission in the central nervous system. The time-course of the associated postsynaptic current has been suggested to be determined principally by the kinetics of glutamate binding and receptor desensitization. Aniracetam and cyclothiazide are drugs capable of selectively preventing desensitization of the AMPA receptor. To investigate the relevance of desensitization to fast synaptic transmission in the cerebellum we have tested these compounds against AMPA-induced depolarizations and postsynaptic potentials using the grease-gap recording technique. Aniracetam (1 microM-5 mM) and cyclothiazide (1 microM-500 microM) both enhanced the depolarising action of AMPA (1 microM) on Purkinje cells in a concentration-dependent manner. At the highest concentrations tested, the increases over controls were approximately 600% and 800% respectively. Aniracetam also increased, in a concentration-dependent manner, the amplitude of the evoked synaptic potentials of both parallel fibre-Purkinje cell and mossy fibre-granule cell pathways, with the highest concentrations tested enhancing the potentials by approximately 60% and 75% respectively. These data suggest that, at two different synapses in the cerebellum, AMPA receptor desensitization occurs physiologically and is likely to contribute to the shape of fast synaptic currents.

Pre-synaptic kainate receptor-mediated facilitation of glutamate release involves PKA and Ca(2+) -calmodulin at thalamocortical synapses.

PubMed

Andrade-Talavera, Yuniesky; Duque-Feria, Paloma; Sihra, Talvinder S; Rodríguez-Moreno, Antonio

2013-09-01

We have investigated the mechanisms underlying the facilitatory modulation mediated by kainate receptor (KAR) activation in the cortex, using isolated nerve terminals (synaptosomes) and slice preparations. In cortical nerve terminals, kainate (KA, 100 μM) produced an increase in 4-aminopyridine (4-AP)-evoked glutamate release. In thalamocortical slices, KA (1 μM) produced an increase in the amplitude of evoked excitatory post-synaptic currents (eEPSCs) at synapses established between thalamic axon terminals from the ventrobasal nucleus onto stellate neurons of L4 of the somatosensory cortex. In both, synaptosomes and slices, the effect of KA was antagonized by 6-cyano-7-nitroquinoxaline-2,3-dione, and persisted after pre-treatment with a cocktail of antagonists of other receptors whose activation could potentially have produced facilitation of release indirectly. Mechanistically, the observed effects of KA appear to be congruent in synaptosomal and slice preparations. Thus, the facilitation by KA of synaptosomal glutamate release and thalamocortical synaptic transmission were suppressed by the inhibition of protein kinase A and occluded by the stimulation of adenylyl cyclase. Dissecting this G-protein-independent regulation further in thalamocortical slices, the KAR-mediated facilitation of synaptic transmission was found to be sensitive to the block of Ca(2+) permeant KARs by philanthotoxin. Intriguingly, the synaptic facilitation was abrogated by depletion of intracellular Ca(2+) stores by thapsigargin, or inhibition of Ca(2+) -induced Ca(2+) -release by ryanodine. Thus, the KA-mediated modulation was contingent on both Ca(2+) entry through Ca(2+) -permeable KARs and liberation of intracellular Ca(2+) stores. Finally, sensitivity to W-7 indicated that the increased cytosolic [Ca(2+) ] underpinning KAR-mediated regulation of synaptic transmission at thalamocortical synapses, requires downstream activation of calmodulin. We conclude that neocortical pre-synaptic

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.

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

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

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

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

Chronic nicotine treatment differentially modifies acute nicotine and alcohol actions on GABA(A) and glutamate receptors in hippocampal brain slices.

PubMed

Proctor, William R; Dobelis, Peter; Moritz, Anna T; Wu, Peter H

2011-03-01

Tobacco and alcohol are often co-abused producing interactive effects in the brain. Although nicotine enhances memory while ethanol impairs it, variable cognitive changes have been reported from concomitant use. This study was designed to determine how nicotine and alcohol interact at synaptic sites to modulate neuronal processes. Acute effects of nicotine, ethanol, and both drugs on synaptic excitatory glutamatergic and inhibitory GABAergic transmission were measured using whole-cell recording in hippocampal CA1 pyramidal neurons from brain slices of mice on control or nicotine-containing diets. Acute nicotine (50 nM) enhanced both GABAergic and glutamatergic synaptic transmission; potentiated GABA(A) receptor currents via activation of α7* and α4β2* nAChRs, and increased N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor currents through α7* receptors. While ethanol (80 mM) also increased GABA(A) currents, it inhibited NMDA currents. Although ethanol had no effect on AMPA currents, it blocked nicotine-induced increases in NMDA and AMPA currents. Following chronic nicotine treatment, acute nicotine or ethanol did not affect NMDA currents, while the effects of GABAergic responses were not altered. Acute ethanol ingestion selectively attenuated nicotine enhancement of excitatory glutamatergic NMDA and AMPA receptor function, suggesting an overall reduction in excitatory output from the hippocampus. It also indicated that ethanol could decrease the beneficial effects of nicotine on memory performance. In addition, chronic nicotine treatment produced tolerance to the effects of nicotine and cross-tolerance to the effects of ethanol on glutamatergic activity, leading to a potential increase in the use of these drugs. British Journal of Pharmacology © 2011 The British Pharmacological Society. No claim to original US government works.

Neurons generated by direct conversion of fibroblasts reproduce synaptic phenotype caused by autism-associated neuroligin-3 mutation.

PubMed

Chanda, Soham; Marro, Samuele; Wernig, Marius; Südhof, Thomas C

2013-10-08

Recent studies suggest that induced neuronal (iN) cells that are directly transdifferentiated from nonneuronal cells provide a powerful opportunity to examine neuropsychiatric diseases. However, the validity of using this approach to examine disease-specific changes has not been demonstrated. Here, we analyze the phenotypes of iN cells that were derived from murine embryonic fibroblasts cultured from littermate wild-type and mutant mice carrying the autism-associated R704C substitution in neuroligin-3. We show that neuroligin-3 R704C-mutant iN cells exhibit a large and selective decrease in AMPA-type glutamate receptor-mediated synaptic transmission without changes in NMDA-type glutamate receptor- or in GABAA receptor-mediated synaptic transmission. Thus, the synaptic phenotype observed in R704C-mutant iN cells replicates the previously observed phenotype of R704C-mutant neurons. Our data show that the effect of the R704C mutation is applicable even to neurons transdifferentiated from fibroblasts and constitute a proof-of-concept demonstration that iN cells can be used for cellular disease modeling.

Neurons generated by direct conversion of fibroblasts reproduce synaptic phenotype caused by autism-associated neuroligin-3 mutation

PubMed Central

Chanda, Soham; Marro, Samuele; Wernig, Marius; Südhof, Thomas C.

2013-01-01

Recent studies suggest that induced neuronal (iN) cells that are directly transdifferentiated from nonneuronal cells provide a powerful opportunity to examine neuropsychiatric diseases. However, the validity of using this approach to examine disease-specific changes has not been demonstrated. Here, we analyze the phenotypes of iN cells that were derived from murine embryonic fibroblasts cultured from littermate wild-type and mutant mice carrying the autism-associated R704C substitution in neuroligin-3. We show that neuroligin-3 R704C-mutant iN cells exhibit a large and selective decrease in AMPA-type glutamate receptor-mediated synaptic transmission without changes in NMDA-type glutamate receptor- or in GABAA receptor-mediated synaptic transmission. Thus, the synaptic phenotype observed in R704C-mutant iN cells replicates the previously observed phenotype of R704C-mutant neurons. Our data show that the effect of the R704C mutation is applicable even to neurons transdifferentiated from fibroblasts and constitute a proof-of-concept demonstration that iN cells can be used for cellular disease modeling. PMID:24046374

The Role of cGMP on Adenosine A1 Receptor-mediated Inhibition of Synaptic Transmission at the Hippocampus

PubMed Central

Pinto, Isa; Serpa, André; Sebastião, Ana M.; Cascalheira, José F.

2016-01-01

Both adenosine A1 receptor and cGMP inhibit synaptic transmission at the hippocampus and recently it was found that A1 receptor increased cGMP levels in hippocampus, but the role of cGMP on A1 receptor-mediated inhibition of synaptic transmission remains to be established. In the present work we investigated if blocking the NOS/sGC/cGMP/PKG pathway using nitric oxide synthase (NOS), protein kinase G (PKG), and soluble guanylyl cyclase (sGC) inhibitors modify the A1 receptor effect on synaptic transmission. Neurotransmission was evaluated by measuring the slope of field excitatory postsynaptic potentials (fEPSPs) evoked by electrical stimulation at hippocampal slices. N6-cyclopentyladenosine (CPA, 15 nM), a selective A1 receptor agonist, reversibly decreased the fEPSPs by 54 ± 5%. Incubation of the slices with an inhibitor of NOS (L-NAME, 200 μM) decreased the CPA effect on fEPSPs by 57 ± 9% in female rats. In males, ODQ (10 μM), an sGC inhibitor, decreased the CPA inhibitory effect on fEPSPs by 23 ± 6%, but only when adenosine deaminase (ADA,1 U/ml) was present; similar results were found in females, where ODQ decreased CPA-induced inhibition of fEPSP slope by 23 ± 7%. In male rats, the presence of the PKG inhibitor (KT5823, 1 nM) decreased the CPA effect by 45.0 ± 9%; similar results were obtained in females, where KT5823 caused a 32 ± 9% decrease on the CPA effect. In conclusion, the results suggest that the inhibitory action of adenosine A1 receptors on synaptic transmission at hippocampus is, in part, mediated by the NOS/sGC/cGMP/PKG pathway. PMID:27148059

Regulation of GABAA and Glutamate Receptor Expression, Synaptic Facilitation and Long-Term Potentiation in the Hippocampus of Prion Mutant Mice

PubMed Central

Rangel, Alejandra; Madroñal, Noelia; Massó, Agnès Gruart i.; Gavín, Rosalina; Llorens, Franc; Sumoy, Lauro; Torres, Juan María; Delgado-García, José María; Río, José Antonio Del

2009-01-01

Background Prionopathies are characterized by spongiform brain degeneration, myoclonia, dementia, and periodic electroencephalographic (EEG) disturbances. The hallmark of prioniopathies is the presence of an abnormal conformational isoform (PrPsc) of the natural cellular prion protein (PrPc) encoded by the Prnp gene. Although several roles have been attributed to PrPc, its putative functions in neuronal excitability are unknown. Although early studies of the behavior of Prnp knockout mice described minor changes, later studies report altered behavior. To date, most functional PrPc studies on synaptic plasticity have been performed in vitro. To our knowledge, only one electrophysiological study has been performed in vivo in anesthetized mice, by Curtis and coworkers. They reported no significant differences in paired-pulse facilitation or LTP in the CA1 region after Schaffer collateral/commissural pathway stimulation. Methodology/Principal Findings Here we explore the role of PrPc expression in neurotransmission and neural excitability using wild-type, Prnp −/− and PrPc-overexpressing mice (Tg20 strain). By correlating histopathology with electrophysiology in living behaving mice, we demonstrate that both Prnp −/− mice but, more relevantly Tg20 mice show increased susceptibility to KA, leading to significant cell death in the hippocampus. This finding correlates with enhanced synaptic facilitation in paired-pulse experiments and hippocampal LTP in living behaving mutant mice. Gene expression profiling using Illumina™ microarrays and Ingenuity pathways analysis showed that 129 genes involved in canonical pathways such as Ubiquitination or Neurotransmission were co-regulated in Prnp −/− and Tg20 mice. Lastly, RT-qPCR of neurotransmission-related genes indicated that subunits of GABAA and AMPA-kainate receptors are co-regulated in both Prnp −/− and Tg20 mice. Conclusions/Significance Present results demonstrate that PrPc is necessary for the proper

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

Effects of central histamine receptors blockade on GABA(A) agonist-induced food intake in broiler cockerels.

PubMed

Morteza, Zendehdel; Vahhab, Babapour; Hossein, Jonaidi

2008-02-01

In this study, the effect of intracerebroventricular (i.c.v) injection of H1, H2 and H3 antagonists on feed intake induced by GABA(A) agonist was evaluated. In Experiment 1, the animals received chloropheniramine, a H1 antagonist and then muscimol, a GABA(A) agonist. In Experiment 2, chickens received famotidine, a H2 receptor antagonist, prior to injection of muscimol. Finally in Experiment 3, the birds were injected with thioperamide, a H3 receptor antagonist and muscimol. Cumulative food intake was measured 15, 30, 45, 60, 90, 120, 150 and 180 min after injections. The results of this study indicated that effects of muscimol on food intake inhibited by pretreatment with chloropheneramine maleate (p < or = 0.05), significantly, while the famotidine and thioperamide were ineffective. These results suggest the existence of H1-receptor mediated histamine-GABA(A) receptor interaction on food intake in broiler cockerels.

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

The TM2 6′ Position of GABAA Receptors Mediates Alcohol Inhibition

PubMed Central

Howard, Rebecca J.; Trudell, James R.; Harris, R. Adron

2012-01-01

Ionotropic GABAA receptors (GABAARs), which mediate inhibitory neurotransmission in the central nervous system, are implicated in the behavioral effects of alcohol and alcoholism. Site-directed mutagenesis studies support the presence of discrete molecular sites involved in alcohol enhancement and, more recently, inhibition of GABAARs. We used Xenopus laevis oocytes to investigate the 6′ position in the second transmembrane region of GABAARs as a site influencing alcohol inhibition. We asked whether modification of the 6′ position by substitution with larger residues or methanethiol labeling [using methyl methanethiosulfonate (MMTS)] of a substituted cysteine, reduced GABA action and/or blocked further inhibition by alcohols. Labeling of the 6′ position in either α2 or β2 subunits reduced responses to GABA. In addition, methanol and ethanol potentiation increased after MMTS labeling or substitution with tryptophan or methionine, consistent with elimination of an inhibitory site for these alcohols. Specific alcohols, but not the anesthetic etomidate, competed with MMTS labeling at the 6′ position. We verified a role for the 6′ position in previously tested α2β2 as well as more physiologically relevant α2β2γ2s GABAARs. Finally, we built a novel molecular model based on the invertebrate glutamate-gated chloride channel receptor, a GABAAR homolog, revealing that the 6′ position residue faces the channel pore, and modification of this residue alters volume and polarity of the pore-facing cavity in this region. These results indicate that the 6′ positions in both α2 and β2 GABAAR subunits mediate inhibition by short-chain alcohols, which is consistent with the presence of multiple counteracting sites of action for alcohols on ligand-gated ion channels. PMID:22072732

Synaptically released zinc triggers metabotropic signaling via a zinc-sensing receptor in the hippocampus.

PubMed

Besser, Limor; Chorin, Ehud; Sekler, Israel; Silverman, William F; Atkin, Stan; Russell, James T; Hershfinkel, Michal

2009-03-04

Zn(2+) is coreleased with glutamate from mossy fiber terminals and can influence synaptic function. Here, we demonstrate that synaptically released Zn(2+) activates a selective postsynaptic Zn(2+)-sensing receptor (ZnR) in the CA3 region of the hippocampus. ZnR activation induced intracellular release of Ca(2+), as well as phosphorylation of extracellular-regulated kinase and Ca(2+)/calmodulin kinase II. Blockade of synaptic transmission by tetrodotoxin or CdCl inhibited the ZnR-mediated Ca(2+) rises. The responses mediated by ZnR were largely attenuated by the extracellular Zn(2+) chelator, CaEDTA, and in slices from mice lacking vesicular Zn(2+), suggesting that synaptically released Zn(2+) triggers the metabotropic activity. Knockdown of the expression of the orphan G-protein-coupled receptor 39 (GPR39) attenuated ZnR activity in a neuronal cell line. Importantly, we observed widespread GPR39 labeling in CA3 neurons, suggesting a role for this receptor in mediating ZnR signaling in the hippocampus. Our results describe a unique role for synaptic Zn(2+) acting as the physiological ligand of a metabotropic receptor and provide a novel pathway by which synaptic Zn(2+) can regulate neuronal function.

GABAA receptors involved in sleep and anaesthesia: β1- versus β3-containing assemblies.

PubMed

Yanovsky, Yevgenij; Schubring, Stephan; Fleischer, Wiebke; Gisselmann, Günter; Zhu, Xin-Ran; Lübbert, Hermann; Hatt, Hanns; Rudolph, Uwe; Haas, Helmut L; Sergeeva, Olga A

2012-01-01

The histaminergic neurons of the posterior hypothalamus (tuberomamillary nucleus-TMN) control wakefulness, and their silencing through activation of GABA(A) receptors (GABA(A)R) induces sleep and is thought to mediate sedation under propofol anaesthesia. We have previously shown that the β1 subunit preferring fragrant dioxane derivatives (FDD) are highly potent modulators of GABA(A)R in TMN neurons. In recombinant receptors containing the β3N265M subunit, FDD action is abolished and GABA potency is reduced. Using rat, wild-type and β3N265M mice, FDD and propofol, we explored the relative contributions of β1- and β3-containing GABA(A)R to synaptic transmission from the GABAergic sleep-on ventrolateral preoptic area neurons to TMN. In β3N265M mice, GABA potency remained unchanged in TMN neurons, but it was decreased in cultured posterior hypothalamic neurons with impaired modulation of GABA(A)R by propofol. Spontaneous and evoked GABAergic synaptic currents (IPSC) showed β1-type pharmacology, with the same effects achieved by 3 μM propofol and 10 μM PI24513. Propofol and the FDD PI24513 suppressed neuronal firing in the majority of neurons at 5 and 100 μM, and in all cells at 10 and 250 μM, respectively. FDD given systemically in mice induced sedation but not anaesthesia. Propofol-induced currents were abolished (1-6 μM) or significantly reduced (12 μM) in β3N265M mice, whereas gating and modulation of GABA(A)R by PI24513 as well as modulation by propofol were unchanged. In conclusion, β1-containing (FDD-sensitive) GABA(A)R represent the major receptor pool in TMN neurons responding to GABA, while β3-containing (FDD-insensitive) receptors are gated by low micromolar doses of propofol. Thus, sleep and anaesthesia depend on different GABA(A)R types.

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.

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

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.

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.

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

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.

Impact of single-site axonal GABAergic synaptic events on cerebellar interneuron activity.

PubMed

de San Martin, Javier Zorrilla; Jalil, Abdelali; Trigo, Federico F

2015-12-01

Axonal ionotropic receptors are present in a variety of neuronal types, and their function has largely been associated with the modulation of axonal activity and synaptic release. It is usually assumed that activation of axonal GABA(A)Rs comes from spillover, but in cerebellar molecular layer interneurons (MLIs) the GABA source is different: in these cells, GABA release activates presynaptic GABA(A) autoreceptors (autoRs) together with postsynaptic targets, producing an autoR-mediated synaptic event. The frequency of presynaptic, autoR-mediated miniature currents is twice that of their somatodendritic counterparts, suggesting that autoR-mediated responses have an important effect on interneuron activity. Here, we used local Ca(2+) photolysis in MLI axons of juvenile rats to evoke GABA release from individual varicosities to study the activation of axonal autoRs in single release sites. Our data show that single-site autoR conductances are similar to postsynaptic dendritic conductances. In conditions of high [Cl(-)](i), autoR-mediated conductances range from 1 to 5 nS; this corresponds to ∼30-150 GABA(A) channels per presynaptic varicosity, a value close to the number of channels in postsynaptic densities. Voltage responses produced by the activation of autoRs in single varicosities are amplified by a Na(v)-dependent mechanism and propagate along the axon with a length constant of 91 µm. Immunolabeling determination of synapse location shows that on average, one third of the synapses produce autoR-mediated signals that are large enough to reach the axon initial segment. Finally, we show that single-site activation of presynaptic GABA(A) autoRs leads to an increase in MLI excitability and thus conveys a strong feedback signal that contributes to spiking activity. © 2015 Zorrilla de San Martin et al.

Impact of single-site axonal GABAergic synaptic events on cerebellar interneuron activity

PubMed Central

Zorrilla de San Martin, Javier; Jalil, Abdelali

2015-01-01

Axonal ionotropic receptors are present in a variety of neuronal types, and their function has largely been associated with the modulation of axonal activity and synaptic release. It is usually assumed that activation of axonal GABAARs comes from spillover, but in cerebellar molecular layer interneurons (MLIs) the GABA source is different: in these cells, GABA release activates presynaptic GABAA autoreceptors (autoRs) together with postsynaptic targets, producing an autoR-mediated synaptic event. The frequency of presynaptic, autoR-mediated miniature currents is twice that of their somatodendritic counterparts, suggesting that autoR-mediated responses have an important effect on interneuron activity. Here, we used local Ca2+ photolysis in MLI axons of juvenile rats to evoke GABA release from individual varicosities to study the activation of axonal autoRs in single release sites. Our data show that single-site autoR conductances are similar to postsynaptic dendritic conductances. In conditions of high [Cl−]i, autoR-mediated conductances range from 1 to 5 nS; this corresponds to ∼30–150 GABAA channels per presynaptic varicosity, a value close to the number of channels in postsynaptic densities. Voltage responses produced by the activation of autoRs in single varicosities are amplified by a Nav-dependent mechanism and propagate along the axon with a length constant of 91 µm. Immunolabeling determination of synapse location shows that on average, one third of the synapses produce autoR-mediated signals that are large enough to reach the axon initial segment. Finally, we show that single-site activation of presynaptic GABAA autoRs leads to an increase in MLI excitability and thus conveys a strong feedback signal that contributes to spiking activity. PMID:26621773

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

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

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.

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

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.

Slow synaptic transmission mediated by TRPV1 channels in CA3 interneurons of the hippocampus.

PubMed

Eguchi, Noriomi; Hishimoto, Akitoyo; Sora, Ichiro; Mori, Masahiro

2016-03-11

Metabotropic glutamate receptors (mGluRs) modulate various neuronal functions in the central nervous system. Many studies reported that mGluRs have linkages to neuronal disorders such as schizophrenia and autism related disorders, indicating that mGluRs are involved in critical functions of the neuronal circuits. To study this possibility further, we recorded mGluR-induced synaptic responses in the interneurons of the CA3 stratum radiatum using rat hippocampal organotypic slice cultures. Electrical stimulation in the CA3 pyramidal cell layer evoked a slow inward current in the interneurons at a holding potential of -70mV in the presence of antagonists for AMPA/kainate receptors, NMDA receptors, GABAA receptors and GABAB receptors. The slow inward current was blocked in the absence of extracellular calcium, suggesting that this was a synaptic response. The slow excitatory postsynaptic current (EPSC) reversed near 0mV, reflecting an increase in a non-selective cationic conductance. The slow EPSC is mediated by group I mGluRs, as it was blocked by AP3, a group I mGluR antagonist. Neither a calcium chelator BAPTA nor a phospholipase C (PLC) inhibitor U73122 affected the slow EPSC. La(3+), a general TRP channel blocker or capsazepine, a selective TRPV1 channel antagonist significantly suppressed the slow EPSC. DHPG, a selective group I mGluRs agonist induced an inward current, which was suppressed by capsazepine. These results indicate that in the interneurons of the hippocampal CA3 stratum radiatum group I mGluRs activate TRPV1 channels independently of PLC and intracellular Ca(2+), resulting in the slow EPSC in the interneurons. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

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.

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.

Mapping General Anesthetic Sites in Heteromeric γ-Aminobutyric Acid Type A Receptors Reveals a Potential For Targeting Receptor Subtypes.

PubMed

Forman, Stuart A; Miller, Keith W

2016-11-01

IV general anesthetics, including propofol, etomidate, alphaxalone, and barbiturates, produce important actions by enhancing γ-aminobutyric acid type A (GABAA) receptor activation. In this article, we review scientific studies that have located and mapped IV anesthetic sites using photoaffinity labeling and substituted cysteine modification protection. These anesthetics bind in transmembrane pockets between subunits of typical synaptic GABAA receptors, and drugs that display stereoselectivity also show remarkably selective interactions with distinct interfacial sites. These results suggest strategies for developing new drugs that selectively modulate distinct GABAA receptor subtypes.

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

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.

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

Iron Mediates N-Methyl-d-aspartate Receptor-dependent Stimulation of Calcium-induced Pathways and Hippocampal Synaptic Plasticity*

PubMed Central

Muñoz, Pablo; Humeres, Alexis; Elgueta, Claudio; Kirkwood, Alfredo; Hidalgo, Cecilia; Núñez, Marco T.

2011-01-01

Iron deficiency hinders hippocampus-dependent learning processes and impairs cognitive performance, but current knowledge on the molecular mechanisms underlying the unique role of iron in neuronal function is sparse. Here, we investigated the participation of iron on calcium signal generation and ERK1/2 stimulation induced by the glutamate agonist N-methyl-d-aspartate (NMDA), and the effects of iron addition/chelation on hippocampal basal synaptic transmission and long-term potentiation (LTP). Addition of NMDA to primary hippocampal cultures elicited persistent calcium signals that required functional NMDA receptors and were independent of calcium influx through L-type calcium channels or α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors; NMDA also promoted ERK1/2 phosphorylation and nuclear translocation. Iron chelation with desferrioxamine or inhibition of ryanodine receptor (RyR)-mediated calcium release with ryanodine-reduced calcium signal duration and prevented NMDA-induced ERK1/2 activation. Iron addition to hippocampal neurons readily increased the intracellular labile iron pool and stimulated reactive oxygen species production; the antioxidant N-acetylcysteine or the hydroxyl radical trapper MCI-186 prevented these responses. Iron addition to primary hippocampal cultures kept in calcium-free medium elicited calcium signals and stimulated ERK1/2 phosphorylation; RyR inhibition abolished these effects. Iron chelation decreased basal synaptic transmission in hippocampal slices, inhibited iron-induced synaptic stimulation, and impaired sustained LTP in hippocampal CA1 neurons induced by strong stimulation. In contrast, iron addition facilitated sustained LTP induction after suboptimal tetanic stimulation. Together, these results suggest that hippocampal neurons require iron to generate RyR-mediated calcium signals after NMDA receptor stimulation, which in turn promotes ERK1/2 activation, an essential step of sustained LTP. PMID:21296883

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.

Ionotropic and metabotropic glutamate receptor mediation of glucocorticoid-induced apoptosis in hippocampal cells and the neuroprotective role of synaptic N-methyl-D-aspartate receptors.

PubMed

Lu, J; Goula, D; Sousa, N; Almeida, O F X

2003-01-01

Glutamate receptors have been proposed to mediate the apoptotic actions of glucocorticoids in hippocampal cells. To further analyze the role of glutamate receptors in this process, we pretreated primary hippocampal cells from neonatal (postnatal day 4) rats with antagonists of ionotropic glutamate receptor (iGluR) and metabotropic glutamate receptor (mGluR) antagonists before exposure to the specific glucocorticoid receptor agonist dexamethasone (DEX) at a dose of 1 microM. Dizocilpine (MK801; a general N-methyl-D-aspartic acid [NMDA] receptor antagonist, NMDAR antagonist) and ifenprodil (a specific ligand of the NMDAR 2B subunit, NR2B), were used to block iGluR; (RS)-alpha-ethyl-4-carboxyphenylglycine (E4CPG) and (RS)-alpha-cyclopropyl-4-phosphonophenyl-glycine (CPPG) were employed as I/II (E4CPG) and II/III (CPPG) mGluR antagonists. Blockade of iGluR resulted in a significant attenuation of DEX-induced cell death; the finding that ifenprodil exerted a similar potency to MK801 demonstrates the involvement of NR2B receptors in glucocorticoid-induced cell death. Apoptosis accounted for a significant amount of the cell loss observed, as detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling histochemistry for the in situ labeling of DNA breaks; apoptotic cells were distinguished from necrosis on the basis of morphological criteria, including chromatin condensation, membrane blebbing and presence of apoptotic bodies. Treatment with E4CPG and CPPG completely abolished the apoptotic response to DEX, thus showing the additional contribution of mGluR to the phenomenon. Further, dose-response studies with NMDA revealed that whereas high (10 microM) doses of NMDA themselves elicit cytotoxic responses, low (1-5 microM) concentrations of NMDA can effectively oppose DEX-induced cell death. Interestingly, the neuroprotective actions of low dose NMDA stimulation were abolished when either synaptic or extrasynaptic NMDA receptors were blocked with MK801

Postsynaptic Synaptotagmins Mediate AMPA Receptor Exocytosis During LTP

PubMed Central

Wu, Dick; Bacaj, Taulant; Morishita, Wade; Goswami, Debanjan; Arendt, Kristin L.; Xu, Wei; Chen, Lu; Malenka, Robert C.; Südhof, Thomas C.

2017-01-01

Strengthening of synaptic connections by NMDA-receptor-dependent long-term potentiation (LTP) shapes neural circuits and mediates learning and memory. During NMDA-receptor-dependent LTP induction, Ca2+-influx stimulates recruitment of synaptic AMPA-receptors, thereby strengthening synapses. How Ca2+ induces AMPA-receptor recruitment, however, remains unclear. Here we show that, in pyramidal neurons of the hippocampal CA1-region, blocking postsynaptic expression of both synaptotagmin-1 and synaptotagmin-7, but not of synaptotagmin-1 or synaptotagmin-7 alone, abolished LTP. LTP was rescued by wild-type but not by Ca2+-binding-deficient mutant synaptotagmin-7. Blocking postsynaptic synaptotagmin-1/7 expression did not impair basal synaptic transmission, synaptic or extrasynaptic AMPA-receptor levels, or other AMPA-receptor trafficking events. Moreover, expression of dominant-negative mutant synaptotagmin-1 that inhibited Ca2+-dependent presynaptic vesicle exocytosis also blocked Ca2+-dependent postsynaptic AMPA-receptor exocytosis, thereby abolishing LTP. Our results suggest that postsynaptic synaptotagmin-1 and synaptotagmin-7 act as redundant Ca2+-sensors for Ca2+-dependent exocytosis of AMPA-receptors during LTP, thus delineating a simple mechanism for the recruitment of AMPA-receptors that mediates LTP. PMID:28355182

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

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

The influence of stress at puberty on mood and learning: Role of the α4βδ GABAA receptor

PubMed Central

Smith, Sheryl S.

2012-01-01

It is well-known that the onset of puberty is associated with changes in mood as well as cognition. Stress can have an impact on these outcomes, which in many cases, can be more influential in females, suggesting that gender differences exist. The adolescent period is a vulnerable time for the onset of certain psychopathologies, including anxiety disorders, depression and eating disorders, which are also more prevalent in females. One factor which may contribute to stress-triggered anxiety at puberty is the GABAA receptor (GABAR), which is known to play a pivotal role in anxiety. Expression of α4βδ GABARs increases on the dendrites of CA1 pyramidal cells at the onset of puberty in the hippocampus, part of the limbic circuitry which governs emotion. This receptor is a sensitive target for the stress steroid THP (3α-OH-5[α]β-pregnan-20-one), which paradoxically reduces inhibition and increases anxiety during the pubertal period (~PND 35–44) of female mice in contrast to its usual effect to enhance inhibition and reduce anxiety. Spatial learning and synaptic plasticity are also adversely impacted at puberty, likely a result of increased expression of α4βδ GABARs on the dendritic spines of CA1 hippocampal pyramidal cells, which are essential for consolidation of memory. This review will focus on the role of these receptors in mediating behavioral changes at puberty. Stress-mediated changes in mood and cognition in early adolescence may have relevance for the expression of psychopathologies in adulthood. PMID:23079628

Alpha-1A Adrenergic receptor activation increases inhibitory tone in CA1 hippocampus

PubMed Central

Hillman, Kristin L.; Lei, Saobo; Doze, Van A.

2009-01-01

The endogenous catecholamine norepinephrine (NE) exhibits anti-epileptic properties, however it is not well understood which adrenergic receptor (AR) mediates this effect. The aim of this study was to investigate α1-adrenergic receptor (AR) activation in region CA1 of the hippocampus, a subcortical structure often implicated in temporal lobe epilepsies. Using cell-attached and whole-cell recordings in rat hippocampal slices, we confirmed that selective α1-AR activation increases action potential firing in a subpopulation of CA1 interneurons. We found that this response is mediated via the α1A-AR subtype, initiated by sodium influx, and appears independent of second messenger signaling. In CA1 pyramidal cells, α1A-AR activation decreases activity due to increased pre-synaptic GABA and somatostatin release. Examination of post-synaptic receptor involvement revealed that while GABAA receptors mediate the majority of α1A-adrenergic effects on CA1 pyramidal cells, significant contributions are also made by GABAB and somatostatin receptors. Finally, to test whether α1A-AR activation could have potential therapeutic implications, we performed AR agonist challenges using two in vitro epileptiform models. When GABAA receptors were available, α1A-AR activation significantly decreased epileptiform bursting in CA1. Together, our findings directly link stimulation of the α1A-AR subtype to release of GABA and somatostatin at the single cell level and suggest that α1A-AR activation may represent one mechanism by which NE exerts anti-epileptic effects within the hippocampus. PMID:19201164

Increased regional cerebral blood flow but normal distribution of GABAA receptor in the visual cortex of subjects with early-onset blindness.

PubMed

Mishina, Masahiro; Senda, Michio; Kiyosawa, Motohiro; Ishiwata, Kiichi; De Volder, Anne G; Nakano, Hideki; Toyama, Hinako; Oda, Kei-ichi; Kimura, Yuichi; Ishii, Kenji; Sasaki, Touru; Ohyama, Masashi; Komaba, Yuichi; Kobayashi, Shirou; Kitamura, Shin; Katayama, Yasuo

2003-05-01

Before the completion of visual development, visual deprivation impairs synaptic elimination in the visual cortex. The purpose of this study was to determine whether the distribution of central benzodiazepine receptor (BZR) is also altered in the visual cortex in subjects with early-onset blindness. Positron emission tomography was carried out with [(15)O]water and [(11)C]flumazenil on six blind subjects and seven sighted controls at rest. We found that the CBF was significantly higher in the visual cortex for the early-onset blind subjects than for the sighted control subjects. However, there was no significant difference in the BZR distribution in the visual cortex for the subject with early-onset blindness than for the sighted control subjects. These results demonstrated that early visual deprivation does not affect the distribution of GABA(A) receptors in the visual cortex with the sensitivity of our measurements. Synaptic elimination may be independent of visual experience in the GABAergic system of the human visual cortex during visual development.

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.

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.

Furosemide suppresses ileal and colonic contractility via interactions with GABA-A receptor in mice.

PubMed

Kaewsaro, Kannaree; Nualplub, Suparp; Bumrungsri, Sara; Khuituan, Pissared

2017-11-01

The loop diuretic furosemide has an action to inhibit Na + -K + -2Cl - co-transporter at the thick ascending limb of Henle's loop resulting in diuresis. Furosemide also has the non-diuretic effects by binding to GABA-A receptor which may involve the gastrointestinal tract. The aim of this study was to investigate the effects of furosemide on smooth muscle contractions in mice ileum and proximal colon. Each intestinal segment suspended in an organ bath was connected to a force transducer. Signal output of mechanical activity was amplified and recorded for analysis using PowerLab System. After equilibration, the intestine was directly exposed to furosemide, GABA, GABA-A receptor agonist (muscimol), or muscarinic receptor antagonist (atropine). Furosemide (50, 100 and 500 μmol L -1 ) acutely reduced the amplitude of ileal and colonic contraction. In the ileum, 1 mmol L -1 GABA and 10-60 μmol L -1 muscimol significantly increased the amplitude, whereas in the colon, 50-100 mmol L -1 GABA and 60 μmol L -1 muscimol decreased the contractions. The contractions were also significantly suppressed by atropine. To investigate the mechanisms underlying the inhibiting effect of furosemide, furosemide was added to the organ bath prior to the addition of muscimol or atropine. A comparison of furosemide combined with muscimol or atropine group and furosemide group showed no significant difference of the ileal contraction, but the amplitude of colonic contraction significantly decreased when compared to adding furosemide alone. These results suggest that furosemide can reduce the ileal and proximal colonic contraction mediated by blocking and supporting of GABA-A receptor, respectively, resulting in decreased acetylcholine release. © 2017 John Wiley & Sons Australia, Ltd.

Acutely increasing δGABA(A) receptor activity impairs memory and inhibits synaptic plasticity in the hippocampus.

PubMed

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 (GABA(A)) receptors that contain the δ subunit (δGABA(A) receptors) are expressed in several brain regions including the dentate gyrus (DG) and CA1 subfields of the hippocampus. Drugs that increase δGABA(A) receptor activity have been proposed as treatments for a variety of disorders including insomnia, epilepsy and chronic pain. Also, long-term pretreatment with the δGABA(A) 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 δGABA(A) 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 δGABA(A) 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 GABA(A) receptor antagonist bicuculline. Thus, acutely increasing δGABA(A) receptor activity impairs memory behaviors and inhibits synaptic plasticity. These results have important implications for the development of therapies aimed at increasing δGABA(A) receptor activity.

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

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

Auditory Thalamic Circuits and GABAA Receptor Function: Putative Mechanisms in Tinnitus Pathology

PubMed Central

Caspary, Donald M.; Llano, Daniel A

2016-01-01

Tinnitus is defined as a phantom sound (ringing in the ears), and can significantly reduce the quality of life for those who suffer its effects. Ten to fifteen percent of the general adult population report symptoms of tinnitus with 1-2% reporting that tinnitus negatively impacts their quality of life. Noise exposure is the most common cause of tinnitus and the military environment presents many challenging high-noise situations. Military noise levels can be so intense that standard hearing protection is not adequate. Recent studies suggest a role for inhibitory neurotransmitter dysfunction in response to noise-induced peripheral deafferentation as a key element in the pathology of tinnitus. The auditory thalamus, or medial geniculate body (MGB), is an obligate auditory brain center in a unique position to gate the percept of sound as it projects to auditory cortex and to limbic structures. Both areas are thought to be involved in those individuals most impacted by tinnitus. For MGB, opposing hypotheses have posited either a tinnitus-related pathologic decrease or pathologic increase in GABAergic inhibition. In sensory thalamus, GABA mediates fast synaptic inhibition via synaptic GABAA receptors (GABAARs) as well as a persistent tonic inhibition via high-affinity extrasynaptic GABAARs and slow synaptic inhibition via GABABRs. Down-regulation of inhibitory neurotransmission, related to partial peripheral deafferentation, is consistently presented as partially underpinning neuronal hyperactivity seen in animal models of tinnitus. This maladaptive plasticity/Gain Control Theory of tinnitus pathology (see Auerbach et al., 2014; Richardson et al., 2012) is characterized by reduced inhibition associated with increased spontaneous and abnormal neuronal activity, including bursting and increased synchrony throughout much of the central auditory pathway. A competing hypothesis suggests that maladaptive oscillations between the MGB and auditory cortex, thalamocortical

Insulin Regulates GABAA Receptor-Mediated Tonic Currents in the Prefrontal Cortex.

PubMed

Trujeque-Ramos, Saraí; Castillo-Rolón, Diego; Galarraga, Elvira; Tapia, Dagoberto; Arenas-López, Gabina; Mihailescu, Stefan; Hernández-López, Salvador

2018-01-01

Recent studies, have shown that insulin increases extrasynaptic GABA A receptor-mediated currents in the hippocampus, causing alterations of neuronal excitability. The prefrontal cortex (PFC) is another brain area which is involved in cognition functions and expresses insulin receptors. Here, we used electrophysiological, molecular, and immunocytochemical techniques to examine the effect of insulin on the extrasynaptic GABA A receptor-mediated tonic currents in brain slices. We found that insulin (20-500 nM) increases GABA A -mediated tonic currents. Our results suggest that insulin promotes the trafficking of extrasynaptic GABA A receptors from the cytoplasm to the cell membrane. Western blot analysis and immunocytochemistry showed that PFC extrasynaptic GABA A receptors contain α-5 and δ subunits. Insulin effect on tonic currents decreased the firing rate and neuronal excitability in layer 5-6 PFC cells. These effects of insulin were dependent on the activation of the PI3K enzyme, a key mediator of the insulin response within the brain. Taken together, these results suggest that insulin modulation of the GABA A -mediated tonic currents can modify the activity of neural circuits within the PFC. These actions could help to explain the alterations of cognitive processes associated with changes in insulin signaling.

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.

Activation of α7 nicotinic acetylcholine receptors persistently enhances hippocampal synaptic transmission and prevents Aß-mediated inhibition of LTP in the rat hippocampus.

PubMed

Ondrejcak, Tomas; Wang, Qinwen; Kew, James N C; Virley, David J; Upton, Neil; Anwyl, Roger; Rowan, Michael J

2012-02-29

Nicotinic acetylcholine receptors mediate fast cholinergic modulation of glutamatergic transmission and synaptic plasticity. Here we investigated the effects of subtype selective activation of the α7 nicotinic acetylcholine receptors on hippocampal transmission and the inhibition of synaptic long-term potentiation by the Alzheimer's disease associated amyloid ß-protein (Aß). The α7 nicotinic acetylcholine receptor agonist "compound A" ((R)-N-(1-azabicyclo[2.2.2]oct-3-yl)(5-(2-pyridyl))thiophene-2-carboxamide) induced a rapid-onset persistent enhancement of synaptic transmission in the dentate gyrus in vitro. Consistent with a requirement for activation of α7 nicotinic acetylcholine receptors, the type II α7-selective positive allosteric modulator PheTQS ((3aR, 4S, 9bS)-4-(4-methylphenyl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinoline-8-sulfonamide) potentiated, and the antagonist methyllycaconitine (MLA) prevented the persistent enhancement. Systemic injection of the agonist also induced a similar MLA-sensitive persistent enhancement of synaptic transmission in the CA1 area in vivo. Remarkably, although compound A did not affect control long-term potentiation (LTP) in vitro, it prevented the inhibition of LTP by Aß1-42 and this effect was inhibited by MLA. These findings strongly indicate that activation of α7 nicotinic acetylcholine receptors is sufficient to persistently enhance hippocampal synaptic transmission and to overcome the inhibition of LTP by Aß. Copyright © 2011 Elsevier B.V. All rights reserved.

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.

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

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

PubMed Central

Moldavan, Mykhaylo G; Allen, Charles N

2013-01-01

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

Correlating Fluorescence and High-Resolution Scanning Electron Microscopy (HRSEM) for the study of GABAA receptor clustering induced by inhibitory synaptic plasticity.

PubMed

Orlando, Marta; Ravasenga, Tiziana; Petrini, Enrica Maria; Falqui, Andrea; Marotta, Roberto; Barberis, Andrea

2017-10-23

Both excitatory and inhibitory synaptic contacts display activity dependent dynamic changes in their efficacy that are globally termed synaptic plasticity. Although the molecular mechanisms underlying glutamatergic synaptic plasticity have been extensively investigated and described, those responsible for inhibitory synaptic plasticity are only beginning to be unveiled. In this framework, the ultrastructural changes of the inhibitory synapses during plasticity have been poorly investigated. Here we combined confocal fluorescence microscopy (CFM) with high resolution scanning electron microscopy (HRSEM) to characterize the fine structural rearrangements of post-synaptic GABA A Receptors (GABA A Rs) at the nanometric scale during the induction of inhibitory long-term potentiation (iLTP). Additional electron tomography (ET) experiments on immunolabelled hippocampal neurons allowed the visualization of synaptic contacts and confirmed the reorganization of post-synaptic GABA A R clusters in response to chemical iLTP inducing protocol. Altogether, these approaches revealed that, following the induction of inhibitory synaptic potentiation, GABA A R clusters increase in size and number at the post-synaptic membrane with no other major structural changes of the pre- and post-synaptic elements.

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

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.

Formation and stability of synaptic receptor domains.

PubMed

Haselwandter, Christoph A; Calamai, Martino; Kardar, Mehran; Triller, Antoine; da Silveira, Rava Azeredo

2011-06-10

Neurotransmitter receptor molecules, concentrated in postsynaptic domains along with scaffold and a number of other molecules, are key regulators of signal transmission across synapses. Combining experiment and theory, we develop a quantitative description of synaptic receptor domains in terms of a reaction-diffusion model. We show that interactions between only receptors and scaffolds, together with the rapid diffusion of receptors on the cell membrane, are sufficient for the formation and stable characteristic size of synaptic receptor domains. Our work reconciles long-term stability of synaptic receptor domains with rapid turnover and diffusion of individual receptors, and suggests novel mechanisms for a form of short-term, postsynaptic plasticity.

Decreased agonist sensitivity of human GABA(A) receptors by an amino acid variant, isoleucine to valine, in the alpha1 subunit.

PubMed

Westh-Hansen, S E; Rasmussen, P B; Hastrup, S; Nabekura, J; Noguchi, K; Akaike, N; Witt, M R; Nielsen, M

1997-06-25

Recombinant human GABA(A) receptors were investigated in vitro by coexpression of cDNAs coding for alpha1, beta2, and gamma2 subunits in the baculovirus/Sf-9 insect cell system. We report that a single amino acid exchange (isoleucine 121 to valine 121) in the N-terminal, extracellular part of the alpha1 subunit induces a marked decrease in agonist GABA(A) receptor ligand sensitivity. The potency of muscimol and GABA to inhibit the binding of the GABA(A) receptor antagonist [3H]SR 95531 (2-(3-carboxypropyl)-3-amino-6-(4-methoxyphenyl)pyridazinium bromide) was higher in receptor complexes of alpha1(ile 121) beta2gamma2 than in those of alpha1(val 121) beta2gamma2 (IC50 values were 32-fold and 26-fold lower for muscimol and GABA, respectively). The apparent affinity of the GABA(A) receptor antagonist bicuculline methiodide to inhibit the binding of [3H]SR 95531 did not differ between the two receptor complex variants. Electrophysiological measurements of GABA induced whole-cell Cl- currents showed a ten-fold decrease in the GABA(A) receptor sensitivity of alpha1 (val 121) beta2gamma2 as compared to alpha1(ile 121) beta2gamma2 receptor complexes. Thus, a relatively small change in the primary structure of the alpha1 subunit leads to a decrease selective for GABA(A) receptor sensitivity to agonist ligands, since no changes were observed in a GABA(A) receptor antagonist affinity and benzodiazepine receptor binding.

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.

Hippocampal GABAA Receptor and Pain Sensitivity during Estrous Cycle in the Rat

PubMed Central

Taherianfard, Mahnaz; Mosavi, Mahnaz

2011-01-01

Background: Estradiol and progesterone as well as hippocampal GABAA receptors are believed to play a role in the modulation of pain. The aim of present study was to investigate the effect of intrahippocampal injections of GABAA receptor agonist (muscimol) and GABAA receptor antagonist (picrotoxin) on pain sensitivity during estrous cycle. Methods: Pain sensitivity was evaluated in rats by formalin test during all stages of estrous cycle. Animals were divided into five groups including; 1- control (intact animal); 2- sham 1 receiving 0.75 µl artificial cerebrospinal fluids (ACSF); 3- sham 2 receiving 0.75 µl alcoholic ACSF; 4- experimental 1 receiving 250 or 500 µg/rat of muscimol in 0.75 µl vehicle, and 5- experimental 2 receiving 20 or 30 µg/rat picrotoxin in 0.75 µl vehicle. Data were analyzed by Kruskal-Wallis followed by Tucky's test for pairwise comparisons using a P value of ≤0.50 for statistical significance. Results: Muscimol significantly (P<0.05) decreased pain sensitivity in all stages of estrous cycle, and the analgesic effect was higher during proestrus and estrus stages of estrous cycle than that during metestrus and diestrus stages. Picrotoxin significantly (P<0.05) increased pain sensitivity in all stages of estrous cycle, and such a hyperalgesic effect was lower during proestrus and estrus stages of estrous cycle than that during metestrus and diestrus stages. Conclusion: The findings of the present study indicate that the role of hippocampal GABAA receptor in the control of the pain sensitivity can be modulated by variation in gonadal steroids during different stages of the estrous cycle. PMID:23115414

Central role for NMDA receptors in redox mediated impairment of synaptic function during aging and Alzheimer's disease.

PubMed

Foster, T C; Kyritsopoulos, C; Kumar, A

2017-03-30

Increased human longevity has magnified the negative impact that aging can have on cognitive integrity of older individuals experiencing some decline in cognitive function. Approximately 30% of the elderly will have cognitive problems that influence their independence. Impaired executive function and memory performance are observed in normal aging and yet can be an early sign of a progressive cognitive impairment of Alzheimer's disease (AD), the most common form of dementia. Brain regions that are vulnerable to aging exhibit the earliest pathology of AD. Senescent synaptic function is observed as a shift in Ca 2+ -dependent synaptic plasticity and similar mechanisms are thought to contribute to the early cognitive deficits associated with AD. In the case of aging, intracellular redox state mediates a shift in Ca 2+ regulation including N-methyl-d-aspartate (NMDA) receptor hypofunction and increased Ca 2+ release from intracellular stores to alter synaptic plasticity. AD can interact with these aging processes such that molecules linked to AD, β-amyloid (Aβ) and mutated presenilin 1 (PS1), can also degrade NMDA receptor function, promote Ca 2+ release from intracellular stores, and may increase oxidative stress. Thus, age is one of the most important predictors of AD and brain aging likely contributes to the onset of AD. The focus of this review article is to provide an update on mechanisms that contribute to the senescent synapse and possible interactions with AD-related molecules, with special emphasis on regulation of NMDA receptors. Copyright © 2016 Elsevier B.V. All rights reserved.

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

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

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.

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

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

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

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.

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

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.

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

MPTP-meditated hippocampal dopamine deprivation modulates synaptic transmission and activity-dependent synaptic plasticity

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

Zhu Guoqi; Chen Ying; Huang Yuying

2011-08-01

Parkinson's disease (PD)-like symptoms including learning deficits are inducible by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Therefore, it is possible that MPTP may disturb hippocampal memory processing by modulation of dopamine (DA)- and activity-dependent synaptic plasticity. We demonstrate here that intraperitoneal (i.p.) MPTP injection reduces the number of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra (SN) within 7 days. Subsequently, the TH expression level in SN and hippocampus and the amount of DA and its metabolite DOPAC in striatum and hippocampus decrease. DA depletion does not alter basal synaptic transmission and changes pair-pulse facilitation (PPF) of field excitatory postsynaptic potentials (fEPSPs) only atmore » the 30 ms inter-pulse interval. In addition, the induction of long-term potentiation (LTP) is impaired whereas the duration of long-term depression (LTD) becomes prolonged. Since both LTP and LTD depend critically on activation of NMDA and DA receptors, we also tested the effect of DA depletion on NMDA receptor-mediated synaptic transmission. Seven days after MPTP injection, the NMDA receptor-mediated fEPSPs are decreased by about 23%. Blocking the NMDA receptor-mediated fEPSP does not mimic the MPTP-LTP. Only co-application of D1/D5 and NMDA receptor antagonists during tetanization resembled the time course of fEPSP potentiation as observed 7 days after i.p. MPTP injection. Together, our data demonstrate that MPTP-induced degeneration of DA neurons and the subsequent hippocampal DA depletion alter NMDA receptor-mediated synaptic transmission and activity-dependent synaptic plasticity. - Highlights: > I.p. MPTP-injection mediates death of dopaminergic neurons. > I.p. MPTP-injection depletes DA and DOPAC in striatum and hippocampus. > I.p. MPTP-injection does not alter basal synaptic transmission. > Reduction of LTP and enhancement of LTD after i.p. MPTP-injection. > Attenuation of NMDA-receptors mediated f

Synaptic transmission at functionally identified synapses in the enteric nervous system: roles for both ionotropic and metabotropic receptors.

PubMed

Gwynne, R M; Bornstein, J C

2007-03-01

Digestion and absorption of nutrients and the secretion and reabsorption of fluid in the gastrointestinal tract are regulated by neurons of the enteric nervous system (ENS), the extensive peripheral nerve network contained within the intestinal wall. The ENS is an important physiological model for the study of neural networks since it is both complex and accessible. At least 20 different neurochemically and functionally distinct classes of enteric neurons have been identified in the guinea pig ileum. These neurons express a wide range of ionotropic and metabotropic receptors. Synaptic potentials mediated by ionotropic receptors such as the nicotinic acetylcholine receptor, P2X purinoceptors and 5-HT(3) receptors are seen in many enteric neurons. However, prominent synaptic potentials mediated by metabotropic receptors, like the P2Y(1) receptor and the NK(1) receptor, are also seen in these neurons. Studies of synaptic transmission between the different neuron classes within the enteric neural pathways have shown that both ionotropic and metabotropic synaptic potentials play major roles at distinct synapses within simple reflex pathways. However, there are still functional synapses at which no known transmitter or receptor has been identified. This review describes the identified roles for both ionotropic and metabotropic neurotransmission at functionally defined synapses within the guinea pig ileum ENS. It is concluded that metabotropic synaptic potentials act as primary transmitters at some synapses. It is suggested identification of the interactions between different synaptic potentials in the production of complex behaviours will require the use of well validated computer models of the enteric neural circuitry.

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.

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

Nootropic agents enhance the recruitment of fast GABAA inhibition in rat neocortex.

PubMed

Ling, Douglas S F; Benardo, Larry S

2005-07-01

It is widely believed that nootropic (cognition-enhancing) agents produce their therapeutic effects by augmenting excitatory synaptic transmission in cortical circuits, primarily through positive modulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate receptors (AMPARs). However, GABA-mediated inhibition is also critical for cognition, and enhanced GABA function may be likewise therapeutic for cognitive disorders. Could nootropics act through such a mechanism as well? To address this question, we examined the effects of nootropic agents on excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) recorded from layer V pyramidal cells in acute slices of somatosensory cortex. Aniracetam, a positive modulator of AMPA/kainate receptors, increased the peak amplitude of evoked EPSCs and the amplitude and duration of polysynaptic fast IPSCs, manifested as a greater total charge carried by IPSCs. As a result, the EPSC/IPSC ratio of total charge was decreased, representing a shift in the excitation-inhibition balance that favors inhibition. Aniracetam did not affect the magnitude of either monosynaptic IPSCs (mono-IPSCs) recorded in the presence of excitatory amino acid receptor antagonists, or miniature IPSCs (mIPSCs) recorded in the presence of tetrodotoxin. However, the duration of both mono-IPSCs and mIPSCs was prolonged, suggesting that aniracetam also directly modulates GABAergic transmission. Cyclothiazide, a preferential modulator of AMPAR function, enhanced the magnitude and duration of polysynaptic IPSCs, similar to aniracetam, but did not affect mono-IPSCs. Concanavalin A, a kainate receptor modulator, had little effect on EPSCs or IPSCs, suggesting there was no contribution from kainate receptor activity. These findings indicate that AMPAR modulators strengthen inhibition in neocortical pyramidal cells, most likely by altering the kinetics of AMPARs on synaptically connected interneurons and possibly by modulating GABA(A) receptor responses

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

Cortical synaptic NMDA receptor deficits in α7 nicotinic acetylcholine receptor gene deletion models: Implications for neuropsychiatric diseases

PubMed Central

Lin, Hong; Hsu, Fu-Chun; Baumann, Bailey H.; Coulter, Douglas A.; Lynch, David R.

2014-01-01

Microdeletion of the human CHRNA7 gene (α7 nicotinic acetylcholine receptor, nAChR) as well as dysfunction in N-methyl-D-aspartate receptors (NMDARs) have been associated with cortical dysfunction in a broad spectrum of neurodevelopmental and neuropsychiatric disorders including schizophrenia. However, the pathophysiological roles of synaptic vs. extrasynaptic NMDARs and their interactions with α7 nAChRs in cortical dysfunction remain largely uncharacterized. Using a combination of in vivo and in vitro models, we demonstrate that α7 nAChR gene deletion leads to specific loss of synaptic NMDARs and their coagonist, D-serine, as well as glutamatergic synaptic deficits in mouse cortex. α7 nAChR null mice had decreased cortical NMDAR expression and glutamatergic synapse formation during postnatal development. Similar reductions in NMDAR expression and glutamatergic synapse formation were revealed in cortical cultures lacking α7 nAChRs. Interestingly, synaptic, but not extrasynaptic, NMDAR currents were specifically diminished in cultured cortical pyramidal neurons as well as in acute prefrontal cortical slices of α7 nAChR null mice. Moreover, D-serine responsive synaptic NMDAR-mediated currents and levels of the D-serine synthetic enzyme serine racemase were both reduced in α7 nAChR null cortical pyramidal neurons. Our findings thus identify specific loss of synaptic NMDARs and their coagonist, D-serine, as well as glutamatergic synaptic deficits in α7 nAChR gene deletion models of cortical dysfunction, thereby implicating α7 nAChR-mediated control of synaptic NMDARs and serine racemase/D-serine pathways in cortical dysfunction underlying many neuropsychiatric and neurodevelopmental disorders, particularly those associated with deletion of human CHRNA7. PMID:24326163

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

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.

Autoantibodies to Synaptic Receptors and Neuronal Cell Surface Proteins in Autoimmune Diseases of the Central Nervous System

PubMed Central

Geis, Christian; Graus, Francesc

2017-01-01

Investigations in the last 10 years have revealed a new category of neurological diseases mediated by antibodies against cell surface and synaptic proteins. There are currently 16 such diseases all characterized by autoantibodies against neuronal proteins involved in synaptic signaling and plasticity. In clinical practice these findings have changed the diagnostic and treatment approach to potentially lethal, but now treatable, neurological and psychiatric syndromes previously considered idiopathic or not even suspected to be immune-mediated. Studies show that patients' antibodies can impair the surface dynamics of the target receptors eliminating them from synapses (e.g., NMDA receptor), block the function of the antigens without changing their synaptic density (e.g., GABAb receptor), interfere with synaptic protein-protein interactions (LGI1, Caspr2), alter synapse formation (e.g., neurexin-3α), or by unclear mechanisms associate to a new form of tauopathy (IgLON5). Here we first trace the process of discovery of these diseases, describing the triggers and symptoms related to each autoantigen, and then review in detail the structural and functional alterations caused by the autoantibodies with special emphasis in those (NMDA receptor, amphiphysin) that have been modeled in animals. PMID:28298428

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.

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

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...

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

Insulin-Independent GABAA Receptor-Mediated Response in the Barrel Cortex of Mice with Impaired Met Activity.

PubMed

Lo, Fu-Sun; Erzurumlu, Reha S; Powell, Elizabeth M

2016-03-30

Autism spectrum disorder (ASD) is a neurodevelopmental disorder caused by genetic variants, susceptibility alleles, and environmental perturbations. The autism associated geneMETtyrosine 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 GABAAreceptor-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 GABAAreceptor-mediated response in the barrel cortex of mice with compromised Met signaling. Thus, lacking insulin effects may be a risk factor in ASD pathogenesis. 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 GABAAreceptors 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. Copyright © 2016 the authors 0270-6474/16/363691-07$15.00/0.

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.

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 desensitization gate of inhibitory Cys-loop receptors

NASA Astrophysics Data System (ADS)

Gielen, Marc; Thomas, Philip; Smart, Trevor G.

2015-04-01

Cys-loop neurotransmitter-gated ion channels are vital for communication throughout the nervous system. Following activation, these receptors enter into a desensitized state in which the ion channel shuts even though the neurotransmitter molecules remain bound. To date, the molecular determinants underlying this most fundamental property of Cys-loop receptors have remained elusive. Here we present a generic mechanism for the desensitization of Cys-loop GABAA (GABAARs) and glycine receptors (GlyRs), which both mediate fast inhibitory synaptic transmission. Desensitization is regulated by interactions between the second and third transmembrane segments, which affect the ion channel lumen near its intracellular end. The GABAAR and GlyR pore blocker picrotoxin prevented desensitization, consistent with its deep channel-binding site overlapping a physical desensitization gate.

PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

PubMed

Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

2015-02-01

The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

Diversity in GABAergic signaling.

PubMed

Vogt, Kaspar

2015-01-01

GABA(A) receptor-mediated synaptic transmission is responsible for inhibitory control of neural function in the brain. Recent progress has shown that GABA(A) receptors also provide a wide range of additional functions beyond simple inhibition. This diversity of functions is mediated by a large variety of different interneuron classes acting on a diverse population of receptor subtypes. Here, I will focus on an additional source of GABAergic signaling diversity, caused by the highly variable ion signaling mechanism of GABA(A) receptors. In concert with the other two sources of GABAergic heterogeneity, this variability in signaling allows for a wide array of GABAergic effects that are crucial for the development of the brain and its function. © 2015 Elsevier Inc. All rights reserved.

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.

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

Synaptic Mechanisms Generating Orientation Selectivity in the ON Pathway of the Rabbit Retina

PubMed Central

Venkataramani, Sowmya

2016-01-01

Neurons that signal the orientation of edges within the visual field have been widely studied in primary visual cortex. Much less is known about the mechanisms of orientation selectivity that arise earlier in the visual stream. Here we examine the synaptic and morphological properties of a subtype of orientation-selective ganglion cell in the rabbit retina. The receptive field has an excitatory ON center, flanked by excitatory OFF regions, a structure similar to simple cell receptive fields in primary visual cortex. Examination of the light-evoked postsynaptic currents in these ON-type orientation-selective ganglion cells (ON-OSGCs) reveals that synaptic input is mediated almost exclusively through the ON pathway. Orientation selectivity is generated by larger excitation for preferred relative to orthogonal stimuli, and conversely larger inhibition for orthogonal relative to preferred stimuli. Excitatory orientation selectivity arises in part from the morphology of the dendritic arbors. Blocking GABAA receptors reduces orientation selectivity of the inhibitory synaptic inputs and the spiking responses. Negative contrast stimuli in the flanking regions produce orientation-selective excitation in part by disinhibition of a tonic NMDA receptor-mediated input arising from ON bipolar cells. Comparison with earlier studies of OFF-type OSGCs indicates that diverse synaptic circuits have evolved in the retina to detect the orientation of edges in the visual input. SIGNIFICANCE STATEMENT A core goal for visual neuroscientists is to understand how neural circuits at each stage of the visual system extract and encode features from the visual scene. This study documents a novel type of orientation-selective ganglion cell in the retina and shows that the receptive field structure is remarkably similar to that of simple cells in primary visual cortex. However, the data indicate that, unlike in the cortex, orientation selectivity in the retina depends on the activity of

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.

Synaptic Glutamate Spillover Due to Impaired Glutamate Uptake Mediates Heroin Relapse

PubMed Central

Scofield, Michael D.; Boger, Heather; Hensley, Megan; Kalivas, Peter W.

2014-01-01

Reducing the enduring vulnerability to relapse is a therapeutic goal in treating drug addiction. Studies with animal models of drug addiction show a marked increase in extrasynaptic glutamate in the core subcompartment of the nucleus accumbens (NAcore) during reinstated drug seeking. However, the synaptic mechanisms linking drug-induced changes in extrasynaptic glutamate to relapse are poorly understood. Here, we discovered impaired glutamate elimination in rats extinguished from heroin self-administration that leads to spillover of synaptically released glutamate into the nonsynaptic extracellular space in NAcore and investigated whether restoration of glutamate transport prevented reinstated heroin seeking. Through multiple functional assays of glutamate uptake and analyzing NMDA receptor-mediated currents, we show that heroin self-administration produced long-lasting downregulation of glutamate uptake and surface expression of the transporter GLT-1. This downregulation was associated with spillover of synaptic glutamate to extrasynaptic NMDA receptors within the NAcore. Ceftriaxone restored glutamate uptake and prevented synaptic glutamate spillover and cue-induced heroin seeking. Ceftriaxone-induced inhibition of reinstated heroin seeking was blocked by morpholino-antisense targeting GLT-1 synthesis. These data reveal that the synaptic glutamate spillover in the NAcore results from reduced glutamate transport and is a critical pathophysiological mechanism underling reinstated drug seeking in rats extinguished from heroin self-administration. PMID:24741055

Transmission to interneurons is via slow excitatory synaptic potentials mediated by P2Y(1) receptors during descending inhibition in guinea-pig ileum.

PubMed

Thornton, Peter D J; Gwynne, Rachel M; McMillan, Darren J; Bornstein, Joel C

2013-01-01

The nature of synaptic transmission at functionally distinct synapses in intestinal reflex pathways has not been fully identified. In this study, we investigated whether transmission between interneurons in the descending inhibitory pathway is mediated by a purine acting at P2Y receptors to produce slow excitatory synaptic potentials (EPSPs). Myenteric neurons from guinea-pig ileum in vitro were impaled with intracellular microelectrodes. Responses to distension 15 mm oral to the recording site, in a separately perfused stimulation chamber and to electrical stimulation of local nerve trunks were recorded. A subset of neurons, previously identified as nitric oxide synthase immunoreactive descending interneurons, responded to both stimuli with slow EPSPs that were reversibly abolished by a high concentration of PPADS (30 μM, P2 receptor antagonist). When added to the central chamber of a three chambered organ bath, PPADS concentration-dependently depressed transmission through that chamber of descending inhibitory reflexes, measured as inhibitory junction potentials in the circular muscle of the anal chamber. Reflexes evoked by distension in the central chamber were unaffected. A similar depression of transmission was seen when the specific P2Y(1) receptor antagonist MRS 2179 (10 μM) was in the central chamber. Blocking either nicotinic receptors (hexamethonium 200 μM) or 5-HT(3) receptors (granisetron 1 μM) together with P2 receptors had no greater effect than blocking P2 receptors alone. Slow EPSPs mediated by P2Y(1) receptors, play a primary role in transmission between descending interneurons of the inhibitory reflexes in the guinea-pig ileum. This is the first demonstration for a primary role of excitatory metabotropic receptors in physiological transmission at a functionally identified synapse.

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

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.

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

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

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

Effects of chronic exposure to an anabolic androgenic steroid cocktail on alpha5-receptor-mediated GABAergic transmission and neural signaling in the forebrain of female mice.

PubMed

Penatti, C A A; Costine, B A; Porter, D M; Henderson, L P

2009-06-30

Anabolic androgenic steroids (AAS) are synthetic derivatives of testosterone that are illicitly self-administered for enhancement of performance and body image, but which also have significant effects on the brain and on behavior. While the stereotypical AAS user is an adult male, AAS abuse in women is rapidly increasing, yet few studies have examined AAS effects in female subjects. We have assessed the effects in female mice of a combination of commonly abused AAS on neuronal activity and neurotransmission mediated by GABA type A (GABA(A)) receptors in the medial preoptic nucleus (MPN); a nexus in the circuits of the hypothalamus and forebrain that are critical for the expression of social behaviors known to be altered in AAS abuse. Our data indicate that chronic exposure to AAS resulted in androgen receptor (AR)-dependent upregulation of alpha(5), beta(3) and delta subunit mRNAs. Acute application of the alpha(5) subunit-selective inverse agonist, L-655,708 (L6), indicated that a significant fraction of the synaptic current is carried by alpha(5)-containing receptors and that AAS treatment may enhance expression of alpha(5)-containing receptors contributing to synaptic, but not tonic, currents in the MPN. AAS treatment also resulted in a significant decrease in action potential frequency in MPN neurons that was also correlated with an increased sensitivity to L-655,708. Our data demonstrate that chronic exposure to multiple AAS elicits significant changes in GABAergic transmission and neuronal activity that are likely to reflect changes in the expression of alpha(5)-containing synaptic receptors within the MPN.

Effects of Chronic Exposure to an Anabolic Androgenic Steroid Cocktail on α5-Receptor Mediated GABAergic Transmission and Neural Signaling in the Forebrain of Female Mice

PubMed Central

Penatti, Carlos A. A.; Costine, Beth A.; Porter, Donna M.; Henderson, Leslie P.

2009-01-01

Anabolic-androgenic steroids (AAS) are synthetic derivatives of testosterone that are illicitly self-administered for enhancement of performance and body image, but which also have significant effects on the brain and on behavior. While the stereotypical AAS user is an adult male, AAS abuse in women is rapidly increasing, yet few studies have examined AAS effects in female subjects. We have assessed the effects in female mice of a combination of commonly abused AAS on neuronal activity and neurotransmission mediated by γ-aminobutyric acid type A (GABAA) receptors in the medial preoptic nucleus (MPN); a nexus in the circuits of the hypothalamus and forebrain that are critical for the expression of social behaviors known to be altered in AAS abuse. Our data indicate that chronic exposure to AAS resulted in androgen receptor (AR)-dependent upregulation of α5, β3 and δ subunit mRNA. Acute application of the α5 subunit-selective inverse agonist, L-655,708, indicated that a significant fraction of the synaptic current is carried by α5-containing receptors and that AAS treatment may enhance expression of α5-containing receptors contributing to synaptic, but not tonic, currents in the MPN. AAS treatment also resulted in a significant decrease in action potential frequency in MPN neurons that was also correlated with an increased sensitivity to L655,708. Our data demonstrate that chronic exposure to multiple AAS elicits significant changes in GABAergic transmission and neuronal activity that are likely to reflect changes in the expression of α5-containing synaptic receptors within the MPN. PMID:19324077

Tau mislocalization to dendritic spines mediates synaptic dysfunction independently of neurodegeneration

PubMed Central

Hoover, Brian R.; Reed, Miranda N.; Su, Jianjun; Penrod, Rachel D.; Kotilinek, Linda A.; Grant, Marianne K.; Pitstick, Rose; Carlson, George A.; Lanier, Lorene M.; Yuan, Li-Lian; Ashe, Karen H.; Liao, Dezhi

2010-01-01

The microtubule-associated protein tau accumulates in Alzheimer’s and other fatal dementias, which manifest when forebrain neurons die. Recent advances in understanding these disorders indicate that brain dysfunction precedes neurodegeneration, but the role of tau is unclear. Here, we show that early tau-related deficits develop not from the loss of synapses or neurons, but rather as a result of synaptic abnormalities caused by the accumulation of hyperphosphorylated tau within intact dendritic spines, where it disrupts synaptic function by impairing glutamate receptor trafficking or synaptic anchoring. Mutagenesis of 14 disease-associated serine and threonine amino acid residues to create pseudohyperphosphorylated tau caused tau mislocalization while creation of phosphorylation-deficient tau blocked the mis-targeting of tau to dendritic spines. Thus, tau phosphorylation plays a critical role in mediating tau mislocalization and subsequent synaptic impairment. These data establish that the locus of early synaptic malfunction caused by tau resides in dendritic spines. PMID:21172610

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.

GABA(B) receptor modulation of feedforward inhibition through hippocampal neurogliaform cells.

PubMed

Price, Christopher J; Scott, Ricardo; Rusakov, Dmitri A; Capogna, Marco

2008-07-02

Feedforward inhibition of neurons is a fundamental component of information flow control in the brain. We studied the roles played by neurogliaform cells (NGFCs) of stratum lacunosum moleculare of the hippocampus in providing feedforward inhibition to CA1 pyramidal cells. We recorded from synaptically coupled pairs of anatomically identified NGFCs and CA1 pyramidal cells and found that, strikingly, a single presynaptic action potential evoked a biphasic unitary IPSC (uIPSC), consisting of two distinct components mediated by GABA(A) and GABA(B) receptors. A GABA(B) receptor-mediated unitary response has not previously been observed in hippocampal excitatory neurons. The decay of the GABA(A) receptor-mediated response was slow (time constant = 50 ms), and was tightly regulated by presynaptic GABA(B) receptors. Surprisingly, the GABA(B) receptor ligands baclofen and (2S)-3-{[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl}(phenylmethyl)phosphinic acid (CGP55845), while affecting the NGFC-mediated uIPSCs, had no effect on action potential-evoked presynaptic Ca2+ signals monitored in individual axonal boutons of NGFCs with two-photon microscopy. In contrast, baclofen clearly depressed presynaptic Ca2+ transients in non-NGF interneurons. Changes in extracellular Ca2+ concentration that mimicked the effects of baclofen or CGP55845 on uIPSCs significantly altered presynaptic Ca2+ transients. Electrophysiological data suggest that GABA(B) receptors expressed by NGFCs contribute to the dynamic control of the excitatory input to CA1 pyramidal neurons from the temporoammonic path. The NGFC-CA1 pyramidal cell connection therefore provides a unique and subtle mechanism to shape the integration time domain for signals arriving via a major excitatory input to CA1 pyramidal cells.

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

Contribution of Resting Conductance, GABAA-Receptor Mediated Miniature Synaptic Currents and Neurosteroid to Chloride Homeostasis in Central Neurons.

PubMed

Yelhekar, Tushar D; Druzin, Michael; Johansson, Staffan

2017-01-01

Maintenance of a low intraneuronal Cl - concentration, [Cl - ] i , is critical for inhibition in the CNS. Here, the contribution of passive, conductive Cl - flux to recovery of [Cl - ] i after a high load was analyzed in mature central neurons from rat. A novel method for quantifying the resting Cl - conductance, important for [Cl - ] i recovery, was developed and the possible contribution of GABA A and glycine receptors and of ClC-2 channels to this conductance was analyzed. The hypothesis that spontaneous, action potential-independent release of GABA is important for [Cl - ] i recovery was tested. [Cl - ] i was examined by gramicidin-perforated patch recordings in medial preoptic neurons. Cells were loaded with Cl - by combining GABA or glycine application with a depolarized voltage, and the time course of [Cl - ] i was followed by measurements of the Cl - equilibrium potential , as obtained from the current recorded during voltage ramps combined with GABA or glycine application. The results show that passive Cl - flux contributes significantly, in the same order of magnitude as does K + -Cl - cotransporter 2 (KCC2), to [Cl - ] i recovery and that Cl - conductance accounts for ∼ 6% of the total resting conductance. A major fraction of this resting Cl - conductance is picrotoxin (PTX)-sensitive and likely due to open GABA A receptors, but ClC-2 channels do not contribute. The results also show that when the decay of GABA A receptor-mediated miniature postsynaptic currents (minis) is slowed by the neurosteroid allopregnanolone, such minis may significantly quicken [Cl - ] i recovery, suggesting a possible steroid-regulated role for minis in the control of Cl - homeostasis.

Neurosteroid modulation of neuronal excitability and synaptic transmission in the rat medial vestibular nuclei.

PubMed

Grassi, Silvarosa; Frondaroli, Adele; Dieni, Cristina; Dutia, Mayank B; Pettorossi, Vito E

2007-07-01

In rat brainstem slices, we investigated the influence of the neurosteroids tetrahydrodeoxycorticosterone (THDOC) and allopregnanolone (ALLO) on the synaptically driven and spontaneous activity of vestibular neurons, by analysing their effects on the amplitude of the field potentials evoked in the medial vestibular nuclei (MVN) by vestibular afferent stimulation and on the spontaneous firing rate of MVN neurons. Furthermore, the interaction with gamma-aminobutyric acid (GABA) and glutamate receptors was analysed by using specific antagonists for GABA(A) (bicuculline), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/ kainate [2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo(f)quinoxaline-7-sulphonamide disodium salt (NBQX)], N-methyl-D-aspartate (NMDA) [D-(-)-2-amino-5-phosphonopentanoic acid (AP-5)] and group I metabotropic glutamate receptors (mGlu-I) [(R,S)-1-aminoindan-1,5-dicarboxylic acid (AIDA)] receptors. THDOC and ALLO evoked two opposite long-lasting effects, consisting of either a potentiation or a reduction of field potential and firing rate, which showed early and late components, occurring in conjunction or separately after neurosteroid application. The depressions depended on GABA(A) receptors, as they were abolished by bicuculline, while early potentiation involved glutamate AMPA/kainate receptors, as NBQX markedly reduced the incidence of early firing rate enhancement and, in the case of ALLO, even provoked depression. This suggests that THDOC and ALLO enhance the GABA(A) inhibitory influence on the MVN neurons and facilitate the AMPA/kainate facilitatory one. Conversely, a late potentiation effect, which was still induced after glutamate and GABA(A) receptor blockade, might involve a different mechanism. We conclude that the modulation of neuronal activity in the MVN by THDOC and ALLO, through their actions on GABA(A) and AMPA/kainate receptors, may have a physiological role in regulating the vestibular system function under normal

Calmodulin activity regulates group I metabotropic glutamate receptor-mediated signal transduction and synaptic depression.

PubMed

Sethna, Ferzin; Zhang, Ming; Kaphzan, Hanoch; Klann, Eric; Autio, Dawn; Cox, Charles L; Wang, Hongbing

2016-05-01

Group I metabotropic glutamate receptors (mGluR), including mGluR1 and mGluR 5 (mGluR1/5), are coupled to Gq and modulate activity-dependent synaptic plasticity. Direct activation of mGluR1/5 causes protein translation-dependent long-term depression (LTD). Although it has been established that intracellular Ca(2+) and the Gq-regulated signaling molecules are required for mGluR1/5 LTD, whether and how Ca(2+) regulates Gq signaling and upregulation of protein expression remain unknown. Through pharmacological inhibition, we tested the function of the Ca(2+) sensor calmodulin (CaM) in intracellular signaling triggered by the activation of mGluR1/5. CaM inhibitor N-[4-aminobutyl]-5-chloro-2-naphthalenesulfonamide hydrochloride (W13) suppressed the mGluR1/5-stimulated activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and p70-S6 kinase 1 (S6K1) in hippocampal neurons. W13 also blocked the mGluR1/5 agonist-induced synaptic depression in hippocampal slices and in anesthetized mice. Consistent with the function of CaM, inhibiting the downstream targets Ca(2+) /CaM-dependent protein kinases (CaMK) blocked ERK1/2 and S6K1 activation. Furthermore, disruption of the CaM-CaMK-ERK1/2 signaling cascade suppressed the mGluR1/5-stimulated upregulation of Arc expression. Altogether, our data suggest CaM as a new Gq signaling component for coupling Ca(2+) and protein upregulation and regulating mGluR1/5-mediated synaptic modification. © 2016 Wiley Periodicals, Inc.

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.

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 NMDA Receptor Properties and Synaptic Transmission by the NR3A Subunit in Mouse Hippocampal and Cerebrocortical Neurons

PubMed Central

Tong, Gary; Takahashi, Hiroto; Tu, Shichun; Shin, Yeonsook; Talantova, Maria; Zago, Wagner; Xia, Peng; Nie, Zhiguo; Goetz, Thomas; Zhang, Dongxian; Lipton, Stuart A.; Nakanishi, Nobuki

2015-01-01

Expression of the NR3A subunit with NR1/NR2 in Xenopus oocytes or mammalian cell lines leads to a reduction in N-methyl-D-aspartate (NMDA)-induced currents and decreased Mg2+ sensitivity and Ca2+ permeability compared with NR1/NR2 receptors. Consistent with these findings, neurons from NR3A knockout (KO) mice exhibit enhanced NMDA-induced currents. Recombinant NR3A can also form excitatory glycine receptors with NR1 in the absence of NR2. However, the effects of NR3A on channel properties in neurons and synaptic transmission have not been fully elucidated. To study physiological roles of NR3A subunits, we generated NR3A transgenic (Tg) mice. Cultured NR3A Tg neurons exhibited two populations of NMDA receptor (NMDAR) channels, reduced Mg2+ sensitivity, and decreased Ca2+ permeability in response to NMDA/glycine, but glycine alone did not elicit excitatory currents. In addition, NMDAR-mediated excitatory postsynaptic currents (EPSCs) in NR3A Tg hippocampal slices showed reduced Mg2+ sensitivity, consistent with the notion that NR3A subunits incorporated into synaptic NMDARs. To study the function of endogenous NR3A subunits, we compared NMDAR-mediated EPSCs in NR3A KO and WT control mice. In NR3A KO mice, the ratio of the amplitudes of the NMDAR-mediated component to α-amino-3-hydroxy-5-methyl-4-isox-azolepropionic acid receptor-mediated component of the EPSC was significantly larger than that seen in WT littermates. This result suggests that NR3A subunits contributed to the NMDAR-mediated component of the EPSC in WT mice. Taken together, these results show that NR3A subunits contribute to NMDAR responses from both synaptic and extra-synaptic receptors, likely composed of NR1, NR2, and NR3 subunits. PMID:18003876

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

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.

Spontaneous Release Regulates Synaptic Scaling in the Embryonic Spinal Network In Vivo

PubMed Central

Garcia-Bereguiain, Miguel Angel; Gonzalez-Islas, Carlos; Lindsly, Casie

2016-01-01

Homeostatic plasticity mechanisms maintain cellular or network spiking activity within a physiologically functional range through compensatory changes in synaptic strength or intrinsic cellular excitability. Synaptic scaling is one form of homeostatic plasticity that is triggered after blockade of spiking or neurotransmission in which the strengths of all synaptic inputs to a cell are multiplicatively scaled upward or downward in a compensatory fashion. We have shown previously that synaptic upscaling could be triggered in chick embryo spinal motoneurons by complete blockade of spiking or GABAA receptor (GABAAR) activation for 2 d in vivo. Here, we alter GABAAR activation in a more physiologically relevant manner by chronically adjusting presynaptic GABA release in vivo using nicotinic modulators or an mGluR2 agonist. Manipulating GABAAR activation in this way triggered scaling in a mechanistically similar manner to scaling induced by complete blockade of GABAARs. Remarkably, we find that altering action-potential (AP)-independent spontaneous release was able to fully account for the observed bidirectional scaling, whereas dramatic changes in spiking activity associated with spontaneous network activity had little effect on quantal amplitude. The reliance of scaling on an AP-independent process challenges the plasticity's relatedness to spiking in the living embryonic spinal network. Our findings have implications for the trigger and function of synaptic scaling and suggest that spontaneous release functions to regulate synaptic strength homeostatically in vivo. SIGNIFICANCE STATEMENT Homeostatic synaptic scaling is thought to prevent inappropriate levels of spiking activity through compensatory adjustments in the strength of synaptic inputs. Therefore, it is thought that perturbations in spike rate trigger scaling. Here, we find that dramatic changes in spiking activity in the embryonic spinal cord have little effect on synaptic scaling; conversely, alterations in

Wnt signaling pathway improves central inhibitory synaptic transmission in a mouse model of Duchenne muscular dystrophy.

PubMed

Fuenzalida, Marco; Espinoza, Claudia; Pérez, Miguel Ángel; Tapia-Rojas, Cheril; Cuitino, Loreto; Brandan, Enrique; Inestrosa, Nibaldo C

2016-02-01

The dystrophin-associated glycoprotein complex (DGC) that connects the cytoskeleton, plasma membrane and the extracellular matrix has been related to the maintenance and stabilization of channels and synaptic receptors, which are both essential for synaptogenesis and synaptic transmission. The dystrophin-deficient (mdx) mouse model of Duchenne muscular dystrophy (DMD) exhibits a significant reduction in hippocampal GABA efficacy, which may underlie the altered synaptic function and abnormal hippocampal long-term plasticity exhibited by mdx mice. Emerging studies have implicated Wnt signaling in the modulation of synaptic efficacy, neuronal plasticity and cognitive function. We report here that the activation of the non-canonical Wnt-5a pathway and Andrographolide, improves hippocampal mdx GABAergic efficacy by increasing the number of inhibitory synapses and GABA(A) receptors or GABA release. These results indicate that Wnt signaling modulates GABA synaptic efficacy and could be a promising novel target for DMD cognitive therapy. Copyright © 2015 Elsevier Inc. All rights reserved.

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.

Extrasynaptic Glycine Receptors of Rodent Dorsal Raphe Serotonergic Neurons: A Sensitive Target for Ethanol

PubMed Central

Maguire, Edward P; Mitchell, Elizabeth A; Greig, Scott J; Corteen, Nicole; Balfour, David J K; Swinny, Jerome D; Lambert, Jeremy J; Belelli, Delia

2014-01-01

Alcohol abuse is a significant medical and social problem. Several neurotransmitter systems are implicated in ethanol's actions, with certain receptors and ion channels emerging as putative targets. The dorsal raphe (DR) nucleus is associated with the behavioral actions of alcohol, but ethanol actions on these neurons are not well understood. Here, using immunohistochemistry and electrophysiology we characterize DR inhibitory transmission and its sensitivity to ethanol. DR neurons exhibit inhibitory ‘phasic' post-synaptic currents mediated primarily by synaptic GABAA receptors (GABAAR) and, to a lesser extent, by synaptic glycine receptors (GlyR). In addition to such phasic transmission mediated by the vesicular release of neurotransmitter, the activity of certain neurons may be governed by a ‘tonic' conductance resulting from ambient GABA activating extrasynaptic GABAARs. However, for DR neurons extrasynaptic GABAARs exert only a limited influence. By contrast, we report that unusually the GlyR antagonist strychnine reveals a large tonic conductance mediated by extrasynaptic GlyRs, which dominates DR inhibition. In agreement, for DR neurons strychnine increases their input resistance, induces membrane depolarization, and consequently augments their excitability. Importantly, this glycinergic conductance is greatly enhanced in a strychnine-sensitive fashion, by behaviorally relevant ethanol concentrations, by drugs used for the treatment of alcohol withdrawal, and by taurine, an ingredient of certain ‘energy drinks' often imbibed with ethanol. These findings identify extrasynaptic GlyRs as critical regulators of DR excitability and a novel molecular target for ethanol. PMID:24264816

Selective effects of aniracetam across receptor types and forms of synaptic facilitation in hippocampus.

PubMed

Xiao, P; Staubli, U; Kessler, M; Lynch, G

1991-10-01

Aniracetam reversibly increased synaptic responses mediated by the AMPA but not the NMDA subclass of glutamate receptors in hippocampus and was considerably more potent than structurally similar nootropics. The drug had greater effects on field excitatory postsynaptic potentials (EPSPs) in the dentate gyrus and CA1 region than it did in the CA3 region, suggesting that it differentiates between variants of the AMPA receptor. Ligand binding to glutamate receptors in synaptosomal membrane fractions was minimally changed by aniracetam. Finally, the percent facilitation produced by aniracetam in the CA1 region was not reduced by any of three treatments (4-aminopyridine, changes in extracellular calcium concentrations, paired-pulse stimulation) that affect release but, in accord with a previous report, was substantially decreased by long-term potentiation. These results support the conclusion that aniracetam selectively increases the conductance of a subgroup of synaptic AMPA receptors in hippocampus and suggest that receptor changes underlie the expression of long-term potentiation.

CPG2 Recruits Endophilin B2 to the Cytoskeleton for Activity-Dependent Endocytosis of Synaptic Glutamate Receptors.

PubMed

Loebrich, Sven; Benoit, Marc Robert; Konopka, Jaclyn Aleksandra; Cottrell, Jeffrey Richard; Gibson, Joanne; Nedivi, Elly

2016-02-08

Internalization of glutamate receptors at the postsynaptic membrane via clathrin-mediated endocytosis (CME) is a key mechanism for regulating synaptic strength. A role for the F-actin cytoskeleton in CME is well established, and recently, PKA-dependent association of candidate plasticity gene 2 (CPG2) with the spine-cytoskeleton has been shown to mediate synaptic glutamate receptor internalization. Yet, how the endocytic machinery is physically coupled to the actin cytoskeleton to facilitate glutamate receptor internalization has not been demonstrated. Moreover, there has been no distinction of endocytic-machinery components that are specific to activity-dependent versus constitutive glutamate receptor internalization. Here, we show that CPG2, through a direct physical interaction, recruits endophilin B2 (EndoB2) to F-actin, thus anchoring the endocytic machinery to the spine cytoskeleton and facilitating glutamate receptor internalization. Regulation of CPG2 binding to the actin cytoskeleton by protein kinase A directly impacts recruitment of EndoB2 and clathrin. Specific disruption of EndoB2 or the CPG2-EndoB2 interaction impairs activity-dependent, but not constitutive, internalization of both NMDA- and AMPA-type glutamate receptors. These results demonstrate that, through direct interactions with F-actin and EndoB2, CPG2 physically bridges the spine cytoskeleton and the endocytic machinery, and this tripartite association is critical specifically for activity-dependent CME of synaptic glutamate receptors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Laminar distribution of GABAA- and glycine-receptor mediated tonic inhibition in the dorsal horn of the rat lumbar spinal cord: effects of picrotoxin and strychnine on expression of Fos-like immunoreactivity.

PubMed

Cronin, John N; Bradbury, Elizabeth J; Lidierth, Malcolm

2004-11-01

Inhibitory mechanisms are essential in suppressing the development of allodynia and hyperalgesia in the normal animal and there is evidence that loss of inhibition can lead to the development of neuropathic pain. We used Fos expression to map the distribution of tonically inhibited cells in the healthy rat lumbar spinal cord. In a control group, Fos-like immunoreactive (Fos-LI) cells were rare, averaging 7.5+/-2.2 cells (mean+/-SEM; N=13 sections) per 20 microm thick section of dorsal horn. This rose to 103+/-11 (mean+/-SEM; N=20) in picrotoxin-treated rats and to 88+/-11 (mean+/-SEM; N=18) in strychnine-treated rats. These changes were significant (ANOVA; P<0.001). There were marked regional variations in the distribution of Fos-LI cells between picrotoxin- and strychnine-treated animals. Picrotoxin induced a significant increase in the number of Fos-LI cells throughout the dorsal horn (lamina I-VI) while strychnine significantly elevated Fos-like immunoreactivity only in deep laminae (III-VI). For both picrotoxin and strychnine, the increase in Fos-like immunoreactivity peaked in lamina V (at 3579+/-319 and 3649+/-375% of control, respectively; mean+/-SEM) but for picrotoxin an additional peak was observed in the outer part of lamina II (1959+/-196%). Intrathecal administration of both GABAA and glycine receptor antagonists has been shown elsewhere to induce tactile allodynia. The present data suggest that this allodynia could arise due to blockade of tonic GABAA and glycine-receptor mediated inhibition in the deep dorsal horn. GABAA antagonists also induce hypersensitivity to noxious inputs. The blockade of tonic inhibition in the superficial dorsal horn shown here may underlie this hyperalgesia.

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.

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 prelimbic medial prefrontal cortex in panic-like elaborated defensive behaviour and innate fear-induced antinociception elicited by GABAA receptor blockade in the dorsomedial and ventromedial hypothalamic nuclei: role of the endocannabinoid CB1 receptor.

PubMed

Freitas, Renato Leonardo de; Salgado-Rohner, Carlos José; Hallak, Jaime Eduardo Cecílio; Crippa, José Alexandre de Souza; Coimbra, Norberto Cysne

2013-09-01

It has been shown that GABAA receptor blockade in the dorsomedial and ventromedial hypothalamic nuclei (DMH and VMH, respectively) induces elaborated defensive behavioural responses accompanied by antinociception, which has been utilized as an experimental model of panic attack. Furthermore, the prelimbic (PL) division of the medial prefrontal cortex (MPFC) has been related to emotional reactions and the processing of nociceptive information. The aim of the present study was to investigate the possible involvement of the PL cortex and the participation of local cannabinoid CB1 receptors in the elaboration of panic-like reactions and in innate fear-induced antinociception. Elaborated fear-induced responses were analysed during a 10-min period in an open-field test arena. Microinjection of the GABAA receptor antagonist bicuculline into the DMH/VMH evoked panic-like behaviour and fear-induced antinociception, which was decreased by microinjection of the non-selective synaptic contact blocker cobalt chloride in the PL cortex. Moreover, microinjection of AM251 (25, 100 or 400 pmol), an endocannabinoid CB1 receptor antagonist, into the PL cortex also attenuated the defensive behavioural responses and the antinociception that follows innate fear behaviour elaborated by DMH/VMH. These data suggest that the PL cortex plays an important role in the organization of elaborated forward escape behaviour and that this cortical area is also involved in the elaboration of innate fear-induced antinociception. Additionally, CB1 receptors in the PL cortex modulate both panic-like behaviours and fear-induced antinociception elicited by disinhibition of the DMH/VMH through microinjection of bicuculline.

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

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

Synaptic Mechanisms Generating Orientation Selectivity in the ON Pathway of the Rabbit Retina.

PubMed

Venkataramani, Sowmya; Taylor, W Rowland

2016-03-16

Neurons that signal the orientation of edges within the visual field have been widely studied in primary visual cortex. Much less is known about the mechanisms of orientation selectivity that arise earlier in the visual stream. Here we examine the synaptic and morphological properties of a subtype of orientation-selective ganglion cell in the rabbit retina. The receptive field has an excitatory ON center, flanked by excitatory OFF regions, a structure similar to simple cell receptive fields in primary visual cortex. Examination of the light-evoked postsynaptic currents in these ON-type orientation-selective ganglion cells (ON-OSGCs) reveals that synaptic input is mediated almost exclusively through the ON pathway. Orientation selectivity is generated by larger excitation for preferred relative to orthogonal stimuli, and conversely larger inhibition for orthogonal relative to preferred stimuli. Excitatory orientation selectivity arises in part from the morphology of the dendritic arbors. Blocking GABAA receptors reduces orientation selectivity of the inhibitory synaptic inputs and the spiking responses. Negative contrast stimuli in the flanking regions produce orientation-selective excitation in part by disinhibition of a tonic NMDA receptor-mediated input arising from ON bipolar cells. Comparison with earlier studies of OFF-type OSGCs indicates that diverse synaptic circuits have evolved in the retina to detect the orientation of edges in the visual input. A core goal for visual neuroscientists is to understand how neural circuits at each stage of the visual system extract and encode features from the visual scene. This study documents a novel type of orientation-selective ganglion cell in the retina and shows that the receptive field structure is remarkably similar to that of simple cells in primary visual cortex. However, the data indicate that, unlike in the cortex, orientation selectivity in the retina depends on the activity of inhibitory interneurons. The

Stereoselective modulatory actions of oleamide on GABAA receptors and voltage-gated Na+ channels in vitro: a putative endogenous ligand for depressant drug sites in CNS

PubMed Central

Verdon, Bernard; Zheng, Jian; Nicholson, Russell A; Ganelli, C Robin; Lees, George

2000-01-01

cis-9,10-octadecenoamide (‘oleamide') accumulates in CSF on sleep deprivation. It induces sleep in animals (the trans form is inactive) but its cellular actions are poorly characterized. We have used electrophysiology in cultures from embryonic rat cortex and biochemical studies in mouse nerve preparations to address these issues. Twenty μM cis-oleamide (but not trans) reversibly enhanced GABAA currents and depressed the frequency of spontaneous excitatory and inhibitory synaptic activity in cultured networks. cis-oleamide stereoselectively blocked veratridine-induced (but not K+-induced) depolarisation of mouse synaptoneurosomes (IC50, 13.9 μM). The cis isomer stereoselectively blocked veratridine-induced (but not K+-induced) [3H]-GABA release from mouse synaptosomes (IC50, 4.6 μM). At 20 μM cis-oleamide, but not trans, produced a marked inhibition of Na+ channel-dependent rises in intrasynaptosomal Ca2+. The physiological significance of these observations was examined by isolating Na+ spikes in cultured pyramidal neurones. Sixty-four μM cis-oleamide did not significantly alter the amplitude, rate of rise or duration of unitary action potentials (1 Hz). cis-Oleamide stereoselectively suppressed sustained repetitive firing (SRF) in these cells with an EC50 of 4.1 μM suggesting a frequency- or state-dependent block of voltage-gated Na+ channels. Oleamide is a stereoselective modulator of both postsynaptic GABAA receptors and presynaptic or somatic voltage-gated Na+ channels which are crucial for synaptic inhibition and conduction. The modulatory actions are strikingly similar to those displayed by sedative or anticonvulsant barbiturates and a variety of general anaesthetics. Oleamide may represent an endogenous modulator for drug receptors and an important regulator of arousal. PMID:10694234

Bidirectional modulation of hippocampal synaptic plasticity by Dopaminergic D4-receptors in the CA1 area of hippocampus.

PubMed

Navakkode, Sheeja; Chew, Katherine C M; Tay, Sabrina Jia Ning; Lin, Qingshu; Behnisch, Thomas; Soong, Tuck Wah

2017-11-14

Long-term potentiation (LTP) is the persistent increase in the strength of the synapses. However, the neural networks would become saturated if there is only synaptic strenghthening. Synaptic weakening could be facilitated by active processes like long-term depression (LTD). Molecular mechanisms that facilitate the weakening of synapses and thereby stabilize the synapses are also important in learning and memory. Here we show that blockade of dopaminergic D4 receptors (D4R) promoted the formation of late-LTP and transformed early-LTP into late-LTP. This effect was dependent on protein synthesis, activation of NMDA-receptors and CaMKII. We also show that GABA A -receptor mediated mechanisms are involved in the enhancement of late-LTP. We could show that short-term plasticity and baseline synaptic transmission were unaffected by D4R inhibition. On the other hand, antagonizing D4R prevented both early and late forms of LTD, showing that activation of D4Rs triggered a dual function. Synaptic tagging experiments on LTD showed that D4Rs act as plasticity related proteins rather than the setting of synaptic tags. D4R activation by PD 168077 induced a slow-onset depression that was protein synthesis, NMDAR and CaMKII dependent. The D4 receptors, thus exert a bidirectional modulation of CA1 pyramidal neurons by restricting synaptic strengthening and facilitating synaptic weakening.

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

JIP1-Mediated JNK Activation Negatively Regulates Synaptic Plasticity and Spatial Memory.

PubMed

Morel, Caroline; Sherrin, Tessi; Kennedy, Norman J; Forest, Kelly H; Avcioglu Barutcu, Seda; Robles, Michael; Carpenter-Hyland, Ezekiel; Alfulaij, Naghum; Standen, Claire L; Nichols, Robert A; Benveniste, Morris; Davis, Roger J; Todorovic, Cedomir

2018-04-11

The c-Jun N-terminal kinase (JNK) signal transduction pathway is implicated in learning and memory. Here, we examined the role of JNK activation mediated by the JNK-interacting protein 1 (JIP1) scaffold protein. We compared male wild-type mice with a mouse model harboring a point mutation in the Jip1 gene that selectively blocks JIP1-mediated JNK activation. These male mutant mice exhibited increased NMDAR currents, increased NMDAR-mediated gene expression, and a lower threshold for induction of hippocampal long-term potentiation. The JIP1 mutant mice also displayed improved hippocampus-dependent spatial memory and enhanced associative fear conditioning. These results were confirmed using a second JIP1 mutant mouse model that suppresses JNK activity. Together, these observations establish that JIP1-mediated JNK activation contributes to the regulation of hippocampus-dependent, NMDAR-mediated synaptic plasticity and learning. SIGNIFICANCE STATEMENT The results of this study demonstrate that c-Jun N-terminal kinase (JNK) activation induced by the JNK-interacting protein 1 (JIP1) scaffold protein negatively regulates the threshold for induction of long-term synaptic plasticity through the NMDA-type glutamate receptor. This change in plasticity threshold influences learning. Indeed, mice with defects in JIP1-mediated JNK activation display enhanced memory in hippocampus-dependent tasks, such as contextual fear conditioning and Morris water maze, indicating that JIP1-JNK constrains spatial memory. This study identifies JIP1-mediated JNK activation as a novel molecular pathway that negatively regulates NMDAR-dependent synaptic plasticity and memory. Copyright © 2018 the authors 0270-6474/18/383708-21$15.00/0.

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

Presynaptic Kainate Receptor Mediation of Frequency Facilitation at Hippocampal Mossy Fiber Synapses

NASA Astrophysics Data System (ADS)

Schmitz, Dietmar; Mellor, Jack; Nicoll, Roger A.

2001-03-01

Inhibition of transmitter release by presynaptic receptors is widespread in the central nervous system and is typically mediated via metabotropic receptors. In contrast, very little is known about facilitatory receptors, and synaptic activation of a facilitatory autoreceptor has not been established. Here we show that activation of presynaptic kainate receptors can facilitate transmitter release from hippocampal mossy fiber synapses. Synaptic activation of these presumed ionotropic kainate receptors is very fast (<10 ms) and lasts for seconds. Thus, these presynaptic kainate receptors contribute to the short-term plasticity characteristics of mossy fiber synapses, which were previously thought to be an intrinsic property of the synapse.

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.

Neurogranin restores amyloid β-mediated synaptic transmission and long-term potentiation deficits.

PubMed

Kaleka, Kanwardeep Singh; Gerges, Nashaat Z

2016-03-01

Amyloid β (Aβ) is widely considered one of the early causes of cognitive deficits observed in Alzheimer's disease. Many of the deficits caused by Aβ are attributed to its disruption of synaptic function represented by its blockade of long-term potentiation (LTP) and its induction of synaptic depression. Identifying pathways that reverse these synaptic deficits may open the door to new therapeutic targets. In this study, we explored the possibility that Neurogranin (Ng)-a postsynaptic calmodulin (CaM) targeting protein that enhances synaptic function-may rescue Aβ-mediated deficits in synaptic function. Our results show that Ng is able to reverse synaptic depression and LTP deficits induced by Aβ. Furthermore, Ng's restoration of synaptic transmission is through the insertion of GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid glutamate receptors (AMPARs). These restorative effects of Ng are dependent on the interaction of Ng and CaM and CaM-dependent activation of CaMKII. Overall, this study identifies a novel mechanism to rescue synaptic deficits induced by Aβ oligomers. It also suggests Ng and CaM signaling as potential therapeutic targets for Alzheimer's disease as well as important tools to further explore the pathophysiology underlying the disease. Copyright © 2015 Elsevier Inc. All rights reserved.

Enhanced pre-synaptic glutamate release in deep-dorsal horn contributes to calcium channel alpha-2-delta-1 protein-mediated spinal sensitization and behavioral hypersensitivity

PubMed Central

Nguyen, David; Deng, Ping; Matthews, Elizabeth A; Kim, Doo-Sik; Feng, Guoping; Dickenson, Anthony H; Xu, Zao C; Luo, Z David

2009-01-01

Nerve injury-induced expression of the spinal calcium channel alpha-2-delta-1 subunit (Cavα2δ1) has been shown to mediate behavioral hypersensitivity through a yet identified mechanism. We examined if this neuroplasticity modulates behavioral hypersensitivity by regulating spinal glutamatergic neurotransmission in injury-free transgenic mice overexpressing the Cavα2δ1 proteins in neuronal tissues. The transgenic mice exhibited hypersensitivity to mechanical stimulation (allodynia) similar to the spinal nerve ligation injury model. Intrathecally delivered antagonists for N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxyl-5-methylisoxazole-4-propionic acid (AMPA)/kainate receptors, but not for the metabotropic glutamate receptors, caused a dose-dependent allodynia reversal in the transgenic mice without changing the behavioral sensitivity in wild-type mice. This suggests that elevated spinal Cavα2δ1 mediates allodynia through a pathway involving activation of selective glutamate receptors. To determine if this is mediated by enhanced spinal neuronal excitability or pre-synaptic glutamate release in deep-dorsal horn, we examined wide-dynamic-range (WDR) neuron excitability with extracellular recording and glutamate-mediated excitatory postsynaptic currents with whole-cell patch recording in deep-dorsal horn of the Cavα2δ1 transgenic mice. Our data indicated that overexpression of Cavα2δ1 in neuronal tissues led to increased frequency, but not amplitude, of miniature excitatory post synaptic currents mediated mainly by AMPA/kainate receptors at physiological membrane potentials, and also by NMDA receptors upon depolarization, without changing the excitability of WDR neurons to high intensity stimulation. Together, these findings support a mechanism of Cavα2δ1-mediated spinal sensitization in which elevated Cavα2δ1 causes increased pre-synaptic glutamate release that leads to reduced excitation thresholds of post-synaptic dorsal horn neurons to innocuous

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.

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.

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

Gravin orchestrates PKA and β2-adrenergic receptor signaling critical for synaptic plasticity and memory

PubMed Central

Havekes, Robbert; Canton, David A.; Park, Alan J.; Huang, Ted; Nie, Ting; Day, Jonathan P.; Guercio, Leonardo A.; Grimes, Quinn; Luczak, Vincent; Gelman, Irwin H.; Baillie, George S.; Scott, John D.; Abel, Ted

2012-01-01

A kinase-anchoring proteins (AKAPs) organize compartmentalized pools of Protein Kinase A (PKA) to enable localized signaling events within neurons. However, it is unclear which of the many expressed AKAPs in neurons target PKA to signaling complexes important for long-lasting forms of synaptic plasticity and memory storage. In the forebrain, the anchoring protein gravin recruits a signaling complex containing PKA, PKC, calmodulin, and PDE4D to the β2-adrenergic receptor. Here, we show that mice lacking the α-isoform of gravin have deficits in PKA-dependent long-lasting forms of hippocampal synaptic plasticity including β2-adrenergic receptor-mediated plasticity, and selective impairments of long-term memory storage. Further, both hippocampal β2-adrenergic receptor phosphorylation by PKA, and learning-induced activation of ERK, are attenuated in the CA1 region of the hippocampus in mice lacking gravin-α. We conclude that gravin compartmentalizes a significant pool of PKA that regulates learning-induced β2-adrenergic receptor signaling and ERK activation in the hippocampus in vivo, organizing molecular interactions between glutamatergic and noradrenergic signaling pathways for long-lasting synaptic plasticity, and memory storage. PMID:23238728

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

Synaptic proteins and receptors defects in autism spectrum disorders

PubMed Central

Chen, Jianling; Yu, Shunying; Fu, Yingmei; Li, Xiaohong

2014-01-01

Recent studies have found that hundreds of genetic variants, including common and rare variants, rare and de novo mutations, and common polymorphisms contribute to the occurrence of autism spectrum disorders (ASDs). The mutations in a number of genes such as neurexin, neuroligin, postsynaptic density protein 95, SH3, and multiple ankyrin repeat domains 3 (SHANK3), synapsin, gephyrin, cadherin, and protocadherin, thousand-and-one-amino acid 2 kinase, and contactin, have been shown to play important roles in the development and function of synapses. In addition, synaptic receptors, such as gamma-aminobutyric acid receptors and glutamate receptors, have also been associated with ASDs. This review will primarily focus on the defects of synaptic proteins and receptors associated with ASDs and their roles in the pathogenesis of ASDs via synaptic pathways. PMID:25309321

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

Selective antagonism of AMPA receptors unmasks kainate receptor-mediated responses in hippocampal neurons.

PubMed

Paternain, A V; Morales, M; Lerma, J

1995-01-01

Although both protein and mRNAs for kainate receptor subunits are abundant in several brain regions, the responsiveness of AMPA receptors to kainate has made it difficult to demonstrate the presence of functional kainate-type receptors in native cells. Recently, however, we have shown that many hippocampal neurons in culture express glutamate receptors of the kainate type. The large nondesensitizing response that kainate induces at AMPA receptors precludes detection and analysis of smaller, rapidly desensitizing currents induced by kainate at kainate receptors. Consequently, the functional significance of these strongly desensitizing glutamate receptors remains enigmatic. We report here that the family of new noncompetitive antagonists of AMPA receptors (GYKI 52466 and 53655) minimally affects kainate-induced responses at kainate receptors while completely blocking AMPA receptor-mediated currents, making it possible to separate the responses mediated by each receptor. These compounds will allow determination of the role played by kainate receptors in synaptic transmission and plasticity in the mammalian brain, as well as evaluation of their involvement in neurotoxicity.

Metabotropic glutamate receptor-mediated use-dependent down-regulation of synaptic excitability involves the fragile X mental retardation protein.

PubMed

Repicky, Sarah; Broadie, Kendal

2009-02-01

Loss of the mRNA-binding protein FMRP results in the most common inherited form of both mental retardation and autism spectrum disorders: fragile X syndrome (FXS). The leading FXS hypothesis proposes that metabotropic glutamate receptor (mGluR) signaling at the synapse controls FMRP function in the regulation of local protein translation to modulate synaptic transmission strength. In this study, we use the Drosophila FXS disease model to test the relationship between Drosophila FMRP (dFMRP) and the sole Drosophila mGluR (dmGluRA) in regulation of synaptic function, using two-electrode voltage-clamp recording at the glutamatergic neuromuscular junction (NMJ). Null dmGluRA mutants show minimal changes in basal synapse properties but pronounced defects during sustained high-frequency stimulation (HFS). The double null dfmr1;dmGluRA mutant shows repression of enhanced augmentation and delayed onset of premature long-term facilitation (LTF) and strongly reduces grossly elevated post-tetanic potentiation (PTP) phenotypes present in dmGluRA-null animals. Null dfmr1 mutants show features of synaptic hyperexcitability, including multiple transmission events in response to a single stimulus and cyclic modulation of transmission amplitude during prolonged HFS. The double null dfmr1;dmGluRA mutant shows amelioration of these defects but does not fully restore wildtype properties in dfmr1-null animals. These data suggest that dmGluRA functions in a negative feedback loop in which excess glutamate released during high-frequency transmission binds the glutamate receptor to dampen synaptic excitability, and dFMRP functions to suppress the translation of proteins regulating this synaptic excitability. Removal of the translational regulator partially compensates for loss of the receptor and, similarly, loss of the receptor weakly compensates for loss of the translational regulator.

Etomidate blocks LTP and impairs learning but does not enhance tonic inhibition in mice carrying the N265M point mutation in the beta3 subunit of the GABAA receptor

PubMed Central

Oh, I; Rau, V; Lor, C; Laha, KT; Jurd, R; Rudolph, U; Eger, EI; Pearce, RA

2015-01-01

Enhancement of tonic inhibition mediated by extrasynaptic α5-subunit containing GABAA receptors (GABAARs) has been proposed as the mechanism by which a variety of anesthetics, including the general anesthetic etomidate, impair learning and memory. Since α5 subunits preferentially partner with β3 subunits, we tested the hypothesis that etomidate acts through β3-subunit containing GABAARs to enhance tonic inhibition, block LTP, and impair memory. We measured the effects of etomidate in wild type mice and in mice carrying a point mutation in the GABAAR β3-subunit (β3-N265M) that renders these receptors insensitive to etomidate. Etomidate enhanced tonic inhibition in CA1 pyramidal cells of the hippocampus in wild type but not in mutant mice, demonstrating that tonic inhibition is mediated by β3-subunit containing GABAARs. However, despite its inability to enhance tonic inhibition, etomidate did block LTP in brain slices from mutant mice as well as in those from wild type mice. Etomidate also impaired fear conditioning to context, with no differences between genotypes. In studies of recombinant receptors expressed in HEK293 cells, α5β1γ2L GABAARs were insensitive to amnestic concentrations of etomidate (1 [.proportional]M and below), whereas α5β2γ2L and α5β3γ2L GABAARs were enhanced. We conclude that etomidate enhances tonic inhibition in pyramidal cells through its action on α5β3-containing GABAA receptors, but blocks LTP and impairs learning by other means - most likely by modulating α5β2-containing GABAA receptors. The critical anesthetic targets underlying amnesia might include other forms of inhibition imposed on pyramidal neurons (e.g. slow phasic inhibition), or inhibitory processes on non-pyramidal cells (e.g. interneurons). PMID:25680234

Endogenous Positive Allosteric Modulation of GABAA Receptors by Diazepam binding inhibitor

PubMed Central

Christian, Catherine A.; Herbert, Anne G.; Holt, Rebecca L.; Peng, Kathy; Sherwood, Kyla D.; Pangratz-Fuehrer, Susanne; Rudolph, Uwe; Huguenard, John R.

2014-01-01

Summary Benzodiazepines (BZs) allosterically modulate γ-aminobutyric acid type-A receptors (GABAARs) to increase inhibitory synaptic strength. Diazepam binding inhibitor (DBI) protein is a BZ site ligand expressed endogenously in the brain, but functional evidence for BZ-mimicking positive modulatory actions has been elusive. We demonstrate an endogenous potentiation of GABAergic synaptic transmission and responses to GABA uncaging in the thalamic reticular nucleus (nRT) that is absent in both nm1054 mice, in which the Dbi gene is deleted, and mice in which BZ binding to α3 subunit-containing GABAARs is disrupted. Viral transduction of DBI into nRT is sufficient to rescue the endogenous potentiation of GABAergic transmission in nm1054 mice. Both mutations enhance thalamocortical spike-and-wave discharges characteristic of absence epilepsy. Together these results indicate that DBI mediates endogenous nucleus-specific BZ-mimicking (“endozepine”) roles to modulate nRT function and suppress thalamocortical oscillations. Enhanced DBI signaling might serve as a novel therapy for epilepsy and other neurological disorders. PMID:23727119

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

Increased anxiety and synaptic plasticity in estrogen receptor -deficient mice

NASA Astrophysics Data System (ADS)

Krel, Wojciech; Dupont, Sonia; Krust, Andrée; Chambon, Pierre; Chapman, Paul F.

2001-10-01

Estrogens are powerful modulators of neuronal physiology and in humans may affect a broad range of functions, including reproductive, emotional, and cognitive behaviors. We studied the contribution of estrogen receptors (ERs) in modulation of emotional processes and analyzed the effects of deleting ER or ER in mice. Behavior consistent with increased anxiety was observed principally in ER mutant females and was associated with a reduced threshold for the induction of synaptic plasticity in the basolateral amygdala. Local increase of 5-hydroxytryptamine 1a receptor expression inmedial amygdala may contribute to these changes. Our data show that, particularly in females, there is an important role for ERβ-mediated estrogen signaling in the processing of emotional behavior.

Dynamic DNA Methylation Controls Glutamate Receptor Trafficking and Synaptic Scaling

PubMed Central

Sweatt, J. David

2016-01-01

Hebbian plasticity, including LTP and LTD, has long been regarded as important for local circuit refinement in the context of memory formation and stabilization. However, circuit development and stabilization additionally relies on non-Hebbian, homoeostatic, forms of plasticity such as synaptic scaling. Synaptic scaling is induced by chronic increases or decreases in neuronal activity. Synaptic scaling is associated with cell-wide adjustments in postsynaptic receptor density, and can occur in a multiplicative manner resulting in preservation of relative synaptic strengths across the entire neuron's population of synapses. Both active DNA methylation and de-methylation have been validated as crucial regulators of gene transcription during learning, and synaptic scaling is known to be transcriptionally dependent. However, it has been unclear whether homeostatic forms of plasticity such as synaptic scaling are regulated via epigenetic mechanisms. This review describes exciting recent work that has demonstrated a role for active changes in neuronal DNA methylation and demethylation as a controller of synaptic scaling and glutamate receptor trafficking. These findings bring together three major categories of memory-associated mechanisms that were previously largely considered separately: DNA methylation, homeostatic plasticity, and glutamate receptor trafficking. PMID:26849493

Synaptic Efficacy as a Function of Ionotropic Receptor Distribution: A Computational Study

PubMed Central

Allam, Sushmita L.; Bouteiller, Jean-Marie C.; Hu, Eric Y.; Ambert, Nicolas; Greget, Renaud; Bischoff, Serge; Baudry, Michel; Berger, Theodore W.

2015-01-01

Glutamatergic synapses are the most prevalent functional elements of information processing in the brain. Changes in pre-synaptic activity and in the function of various post-synaptic elements contribute to generate a large variety of synaptic responses. Previous studies have explored postsynaptic factors responsible for regulating synaptic strength variations, but have given far less importance to synaptic geometry, and more specifically to the subcellular distribution of ionotropic receptors. We analyzed the functional effects resulting from changing the subsynaptic localization of ionotropic receptors by using a hippocampal synaptic computational framework. The present study was performed using the EONS (Elementary Objects of the Nervous System) synaptic modeling platform, which was specifically developed to explore the roles of subsynaptic elements as well as their interactions, and that of synaptic geometry. More specifically, we determined the effects of changing the localization of ionotropic receptors relative to the presynaptic glutamate release site, on synaptic efficacy and its variations following single pulse and paired-pulse stimulation protocols. The results indicate that changes in synaptic geometry do have consequences on synaptic efficacy and its dynamics. PMID:26480028

Synaptic Efficacy as a Function of Ionotropic Receptor Distribution: A Computational Study.

PubMed

Allam, Sushmita L; Bouteiller, Jean-Marie C; Hu, Eric Y; Ambert, Nicolas; Greget, Renaud; Bischoff, Serge; Baudry, Michel; Berger, Theodore W

2015-01-01

Glutamatergic synapses are the most prevalent functional elements of information processing in the brain. Changes in pre-synaptic activity and in the function of various post-synaptic elements contribute to generate a large variety of synaptic responses. Previous studies have explored postsynaptic factors responsible for regulating synaptic strength variations, but have given far less importance to synaptic geometry, and more specifically to the subcellular distribution of ionotropic receptors. We analyzed the functional effects resulting from changing the subsynaptic localization of ionotropic receptors by using a hippocampal synaptic computational framework. The present study was performed using the EONS (Elementary Objects of the Nervous System) synaptic modeling platform, which was specifically developed to explore the roles of subsynaptic elements as well as their interactions, and that of synaptic geometry. More specifically, we determined the effects of changing the localization of ionotropic receptors relative to the presynaptic glutamate release site, on synaptic efficacy and its variations following single pulse and paired-pulse stimulation protocols. The results indicate that changes in synaptic geometry do have consequences on synaptic efficacy and its dynamics.

Early-life seizures alter synaptic calcium-permeable AMPA receptor function and plasticity

PubMed Central

Lippman-Bell, Jocelyn J.; Zhou, Chengwen; Sun, Hongyu; Feske, Joel S.; Jensen, Frances E.

2016-01-01

Calcium (Ca2+)-mediated1 signaling pathways are critical to synaptic plasticity. In adults, the NMDA glutamate receptor (NMDAR) represents a major route for activity-dependent synaptic Ca2+ entry. However, during neonatal development, when synaptic plasticity is high, many AMPA glutamate receptors (AMPARs) are also permeable to Ca2+ (CP-AMPAR) due to low GluA2 subunit expression, providing an additional route for activity- and glutamate-dependent Ca2+ influx and subsequent signaling. Therefore, altered hippocampal Ca2+ signaling may represent an age-specific pathogenic mechanism. We thus aimed to assess Ca2+ responses 48 hours after hypoxia-induced neonatal seizures (HS) in postnatal day (P)10 rats, a post-seizure time point at which we previously reported LTP attenuation. We found that Ca2+ responses were higher in brain slices from post-HS rats than in controls and this increase was CP-AMPAR-dependent. To determine whether synaptic CP-AMPAR expression was also altered post-HS, we assessed the expression of GluA2 at hippocampal synapses and the expression of long-term depression (LTD), which has been linked to the presence of synaptic GluA2. Here we report a decrease 48 hours after HS in synaptic GluA2 expression at synapses and LTD in hippocampal CA1. Given the potentially critical role of AMPAR trafficking in disease progression, we aimed to establish whether post-seizure in vivo AMPAR antagonist treatment prevented the enhanced Ca2+ responses, changes in GluA2 synaptic expression, and diminished LTD. We found that NBQX treatment prevents all three of these post-seizure consequences, further supporting a critical role for AMPARs as an age-specific therapeutic target. PMID:27521497

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

Aniracetam reduces glutamate receptor desensitization and slows the decay of fast excitatory synaptic currents in the hippocampus.

PubMed Central

Isaacson, J S; Nicoll, R A

1991-01-01

Aniracetam is a nootropic drug that has been shown to selectively enhance quisqualate receptor-mediated responses in Xenopus oocytes injected with brain mRNA and in hippocampal pyramidal cells [Ito, I., Tanabe, S., Kohda, A. & Sugiyama, H. (1990) J. Physiol. (London) 424, 533-544]. We have used patch clamp recording techniques in hippocampal slices to elucidate the mechanism for this selective action. We find that aniracetam enhances glutamate-evoked currents in whole-cell recordings and, in outside-out patches, strongly reduces glutamate receptor desensitization. In addition, aniracetam selectively prolongs the time course and increases the peak amplitude of fast synaptic currents. These findings indicate that aniracetam slows the kinetics of fast synaptic transmission and are consistent with the proposal [Trussell, L. O. & Fischbach, G. D. (1989) Neuron 3, 209-218; Tang, C.-M., Dichter, M. & Morad, M. (1989) Science 243, 1474-1477] that receptor desensitization governs the strength of fast excitatory synaptic transmission in the brain. PMID:1660156

Aniracetam reduces glutamate receptor desensitization and slows the decay of fast excitatory synaptic currents in the hippocampus.

PubMed

Isaacson, J S; Nicoll, R A

1991-12-01

Aniracetam is a nootropic drug that has been shown to selectively enhance quisqualate receptor-mediated responses in Xenopus oocytes injected with brain mRNA and in hippocampal pyramidal cells [Ito, I., Tanabe, S., Kohda, A. & Sugiyama, H. (1990) J. Physiol. (London) 424, 533-544]. We have used patch clamp recording techniques in hippocampal slices to elucidate the mechanism for this selective action. We find that aniracetam enhances glutamate-evoked currents in whole-cell recordings and, in outside-out patches, strongly reduces glutamate receptor desensitization. In addition, aniracetam selectively prolongs the time course and increases the peak amplitude of fast synaptic currents. These findings indicate that aniracetam slows the kinetics of fast synaptic transmission and are consistent with the proposal [Trussell, L. O. & Fischbach, G. D. (1989) Neuron 3, 209-218; Tang, C.-M., Dichter, M. & Morad, M. (1989) Science 243, 1474-1477] that receptor desensitization governs the strength of fast excitatory synaptic transmission in the brain.

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

The influence of synaptic size on AMPA receptor activation: a Monte Carlo model.

PubMed

Montes, Jesus; Peña, Jose M; DeFelipe, Javier; Herreras, Oscar; Merchan-Perez, Angel

2015-01-01

Physiological and electron microscope studies have shown that synapses are functionally and morphologically heterogeneous and that variations in size of synaptic junctions are related to characteristics such as release probability and density of postsynaptic AMPA receptors. The present article focuses on how these morphological variations impact synaptic transmission. We based our study on Monte Carlo computational simulations of simplified model synapses whose morphological features have been extracted from hundreds of actual synaptic junctions reconstructed by three-dimensional electron microscopy. We have examined the effects that parameters such as synaptic size or density of AMPA receptors have on the number of receptors that open after release of a single synaptic vesicle. Our results indicate that the maximum number of receptors that will open after the release of a single synaptic vesicle may show a ten-fold variation in the whole population of synapses. When individual synapses are considered, there is also a stochastical variability that is maximal in small synapses with low numbers of receptors. The number of postsynaptic receptors and the size of the synaptic junction are the most influential parameters, while the packing density of receptors or the concentration of extrasynaptic transporters have little or no influence on the opening of AMPA receptors.

The Influence of Synaptic Size on AMPA Receptor Activation: A Monte Carlo Model

PubMed Central

Montes, Jesus; Peña, Jose M.; DeFelipe, Javier; Herreras, Oscar; Merchan-Perez, Angel

2015-01-01

Physiological and electron microscope studies have shown that synapses are functionally and morphologically heterogeneous and that variations in size of synaptic junctions are related to characteristics such as release probability and density of postsynaptic AMPA receptors. The present article focuses on how these morphological variations impact synaptic transmission. We based our study on Monte Carlo computational simulations of simplified model synapses whose morphological features have been extracted from hundreds of actual synaptic junctions reconstructed by three-dimensional electron microscopy. We have examined the effects that parameters such as synaptic size or density of AMPA receptors have on the number of receptors that open after release of a single synaptic vesicle. Our results indicate that the maximum number of receptors that will open after the release of a single synaptic vesicle may show a ten-fold variation in the whole population of synapses. When individual synapses are considered, there is also a stochastical variability that is maximal in small synapses with low numbers of receptors. The number of postsynaptic receptors and the size of the synaptic junction are the most influential parameters, while the packing density of receptors or the concentration of extrasynaptic transporters have little or no influence on the opening of AMPA receptors. PMID:26107874

Laser photolysis of DPNI-GABA, a tool for investigating the properties and distribution of GABA receptors and for silencing neurons in situ.

PubMed

Trigo, Federico F; Papageorgiou, George; Corrie, John E T; Ogden, David

2009-07-30

Laser photolysis to release GABA at precisely defined times and locations permits investigation of the distribution of functional GABA(A) receptors in neuronal compartments, the activation kinetics and pharmacology of GABA(A) receptors in situ, and the role of individual neurons in neural circuits by selective silencing with low GABA concentrations. We describe the experimental evaluation and applications of a new nitroindoline-caged GABA, DPNI-GABA, modified to minimize the pharmacological interference commonly found with caged GABA reagents, but retaining the advantages of nitroindoline cages. Unlike the 5-methoxycarbonylmethyl-7-nitroindolinyl-GABA tested previously, DPNI-GABA inhibited GABA(A) receptors with much lower affinity, reducing peak GABA-evoked responses with an IC(50) of approximately 0.5 mM. Most importantly, the kinetics of receptor activation, determined as 10-90% rise-times, were comparable to synaptic events and were little affected by DPNI-GABA present at 1mM concentration, permitting photolysis of DPNI-GABA to mimic synaptic activation of GABA(A) receptors. With a laser spot of 1 microm applied to cerebellar molecular layer interneurons, the spatial resolution of uncaging DPNI-GABA in dendrites was estimated as 2 microm laterally and 7.5 microm focally. Finally, at low DPNI-GABA concentration, photorelease restricted to the area of the soma suppressed spiking in single Purkinje neurons or molecular layer interneurons for periods controlled by the flash intensity and duration. DPNI-GABA has properties better adapted for fast kinetic studies with laser photolysis at GABA(A) receptors than previously reported caged GABA reagents, and can be used in experiments where spatial resolution is determined by the dimensions of the laser light spot.

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

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.

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.

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.

Potentiation of Schaffer-Collateral CA1 Synaptic Transmission by eEF2K and p38 MAPK Mediated Mechanisms.

PubMed

Weng, Weiguang; Chen, Ying; Wang, Man; Zhuang, Yinghan; Behnisch, Thomas

2016-01-01

The elongation factor 2 kinase (eEF2K), likewise known as CaMKIII, has been demonstrated to be involved in antidepressant responses of NMDA receptor antagonists. Even so, it remains open whether direct inhibition of eEF2K without altering up-stream or other signaling pathways affects hippocampal synaptic transmission and neuronal network synchrony. Inhibition of eEF2K by the selective and potent eEF2K inhibitor A-484954 induced a fast pre-synaptically mediated enhancement of synaptic transmission and synchronization of neural network activity. The eEF2K-inhibition mediated potentiation of synaptic transmission of hippocampal CA1 neurons is most notably independent of protein synthesis and does not rely on protein kinase C, protein kinase A or mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase 1/2. Moreover, the strengthening of synaptic transmission in the response to the inhibition of eEF2K was strongly attenuated by the inhibition of p38 MAPK. In addition, we show the involvement of barium-sensitive and more specific the TWIK-related potassium-1 (TREK-1) channels in the eEF2K-inhibition mediated potentiation of synaptic transmission. These findings reveal a novel pathway of eEF2K mediated regulation of hippocampal synaptic transmission. Further research is required to study whether such compounds could be beneficial for the development of mood disorder treatments with a fast-acting antidepressant response.

Extracellular alpha-synuclein oligomers modulate synaptic transmission and impair LTP via NMDA-receptor activation.

PubMed

Diógenes, Maria José; Dias, Raquel B; Rombo, Diogo M; Vicente Miranda, Hugo; Maiolino, Francesca; Guerreiro, Patrícia; Näsström, Thomas; Franquelim, Henri G; Oliveira, Luís M A; Castanho, Miguel A R B; Lannfelt, Lars; Bergström, Joakim; Ingelsson, Martin; Quintas, Alexandre; Sebastião, Ana M; Lopes, Luísa V; Outeiro, Tiago Fleming

2012-08-22

Parkinson's disease (PD) is the most common representative of a group of disorders known as synucleinopathies, in which misfolding and aggregation of α-synuclein (a-syn) in various brain regions is the major pathological hallmark. Indeed, the motor symptoms in PD are caused by a heterogeneous degeneration of brain neurons not only in substantia nigra pars compacta but also in other extrastriatal areas of the brain. In addition to the well known motor dysfunction in PD patients, cognitive deficits and memory impairment are also an important part of the disorder, probably due to disruption of synaptic transmission and plasticity in extrastriatal areas, including the hippocampus. Here, we investigated the impact of a-syn aggregation on AMPA and NMDA receptor-mediated rat hippocampal (CA3-CA1) synaptic transmission and long-term potentiation (LTP), the neurophysiological basis for learning and memory. Our data show that prolonged exposure to a-syn oligomers, but not monomers or fibrils, increases basal synaptic transmission through NMDA receptor activation, triggering enhanced contribution of calcium-permeable AMPA receptors. Slices treated with a-syn oligomers were unable to respond with further potentiation to theta-burst stimulation, leading to impaired LTP. Prior delivery of a low-frequency train reinstated the ability to express LTP, implying that exposure to a-syn oligomers drives the increase of glutamatergic synaptic transmission, preventing further potentiation by physiological stimuli. Our novel findings provide mechanistic insight on how a-syn oligomers may trigger neuronal dysfunction and toxicity in PD and other synucleinopathies.

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

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

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

Quantitative Electroencephalography Within Sleep/Wake States Differentiates GABAA Modulators Eszopiclone and Zolpidem From Dual Orexin Receptor Antagonists in Rats

PubMed Central

Fox, Steven V; Gotter, Anthony L; Tye, Spencer J; Garson, Susan L; Savitz, Alan T; Uslaner, Jason M; Brunner, Joseph I; Tannenbaum, Pamela L; McDonald, Terrence P; Hodgson, Robert; Yao, Lihang; Bowlby, Mark R; Kuduk, Scott D; Coleman, Paul J; Hargreaves, Richard; Winrow, Christopher J; Renger, John J

2013-01-01

Dual orexin receptor antagonists (DORAs) induce sleep by blocking orexin 1 and orexin 2 receptor-mediated activities responsible for regulating wakefulness. DORAs represent a potential alternative mechanism to the current standard of care that includes the γ-aminobutyric acid (GABA)A receptor-positive allosteric modulators, eszopiclone and zolpidem. This work uses an innovative method to analyze electroencephalogram (EEG) spectral frequencies within sleep/wake states to differentiate the effects of GABAA modulators from DORA-22, an analog of the DORA MK-6096, in Sprague–Dawley rats. The effects of low, intermediate, and high doses of eszopiclone, zolpidem, and DORA-22 were examined after first defining each compound's ability to promote sleep during active-phase dosing. The EEG spectral frequency power within specific sleep stages was calculated in 1-Hz intervals from 1 to 100 Hz within each sleep/wake state for the first 4 h after the dose. Eszopiclone and zolpidem produced marked, dose-responsive disruptions in sleep stage-specific EEG spectral profiles compared with vehicle treatment. In marked contrast, DORA-22 exhibited marginal changes in the spectral profile, observed only during rapid eye movement sleep, and only at the highest dose tested. Moreover, while eszopiclone- and zolpidem-induced changes were evident in the inactive period, the EEG spectral responses to DORA-22 were absent during this phase. These results suggest that DORA-22 differs from eszopiclone and zolpidem whereby DORA-22 promotes somnolence without altering the neuronal network EEG activity observed during normal sleep. PMID:23722242

Sex differences in cerebellar synaptic transmission and sex-specific responses to autism-linked Gabrb3 mutations in mice.

PubMed

Mercer, Audrey A; Palarz, Kristin J; Tabatadze, Nino; Woolley, Catherine S; Raman, Indira M

2016-04-14

Neurons of the cerebellar nuclei (CbN) transmit cerebellar signals to premotor areas. The cerebellum expresses several autism-linked genes, including GABRB3, which encodes GABAA receptor β3 subunits and is among the maternal alleles deleted in Angelman syndrome. We tested how this Gabrb3 m-/p+ mutation affects CbN physiology in mice, separating responses of males and females. Wild-type mice showed sex differences in synaptic excitation, inhibition, and intrinsic properties. Relative to females, CbN cells of males had smaller synaptically evoked mGluR1/5-dependent currents, slower Purkinje-mediated IPSCs, and lower spontaneous firing rates, but rotarod performances were indistinguishable. In mutant CbN cells, IPSC kinetics were unchanged, but mutant males, unlike females, showed enlarged mGluR1/5 responses and accelerated spontaneous firing. These changes appear compensatory, since mutant males but not females performed indistinguishably from wild-type siblings on the rotarod task. Thus, sex differences in cerebellar physiology produce similar behavioral output, but provide distinct baselines for responses to mutations.

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

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.

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.

Synaptic Bistability Due to Nucleation and Evaporation of Receptor Clusters

NASA Astrophysics Data System (ADS)

Burlakov, V. M.; Emptage, N.; Goriely, A.; Bressloff, P. C.

2012-01-01

We introduce a bistability mechanism for long-term synaptic plasticity based on switching between two metastable states that contain significantly different numbers of synaptic receptors. One state is characterized by a two-dimensional gas of mobile interacting receptors and is stabilized against clustering by a high nucleation barrier. The other state contains a receptor gas in equilibrium with a large cluster of immobile receptors, which is stabilized by the turnover rate of receptors into and out of the synapse. Transitions between the two states can be initiated by either an increase (potentiation) or a decrease (depotentiation) of the net receptor flux into the synapse. This changes the saturation level of the receptor gas and triggers nucleation or evaporation of receptor clusters.

Acute suppression of spontaneous neurotransmission drives synaptic potentiation.

PubMed

Nosyreva, Elena; Szabla, Kristen; Autry, Anita E; Ryazanov, Alexey G; Monteggia, Lisa M; Kavalali, Ege T

2013-04-17

The impact of spontaneous neurotransmission on neuronal plasticity remains poorly understood. Here, we show that acute suppression of spontaneous NMDA receptor-mediated (NMDAR-mediated) neurotransmission potentiates synaptic responses in the CA1 regions of rat and mouse hippocampus. This potentiation requires protein synthesis, brain-derived neurotrophic factor expression, eukaryotic elongation factor-2 kinase function, and increased surface expression of AMPA receptors. Our behavioral studies link this same synaptic signaling pathway to the fast-acting antidepressant responses elicited by ketamine. We also show that selective neurotransmitter depletion from spontaneously recycling vesicles triggers synaptic potentiation via the same pathway as NMDAR blockade, demonstrating that presynaptic impairment of spontaneous release, without manipulation of evoked neurotransmission, is sufficient to elicit postsynaptic plasticity. These findings uncover an unexpectedly dynamic impact of spontaneous glutamate release on synaptic efficacy and provide new insight into a key synaptic substrate for rapid antidepressant action.

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.

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

Regulation of glutamate receptor internalization by the spine cytoskeleton is mediated by its PKA-dependent association with CPG2

PubMed Central

Loebrich, Sven; Djukic, Biljana; Tong, Zachary J.; Cottrell, Jeffrey R.; Turrigiano, Gina G.; Nedivi, Elly

2013-01-01

A key neuronal mechanism for adjusting excitatory synaptic strength is clathrin-mediated endocytosis of postsynaptic glutamate receptors (GluRs). The actin cytoskeleton is critical for clathrin-mediated endocytosis, yet we lack a mechanistic understanding of its interaction with the endocytic process and how it may be regulated. Here we show that F-actin in dendritic spines physically binds the synaptic nuclear envelope 1 gene product candidate plasticity gene 2 (CPG2) in a PKA-dependent manner, and that this association is required for synaptic GluR internalization. Mutating two PKA sites on CPG2 disrupts its cytoskeletal association, attenuating GluR endocytosis and affecting the efficacy of synaptic transmission in vivo. These results identify CPG2 as an F-actin binding partner that functionally mediates interaction of the spine cytoskeleton with postsynaptic endocytosis. Further, the regulation of CPG2/F-actin association by PKA provides a gateway for cellular control of synaptic receptor internalization through second messenger signaling pathways. Recent identification of human synaptic nuclear envelope 1 as a risk locus for bipolar disorder suggests that CPG2 could play a role in synaptic dysfunction underlying neuropsychiatric disease. PMID:24191017

Modulation of transient receptor potential vanilloid 4-mediated membrane currents and synaptic transmission by protein kinase C

PubMed Central

Cao, De-Shou; Yu, Shuang-Quan; Premkumar, Louis S

2009-01-01

Background Transient receptor potential Vanilloid (TRPV) receptors are involved in nociception and are expressed predominantly in sensory neurons. TRPV1, a non-selective cation channel has been extensively studied and is responsible for inflammatory thermal hypersensitivity. In this study, the expression and function of TRPV4 have been characterized and compared with those of TRPV1. Results Immunohistochemical studies revealed that both TRPV1 and TRPV4 were co-expressed in dorsal root ganglion (DRG) neuronal cell bodies and in the central terminals of laminae I and II of the spinal dorsal horn (DH). In Ca2+ fluorescence imaging and whole-cell patch-clamp experiments, TRPV1- and TRPV4-mediated responses were observed in a population of the same DRG neurons. Sensitization of TRPV1 has been shown to be involved in inflammatory pain conditions. Incubation with phorbol 12, 13-dibutyrate (PDBu), a PKC activator, resulted in a significant potentiation of TRPV4 currents in DRG neurons. In TRPV4 expressing HEK 293T cells, PDBu increased 4α-phorbol 12, 13-didecanoate (4α-PDD)-induced single-channel activity in cell-attached patches, which was abrogated by bisindolylmaleimide (BIM), a selective PKC inhibitor. TRPV4 is also expressed at the central terminals of sensory neurons. Activation of TRPV4 by 4α-PDD increased the frequency of miniature excitatory post synaptic currents (mEPSCs) in DRG-DH neuronal co-cultures. 4α-PDD-induced increase in the frequency of mEPSCs was further enhanced by PDBu. The expression of TRP channels has been shown in other areas of the CNS; application of 4α-PDD significantly increased the mEPSC frequency in cultured hippocampal neurons, which was further potentiated by PDBu, whereas, TRPV1 agonist capsaicin did not modulate synaptic transmission. Conclusion These results indicate that TRPV4 and TRPV1 are co-expressed in certain DRG neurons and TRPV4 can be sensitized by PKC not only in DRG neuronal cell bodies, but also in the central sensory

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.

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

Spinal Endocannabinoids and CB1 Receptors Mediate C-Fiber-Induced Heterosynaptic Pain Plasticity

PubMed Central

Pernía-Andrade, Alejandro J.; Kato, Ako; Witschi, Robert; Nyilas, Rita; Katona, István; Freund, Tamás F.; Watanabe, Masahiko; Filitz, Jörg; Koppert, Wolfgang; Schüttler, Jürgen; Ji, Guangchen; Neugebauer, Volker; Marsicano, Giovanni; Lutz, Beat; Vanegas, Horacio; Zeilhofer, Hanns Ulrich

2010-01-01

Diminished synaptic inhibition in the spinal dorsal horn is a major contributor to chronic pain. Pathways, which reduce synaptic inhibition in inflammatory and neuropathic pain states, have been identified, but central hyperalgesia and diminished dorsal horn synaptic inhibition also occur in the absence of inflammation or neuropathy, solely triggered by intense nociceptive (C–fiber) input to the spinal dorsal horn. We found that endocannabinoids produced upon strong nociceptive stimulation activated CB1 receptors on inhibitory dorsal horn neurons to reduce the synaptic release of GABA and glycine and thus rendered nociceptive neurons excitable by non-painful stimuli. Spinal endocannabinoids and CB1 receptors on inhibitory dorsal horn interneurons act as mediators of heterosynaptic pain sensitization and play an unexpected role in dorsal horn pain controlling circuits. PMID:19661434

The laminar organization of the Drosophila ellipsoid body is semaphorin-dependent and prevents the formation of ectopic synaptic connections

PubMed Central

Xie, Xiaojun; Tabuchi, Masashi; Brown, Matthew P; Mitchell, Sarah P; Wu, Mark N; Kolodkin, Alex L

2017-01-01

The ellipsoid body (EB) in the Drosophila brain is a central complex (CX) substructure that harbors circumferentially laminated ring (R) neuron axons and mediates multifaceted sensory integration and motor coordination functions. However, what regulates R axon lamination and how lamination affects R neuron function remain unknown. We show here that the EB is sequentially innervated by small-field and large-field neurons and that early developing EB neurons play an important regulatory role in EB laminae formation. The transmembrane proteins semaphorin-1a (Sema-1a) and plexin A function together to regulate R axon lamination. R neurons recruit both GABA and GABA-A receptors to their axon terminals in the EB, and optogenetic stimulation coupled with electrophysiological recordings show that Sema-1a-dependent R axon lamination is required for preventing the spread of synaptic inhibition between adjacent EB lamina. These results provide direct evidence that EB lamination is critical for local pre-synaptic inhibitory circuit organization. DOI: http://dx.doi.org/10.7554/eLife.25328.001 PMID:28632130

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.

Context-dependent modulation of alphabetagamma and alphabetadelta GABA A receptors by penicillin: implications for phasic and tonic inhibition.

PubMed

Feng, Hua-Jun; Botzolakis, Emmanuel J; Macdonald, Robert L

2009-01-01

Penicillin, an open-channel blocker of GABA(A) receptors, was recently reported to inhibit phasic, but not tonic, currents in hippocampal neurons. To distinguish between isoform-specific and context-dependent modulation as possible explanations for this selectivity, the effects of penicillin were evaluated on recombinant GABA(A) receptors expressed in HEK293T cells. When co-applied with saturating GABA, penicillin decreased peak amplitude, induced rebound, and prolonged deactivation of currents evoked from both synaptic and extrasynaptic receptor isoforms. However, penicillin had isoform-specific effects on the extent of desensitization, reflecting its ability to differentially modulate peak (non-equilibrium) and residual (near-equilibrium) currents. This suggested that the context of activation could determine the apparent sensitivity of a given receptor isoform to penicillin. To test this hypothesis, we explored the ability of penicillin to modulate synaptic and extrasynaptic isoform currents that were activated under more physiologically relevant conditions. Interestingly, while currents evoked from synaptic isoforms under phasic conditions (transient activation by a saturating concentration of GABA) were substantially inhibited by penicillin, currents evoked from extrasynaptic isoforms under tonic conditions (prolonged application by a sub-saturating concentration of GABA) were minimally affected. We therefore concluded that the reported inability of penicillin to modulate tonic currents could not simply be attributed to insensitivity of extrasynaptic receptors, but rather, reflected an inability to modulate these receptors in their native context of activation.

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

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.

EphA4 Activation of c-Abl Mediates Synaptic Loss and LTP Blockade Caused by Amyloid-β Oligomers

PubMed Central

M. Vargas, Lina; Leal, Nancy; Estrada, Lisbell D.; González, Adrian; Serrano, Felipe; Araya, Katherine; Gysling, Katia; Inestrosa, Nibaldo C.; Pasquale, Elena B.; Alvarez, Alejandra R.

2014-01-01

The early stages of Alzheimer's disease are characterised by impaired synaptic plasticity and synapse loss. Here, we show that amyloid-β oligomers (AβOs) activate the c-Abl kinase in dendritic spines of cultured hippocampal neurons and that c-Abl kinase activity is required for AβOs-induced synaptic loss. We also show that the EphA4 receptor tyrosine kinase is upstream of c-Abl activation by AβOs. EphA4 tyrosine phosphorylation (activation) is increased in cultured neurons and synaptoneurosomes exposed to AβOs, and in Alzheimer-transgenic mice brain. We do not detect c-Abl activation in EphA4-knockout neurons exposed to AβOs. More interestingly, we demonstrate EphA4/c-Abl activation is a key-signalling event that mediates the synaptic damage induced by AβOs. According to this results, the EphA4 antagonistic peptide KYL and c-Abl inhibitor STI prevented i) dendritic spine reduction, ii) the blocking of LTP induction and iii) neuronal apoptosis caused by AβOs. Moreover, EphA4-/- neurons or sh-EphA4-transfected neurons showed reduced synaptotoxicity by AβOs. Our results are consistent with EphA4 being a novel receptor that mediates synaptic damage induced by AβOs. EphA4/c-Abl signalling could be a relevant pathway involved in the early cognitive decline observed in Alzheimer's disease patients. PMID:24658113

Distinct kinetics of inhibitory currents in thalamocortical neurons that arise from dendritic or axonal origin.

PubMed

Yang, Sunggu; Govindaiah, Gubbi; Lee, Sang-Hun; Yang, Sungchil; Cox, Charles L

2017-01-01

Thalamocortical neurons in the dorsal lateral geniculate nucleus (dLGN) transfer visual information from retina to primary visual cortex. This information is modulated by inhibitory input arising from local interneurons and thalamic reticular nucleus (TRN) neurons, leading to alterations of receptive field properties of thalamocortical neurons. Local GABAergic interneurons provide two distinct synaptic outputs: axonal (F1 terminals) and dendritic (F2 terminals) onto dLGN thalamocortical neurons. By contrast, TRN neurons provide only axonal output (F1 terminals) onto dLGN thalamocortical neurons. It is unclear if GABAA receptor-mediated currents originating from F1 and F2 terminals have different characteristics. In the present study, we examined multiple characteristics (rise time, slope, halfwidth and decay τ) of GABAA receptor-mediated miniature inhibitory postsynaptic synaptic currents (mIPSCs) originating from F1 and F2 terminals. The mIPSCs arising from F2 terminals showed slower kinetics relative to those from F1 terminals. Such differential kinetics of GABAAR-mediated responses could be an important role in temporal coding of visual signals.

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.

The differential role of α1- and α5-containing GABAA receptors in mediating diazepam effects on spontaneous locomotor activity and water-maze learning and memory in rats

PubMed Central

Savić, Miroslav M.; Milinković, Marija M.; Rallapalli, Sundari; Clayton, Terry; Joksimović, Srðan; Van Linn, Michael; Cook, James M.

2009-01-01

The clinical use of benzodiazepines (BZs) is hampered by sedation and cognitive deterioration. Although genetic and pharmacological studies suggest that α1- and α5-containing GABAA receptors mediate and/or modulate these effects, their molecular substrate is not fully elucidated. By the use of two selective ligands : the α1-subunit affinity-selective antagonist β-CCt, and the α5-subunit affinity- and efficacy-selective antagonist XLi093, we examined the mechanisms of behavioural effects of diazepam in the tests of spontaneous locomotor activity and water-maze acquisition and recall, the two paradigms indicative of sedative- and cognition-impairing effects of BZs, respectively. The locomotor-activity decreasing propensity of diazepam (significant at 1.5 and 5 mg/kg) was antagonized by β-CCt (5 and 15 mg/kg), while it tended to be potentiated by XLi093 in doses of 10 mg/kg, and especially 20 mg/kg. Diazepam decreased acquisition and recall in the water maze, with a minimum effective dose of 1.5 mg/kg. Both antagonists reversed the thigmotaxis induced by 2 mg/kg diazepam throughout the test, suggesting that both GABAA receptor subtypes participate in BZ effects on the procedural component of the task. Diazepam-induced impairment in the declarative component of the task, as assessed by path efficiency, the latency and distance before finding the platform across acquisition trials, and also by the spatial parameters in the probe trial, was partially prevented by both, 15 mg/kg β-CCt and 10 mg/kg XLi093. Combining a BZ with β-CCt results in the near to control level of performance of a cognitive task, without sedation, and may be worth testing on human subjects. PMID:19265570

Synaptic activity regulates AMPA receptor trafficking through different recycling pathways

PubMed Central

Zheng, Ning; Jeyifous, Okunola; Munro, Charlotte; Montgomery, Johanna M; Green, William N

2015-01-01

Changes in glutamatergic synaptic strength in brain are dependent on AMPA-type glutamate receptor (AMPAR) recycling, which is assumed to occur through a single local pathway. In this study, we present evidence that AMPAR recycling occurs through different pathways regulated by synaptic activity. Without synaptic stimulation, most AMPARs recycled in dynamin-independent endosomes containing the GTPase, Arf6. Few AMPARs recycled in dynamin-dependent endosomes labeled by transferrin receptors (TfRs). AMPAR recycling was blocked by alterations in the GTPase, TC10, which co-localized with Arf6 endosomes. TC10 mutants that reduced AMPAR recycling had no effect on increased AMPAR levels with long-term potentiation (LTP) and little effect on decreased AMPAR levels with long-term depression. However, internalized AMPAR levels in TfR-containing recycling endosomes increased after LTP, indicating increased AMPAR recycling through the dynamin-dependent pathway with synaptic plasticity. LTP-induced AMPAR endocytosis is inconsistent with local recycling as a source of increased surface receptors, suggesting AMPARs are trafficked from other sites. DOI: http://dx.doi.org/10.7554/eLife.06878.001 PMID:25970033

Endocannabinoid signaling and synaptic function

PubMed Central

Castillo, Pablo E.; Younts, Thomas J.; Chávez, Andrés E.; Hashimotodani, Yuki

2012-01-01

Endocannabinoids are key modulators of synaptic function. By activating cannabinoid receptors expressed in the central nervous system, these lipid messengers can regulate several neural functions and behaviors. As experimental tools advance, the repertoire of known endocannabinoid-mediated effects at the synapse, and their underlying mechanism, continues to expand. Retrograde signaling is the principal mode by which endocannabinoids mediate short- and long-term forms of plasticity at both excitatory and inhibitory synapses. However, growing evidence suggests that endocannabinoids can also signal in a non-retrograde manner. In addition to mediating synaptic plasticity, the endocannabinoid system is itself subject to plastic changes. Multiple points of interaction with other neuromodulatory and signaling systems have now been identified. Synaptic endocannabinoid signaling is thus mechanistically more complex and diverse than originally thought. In this review, we focus on new advances in endocannabinoid signaling and highlight their role as potent regulators of synaptic function in the mammalian brain. PMID:23040807

Endocannabinoid signaling and synaptic function.

PubMed

Castillo, Pablo E; Younts, Thomas J; Chávez, Andrés E; Hashimotodani, Yuki

2012-10-04

Endocannabinoids are key modulators of synaptic function. By activating cannabinoid receptors expressed in the central nervous system, these lipid messengers can regulate several neural functions and behaviors. As experimental tools advance, the repertoire of known endocannabinoid-mediated effects at the synapse, and their underlying mechanism, continues to expand. Retrograde signaling is the principal mode by which endocannabinoids mediate short- and long-term forms of plasticity at both excitatory and inhibitory synapses. However, growing evidence suggests that endocannabinoids can also signal in a nonretrograde manner. In addition to mediating synaptic plasticity, the endocannabinoid system is itself subject to plastic changes. Multiple points of interaction with other neuromodulatory and signaling systems have now been identified. In this Review, we focus on new advances in synaptic endocannabinoid signaling in the mammalian brain. The emerging picture not only reinforces endocannabinoids as potent regulators of synaptic function but also reveals that endocannabinoid signaling is mechanistically more complex and diverse than originally thought. Copyright © 2012 Elsevier Inc. All rights reserved.

Antagonism of GABA-B but not GABA-A receptors in the VTA prevents stress- and intra-VTA CRF-induced reinstatement of extinguished cocaine seeking in rats

PubMed Central

Blacktop, Jordan M.; Vranjkovic, Oliver; Mayer, Matthieu; Van Hoof, Matthew; Baker, David A.; Mantsch, John R.

2015-01-01

Stress-induced reinstatement of cocaine seeking requires corticotropin releasing factor (CRF) actions in the ventral tegmental area (VTA). However the mechanisms through which CRF regulates VTA function to promote cocaine use are not fully understood. Here we examined the role of GABAergic neurotransmission in the VTA mediated by GABA-A or GABA-B receptors in the reinstatement of extinguished cocaine seeking by a stressor, uncontrollable intermittent footshock, or bilateral intra-VTA administration of CRF. Rats underwent repeated daily cocaine self-administration (1.0 mg/kg/ing; 14 × 6 hrs/day) and extinction and were tested for reinstatement in response to footshock (0.5 mA, 0.5” duration, average every 40 sec; range 10–70 sec) or intra-VTA CRF delivery (500 ng/side) following intra-VTA pretreatment with the GABA-A antagonist, bicuculline, the GABA-B antagonist, 2-hydroxysaclofen or vehicle. Intra-VTA bicuculline (1, 10 or 20 ng/side) failed to block footshock- or CRF-induced cocaine seeking at either dose tested. By contrast, 2-hydroxysaclofen (0.2 or 2 µg/side) prevented reinstatement by both footshock and intra-VTA CRF at a concentration that failed to attenuate food-reinforced lever pressing (45 mg sucrose-sweetened pellets; FR4 schedule) in a separate group of rats. These data suggest that GABA-B receptor-dependent CRF actions in the VTA mediate stress-induced cocaine seeking and that GABA-B receptor antagonists may have utility for the management of stress-induced relapse in cocaine addicts. PMID:26596556

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

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

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

GABA, its receptors, and GABAergic inhibition in mouse taste buds.

PubMed

Dvoryanchikov, Gennady; Huang, Yijen A; Barro-Soria, Rene; Chaudhari, Nirupa; Roper, Stephen D

2011-04-13

Taste buds consist of at least three principal cell types that have different functions in processing gustatory signals: glial-like (type I) cells, receptor (type II) cells, and presynaptic (type III) cells. Using a combination of Ca2+ imaging, single-cell reverse transcriptase-PCR and immunostaining, we show that GABA is an inhibitory transmitter in mouse taste buds, acting on GABA(A) and GABA(B) receptors to suppress transmitter (ATP) secretion from receptor cells during taste stimulation. Specifically, receptor cells express GABA(A) receptor subunits β2, δ, and π, as well as GABA(B) receptors. In contrast, presynaptic cells express the GABA(A) β3 subunit and only occasionally GABA(B) receptors. In keeping with the distinct expression pattern of GABA receptors in presynaptic cells, we detected no GABAergic suppression of transmitter release from presynaptic cells. We suggest that GABA may serve function(s) in taste buds in addition to synaptic inhibition. Finally, we also defined the source of GABA in taste buds: GABA is synthesized by GAD65 in type I taste cells as well as by GAD67 in presynaptic (type III) taste cells and is stored in both those two cell types. We conclude that GABA is an inhibitory transmitter released during taste stimulation and possibly also during growth and differentiation of taste buds.

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

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

Virus-mediated swapping of zolpidem-insensitive with zolpidem-sensitive GABA(A) receptors in cortical pyramidal cells.

PubMed

Sumegi, Mate; Fukazawa, Yugo; Matsui, Ko; Lorincz, Andrea; Eyre, Mark D; Nusser, Zoltan; Shigemoto, Ryuichi

2012-04-01

Recently developed pharmacogenetic and optogenetic approaches, with their own advantages and disadvantages, have become indispensable tools in modern neuroscience. Here, we employed a previously described knock-in mouse line (GABA(A)Rγ2(77I)lox) in which the γ2 subunit of the GABA(A) receptor (GABA(A)R) was mutated to become zolpidem insensitive (γ2(77I)) and used viral vectors to swap γ2(77I) with wild-type, zolpidem-sensitive γ2 subunits (γ2(77F)). The verification of unaltered density and subcellular distribution of the virally introduced γ2 subunits requires their selective labelling. For this we generated six N- and six C-terminal-tagged γ2 subunits, with which cortical cultures of GABA(A)Rγ2(−/−) mice were transduced using lentiviruses. We found that the N-terminal AU1 tag resulted in excellent immunodetection and unimpaired synaptic localization. Unaltered kinetic properties of the AU1-tagged γ2 ((AU1)γ2(77F)) channels were demonstrated with whole-cell patch-clamp recordings of spontaneous IPSCs from cultured cells. Next, we carried out stereotaxic injections of lenti- and adeno-associated viruses containing Cre-recombinase and the (AU1)γ2(77F) subunit (Cre-2A-(AU1)γ2(77F)) into the neocortex of GABA(A)Rγ2(77I)lox mice. Light microscopic immunofluorescence and electron microscopic freeze-fracture replica immunogold labelling demonstrated the efficient immunodetection of the AU1 tag and the normal enrichment of the (AU1)γ2(77F) subunits in perisomatic GABAergic synapses. In line with this,miniature and action potential-evoked IPSCs whole-cell recorded from transduced cells had unaltered amplitudes, kinetics and restored zolpidem sensitivity. Our results obtained with a wide range of structural and functional verification methods reveal unaltered subcellular distributions and functional properties of γ2(77I) and (AU1)γ2(77F) GABA(A)Rs in cortical pyramidal cells. This transgenic–viral pharmacogenetic approach has the advantage that it

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

γ-Aminobutyric Acid Type A Receptor Potentiation Inhibits Learning in a Computational Network Model.

PubMed

Storer, Kingsley P; Reeke, George N

2018-04-17

Propofol produces memory impairment at concentrations well below those abolishing consciousness. Episodic memory, mediated by the hippocampus, is most sensitive. Two potentially overlapping scenarios may explain how γ-aminobutyric acid receptor type A (GABAA) potentiation by propofol disrupts episodic memory-the first mediated by shifting the balance from excitation to inhibition while the second involves disruption of rhythmic oscillations. We use a hippocampal network model to explore these scenarios. The basis for these experiments is the proposal that the brain represents memories as groups of anatomically dispersed strongly connected neurons. A neuronal network with connections modified by synaptic plasticity was exposed to patterned stimuli, after which spiking output demonstrated evidence of stimulus-related neuronal group development analogous to memory formation. The effect of GABAA potentiation on this memory model was studied in 100 unique networks. GABAA potentiation consistent with moderate propofol effects reduced neuronal group size formed in response to a patterned stimulus by around 70%. Concurrently, accuracy of a Bayesian classifier in identifying learned patterns in the network output was reduced. Greater potentiation led to near total failure of group formation. Theta rhythm variations had no effect on group size or classifier accuracy. Memory formation is widely thought to depend on changes in neuronal connection strengths during learning that enable neuronal groups to respond with greater facility to familiar stimuli. This experiment suggests the ability to form such groups is sensitive to alteration in the balance between excitation and inhibition such as that resulting from administration of a γ-aminobutyric acid-mediated anesthetic agent.

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.

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.

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.

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

GABAB receptor-mediated, layer-specific synaptic plasticity reorganizes gamma-frequency neocortical response to stimulation

PubMed Central

Ainsworth, Matthew; Lee, Shane; Kaiser, Marcus; Simonotto, Jennifer; Kopell, Nancy J.

2016-01-01

Repeated presentations of sensory stimuli generate transient gamma-frequency (30–80 Hz) responses in neocortex that show plasticity in a task-dependent manner. Complex relationships between individual neuronal outputs and the mean, local field potential (population activity) accompany these changes, but little is known about the underlying mechanisms responsible. Here we show that transient stimulation of input layer 4 sufficient to generate gamma oscillations induced two different, lamina-specific plastic processes that correlated with lamina-specific changes in responses to further, repeated stimulation: Unit rates and recruitment showed overall enhancement in supragranular layers and suppression in infragranular layers associated with excitatory or inhibitory synaptic potentiation onto principal cells, respectively. Both synaptic processes were critically dependent on activation of GABAB receptors and, together, appeared to temporally segregate the cortical representation. These data suggest that adaptation to repetitive sensory input dramatically alters the spatiotemporal properties of the neocortical response in a manner that may both refine and minimize cortical output simultaneously. PMID:27118845

GABAB receptor-mediated, layer-specific synaptic plasticity reorganizes gamma-frequency neocortical response to stimulation.

PubMed

Ainsworth, Matthew; Lee, Shane; Kaiser, Marcus; Simonotto, Jennifer; Kopell, Nancy J; Whittington, Miles A

2016-05-10

Repeated presentations of sensory stimuli generate transient gamma-frequency (30-80 Hz) responses in neocortex that show plasticity in a task-dependent manner. Complex relationships between individual neuronal outputs and the mean, local field potential (population activity) accompany these changes, but little is known about the underlying mechanisms responsible. Here we show that transient stimulation of input layer 4 sufficient to generate gamma oscillations induced two different, lamina-specific plastic processes that correlated with lamina-specific changes in responses to further, repeated stimulation: Unit rates and recruitment showed overall enhancement in supragranular layers and suppression in infragranular layers associated with excitatory or inhibitory synaptic potentiation onto principal cells, respectively. Both synaptic processes were critically dependent on activation of GABAB receptors and, together, appeared to temporally segregate the cortical representation. These data suggest that adaptation to repetitive sensory input dramatically alters the spatiotemporal properties of the neocortical response in a manner that may both refine and minimize cortical output simultaneously.

Crucial Role of Postsynaptic Syntaxin 4 in Mediating Basal Neurotransmission and Synaptic Plasticity in Hippocampal CA1 Neurons.

PubMed

Bin, Na-Ryum; Ma, Ke; Harada, Hidekiyo; Tien, Chi-Wei; Bergin, Fiona; Sugita, Kyoko; Luyben, Thomas T; Narimatsu, Masahiro; Jia, Zhengping; Wrana, Jeffrey L; Monnier, Philippe P; Zhang, Liang; Okamoto, Kenichi; Sugita, Shuzo

2018-06-05

Trafficking of neurotransmitter receptors on postsynaptic membranes is critical for basal neurotransmission and synaptic plasticity, yet the underlying mechanisms remain elusive. Here, we investigated the role of syntaxin 4 in postsynaptic hippocampal CA1 neurons by analyzing conditional knockout (syntaxin 4 cKO) mice. We show that syntaxin 4 cKO resulted in reduction of basal neurotransmission without changes in paired-pulse ratios. Both α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-d-aspartic acid (NMDA) receptor-mediated charge transfers were diminished. Patch-clamp experiments revealed that amplitudes, but not frequencies, of spontaneous excitatory postsynaptic currents are reduced. Syntaxin 4 knockout (KO) caused drastic reduction in expression of surface α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-d-aspartic acid (NMDA) receptors in cultured hippocampal neurons. Furthermore, cKO caused defects in theta-burst stimulation induced long-term potentiation and spatial learning as assessed by a water maze task, indicating that synaptic plasticity was altered. Our data reveal a crucial role of syntaxin 4 in trafficking of ionotropic glutamate receptors that are essential for basal neurotransmission, synaptic plasticity, and spatial memory. Copyright © 2018 The Author(s). Published by Elsevier Inc. 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

Deficits in cognition and synaptic plasticity in a mouse model of Down syndrome ameliorated by GABAB receptor antagonists

PubMed Central

Kleschevnikov, A.M.; Belichenko, P.V.; Faizi, M.; Jacobs, L.F.; Htun, K.; Shamloo, M.; Mobley, W.C.

2012-01-01

Cognitive impairment in Down syndrome (DS) is characterized by deficient learning and memory. Mouse genetic models of DS exhibit impaired cognition in hippocampally mediated behavioral tasks and reduced synaptic plasticity of hippocampal pathways. Enhanced efficiency of GABAergic neurotransmission was implicated in those changes. We have recently shown that signaling through postsynaptic GABAB receptors is significantly increased in the dentate gyrus (DG) of Ts65Dn mice, a genetic model of DS. Here we examined a role for GABAB receptors in cognitive deficits in DS by defining the effect of selective GABAB receptor antagonists on behavior and synaptic plasticity of adult Ts65Dn mice. Treatment with the GABAB receptor antagonist CGP55845 restored memory of Ts65Dn mice in the novel place recognition, novel object recognition and contextual fear conditioning tasks, but did not affect locomotion and performance in T-maze. The treatment increased hippocampal levels of brain-derived neurotrophic factor (BDNF), equally in 2N and Ts65Dn mice. In hippocampal slices, treatment with the GABAB receptor antagonists CGP55845 or CGP52432 enhanced long-term potentiation (LTP) in the Ts65Dn DG. The enhancement of LTP was accompanied by an increase in the NMDA receptor-mediated component of the tetanus-evoked responses. These findings are evidence for a contribution of GABAB receptors to changes in hippocampal-based cognition in the Ts65Dn mouse. The ability to rescue cognitive performance through treatment with selective GABAB receptor antagonists motivates studies to further explore the therapeutic potential of these compounds in people with DS. PMID:22764230

Neurexin and Neuroligin Mediate Retrograde Synaptic Inhibition in C. elegans

PubMed Central

Hu, Zhitao; Hom, Sabrina; Kudze, Tambudzai; Tong, Xia-Jing; Choi, Seungwon; Aramuni, Gayane; Zhang, Weiqi; Kaplan, Joshua M.

2013-01-01

The synaptic adhesion molecules Neurexin and Neuroligin alter the development and function of synapses and are linked to autism in humans. We find that C. elegans Neurexin (NRX-1) and Neuroligin (NLG-1) mediate a retrograde synaptic signal that inhibits neurotransmitter release at neuromuscular junctions. Retrograde signaling was induced in mutants lacking a muscle microRNA (miR-1) and was blocked in mutants lacking NLG-1 or NRX-1. Release was rapid and abbreviated when the retrograde signal was on whereas release was slow and prolonged when retrograde signaling was blocked. The retrograde signal adjusted release kinetics by inhibiting exocytosis of synaptic vesicles (SVs) that are distal to the site of calcium entry. Inhibition of release was mediated by increased pre-synaptic levels of Tomosyn, an inhibitor of SV fusion. PMID:22859820

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

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

Treatment with a Clinically-Relevant Dose of Methylphenidate Alters NMDA Receptor Composition and Synaptic Plasticity in the Juvenile Rat Prefrontal Cortex

PubMed Central

Urban, Kimberly R.; Li, Yan-Chun; Gao, Wen-Jun

2013-01-01

Methylphenidate (Ritalin, MPH) is the most commonly prescribed psychoactive drug for children. Used to treat attention-deficit/hyperactivity disorder (ADHD) and for cognitive enhancement in healthy individuals, its cellular mechanisms of action and potential long-term effects are poorly understood. We recently reported that a clinically relevant (1 mg/kg i.p., single injection) dose of MPH significantly decreased neuronal excitability in the juvenile rat prefrontal cortical neurons. Here we further explore the actions of acute treatment with MPH on the level of NMDA receptor subunits and NMDA receptor-mediated short- and long-term synaptic plasticity in the juvenile rat prefrontal cortical neurons. We found that a single dose of MPH treatment (1 mg/kg, intraperitoneal) significantly decreased the surface and total protein levels of NMDA receptor subunits NR1 and NR2B, but not NR2A, in the juvenile prefrontal cortex. In addition, the amplitude, decay time and charge transfer of NMDA receptor-mediated EPSCs were significantly decreased whereas the amplitude and short-term depression of AMPA receptor-mediated EPSCs were significantly increased in the prefrontal neurons. Furthermore, MPH treatment also significantly increased the probability and magnitude of LTP induction, but had only a small effect on LTD induction in juvenile rat prefrontal cortical neurons. Our data thus present a novel mechanism of action of MPH, i.e., changes in glutamatergic receptor-mediated synaptic plasticity following early-life treatment. Furthermore, since a single dosage resulted in significant changes in NMDA receptors, off-label usage by healthy individuals, especially children and adolescents, may result in altered potential for plastic learning. PMID:23333502

Muscimol increases acetylcholine release by directly stimulating adult striatal cholinergic interneurons.

PubMed

Login, I S; Pal, S N; Adams, D T; Gold, P E

1998-01-01

Because GabaA ligands increase acetylcholine (ACh) release from adult striatal slices, we hypothesized that activation of GabaA receptors on striatal cholinergic interneurons directly stimulates ACh secretion. Fractional [3H]ACh release was recorded during perifusion of acutely dissociated, [3H]choline-labeled, adult male rat striata. The GabaA agonist, muscimol, immediately stimulated release maximally approximately 300% with EC50 = approximately 1 microM. This action was enhanced by the allosteric GabaA receptor modulators, diazepam and secobarbital, and inhibited by the GabaA antagonist, bicuculline, by ligands for D2 or muscarinic cholinergic receptors or by low calcium buffer, tetrodotoxin or vesamicol. Membrane depolarization inversely regulated muscimol-stimulated secretion. Release of endogenous and newly synthesized ACh was stimulated in parallel by muscimol without changing choline release. Muscimol pretreatment inhibited release evoked by K+ depolarization or by receptor-mediated stimulation with glutamate. Thus, GabaA receptors on adult striatal cholinergic interneurons directly stimulate voltage- and calcium-dependent exocytosis of ACh stored in vesamicol-sensitive synaptic vesicles. The action depends on the state of membrane polarization and apparently depolarizes the membrane in turn. This functional assay demonstrates that excitatory GabaA actions are not limited to neonatal tissues. GabaA-stimulated ACh release may be prevented in situ by normal tonic dopaminergic and muscarinic input to cholinergic neurons.

Sex differences in cerebellar synaptic transmission and sex-specific responses to autism-linked Gabrb3 mutations in mice

PubMed Central

Mercer, Audrey A; Palarz, Kristin J; Tabatadze, Nino; Woolley, Catherine S; Raman, Indira M

2016-01-01

Neurons of the cerebellar nuclei (CbN) transmit cerebellar signals to premotor areas. The cerebellum expresses several autism-linked genes, including GABRB3, which encodes GABAA receptor β3 subunits and is among the maternal alleles deleted in Angelman syndrome. We tested how this Gabrb3 m-/p+ mutation affects CbN physiology in mice, separating responses of males and females. Wild-type mice showed sex differences in synaptic excitation, inhibition, and intrinsic properties. Relative to females, CbN cells of males had smaller synaptically evoked mGluR1/5-dependent currents, slower Purkinje-mediated IPSCs, and lower spontaneous firing rates, but rotarod performances were indistinguishable. In mutant CbN cells, IPSC kinetics were unchanged, but mutant males, unlike females, showed enlarged mGluR1/5 responses and accelerated spontaneous firing. These changes appear compensatory, since mutant males but not females performed indistinguishably from wild-type siblings on the rotarod task. Thus, sex differences in cerebellar physiology produce similar behavioral output, but provide distinct baselines for responses to mutations. DOI: http://dx.doi.org/10.7554/eLife.07596.001 PMID:27077953

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.

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

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.

Non-synaptic receptors and transporters involved in brain functions and targets of drug treatment.

PubMed

Vizi, E S; Fekete, A; Karoly, R; Mike, A

2010-06-01

Beyond direct synaptic communication, neurons are able to talk to each other without making synapses. They are able to send chemical messages by means of diffusion to target cells via the extracellular space, provided that the target neurons are equipped with high-affinity receptors. While synaptic transmission is responsible for the 'what' of brain function, the 'how' of brain function (mood, attention, level of arousal, general excitability, etc.) is mainly controlled non-synaptically using the extracellular space as communication channel. It is principally the 'how' that can be modulated by medicine. In this paper, we discuss different forms of non-synaptic transmission, localized spillover of synaptic transmitters, local presynaptic modulation and tonic influence of ambient transmitter levels on the activity of vast neuronal populations. We consider different aspects of non-synaptic transmission, such as synaptic-extrasynaptic receptor trafficking, neuron-glia communication and retrograde signalling. We review structural and functional aspects of non-synaptic transmission, including (i) anatomical arrangement of non-synaptic release sites, receptors and transporters, (ii) intravesicular, intra- and extracellular concentrations of neurotransmitters, as well as the spatiotemporal pattern of transmitter diffusion. We propose that an effective general strategy for efficient pharmacological intervention could include the identification of specific non-synaptic targets and the subsequent development of selective pharmacological tools to influence them.

Antagonism of GABA-B but not GABA-A receptors in the VTA prevents stress- and intra-VTA CRF-induced reinstatement of extinguished cocaine seeking in rats.

PubMed

Blacktop, Jordan M; Vranjkovic, Oliver; Mayer, Matthieu; Van Hoof, Matthew; Baker, David A; Mantsch, John R

2016-03-01

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

Physical exercise improves synaptic dysfunction and recovers the loss of survival factors in 3xTg-AD mouse brain.

PubMed

Revilla, Susana; Suñol, Cristina; García-Mesa, Yoelvis; Giménez-Llort, Lydia; Sanfeliu, Coral; Cristòfol, Rosa

2014-06-01

Physical exercise has become a potentially beneficial therapy for reducing neurodegeneration symptoms in Alzheimer's disease. Previous studies have shown that cognitive deterioration, anxiety and the startle response observed in 7-month-old 3xTg-AD mice were ameliorated after 6 months of free access to a running wheel. Also, alterations in synaptic response to paired-pulse stimulation were improved. The present study further investigated some molecular mechanisms underlying the beneficial effects of 6 months of voluntary exercise on synaptic plasticity in 7-month-old 3xTg-AD mice. Changes in binding parameters of [(3)H]-flunitrazepam to GABAA receptor and of [(3)H]-MK-801 to NMDA receptor in cerebral cortex of 3xTgAD mice were restored by voluntary exercise. In addition, reduced expression levels of NMDA receptor NR2B subunit were reestablished. The synaptic proteins synaptophysin and PSD-95 and the neuroprotective proteins GDNF and SIRT1 were downregulated in 3xTgAD mice and were recovered by exercise treatment. Overall, in this paper we highlight the fact that different interrelated mechanisms are involved in the beneficial effects of exercise on synaptic plasticity alterations in the 3xTg-AD mouse model. Copyright © 2014 Elsevier Ltd. All rights reserved.

Characterization of auditory synaptic inputs to gerbil perirhinal cortex

PubMed Central

Kotak, Vibhakar C.; Mowery, Todd M.; Sanes, Dan H.

2015-01-01

The representation of acoustic cues involves regions downstream from the auditory cortex (ACx). One such area, the perirhinal cortex (PRh), processes sensory signals containing mnemonic information. Therefore, our goal was to assess whether PRh receives auditory inputs from the auditory thalamus (MG) and ACx in an auditory thalamocortical brain slice preparation and characterize these afferent-driven synaptic properties. When the MG or ACx was electrically stimulated, synaptic responses were recorded from the PRh neurons. Blockade of type A gamma-aminobutyric acid (GABA-A) receptors dramatically increased the amplitude of evoked excitatory potentials. Stimulation of the MG or ACx also evoked calcium transients in most PRh neurons. Separately, when fluoro ruby was injected in ACx in vivo, anterogradely labeled axons and terminals were observed in the PRh. Collectively, these data show that the PRh integrates auditory information from the MG and ACx and that auditory driven inhibition dominates the postsynaptic responses in a non-sensory cortical region downstream from the ACx. PMID:26321918

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

Δ9-THC-caused synaptic and memory impairments are mediated through COX-2 signaling

PubMed Central

Yang, Hongwei; Tang, Ya-ping; Sun, Hao; Song, Yunping; Chen, Chu

2013-01-01

SUMMARY Marijuana has been used for thousands of years as a treatment for medical conditions. However, untoward side effects limit its medical value. Here we show that synaptic and cognitive impairments following repeated exposure to Δ9-tetrahydrocannabinol (Δ9-THC) are associated with the induction of cyclooxygenase-2 (COX-2), an inducible enzyme that converts arachidonic acid to prostanoids, in the brain. COX-2 induction by Δ9-THC is mediated via CB1 receptor-coupled G-protein βγ subunits. Pharmacological or genetic inhibition of COX-2 blocks down-regulation and internalization of glutamate receptor subunits and alterations of the dendritic spine density of hippocampal neurons induced by repeated Δ9-THC exposures. Ablation of COX-2 also eliminates Δ9-THC-impaired hippocampal long-term synaptic plasticity, spatial, and fear memories. Importantly, the beneficial effects of decreasing β-amyloid plaques and neurodegeneration by Δ9-THC in Alzheimer’s disease animals are retained in the presence of COX-2 inhibition. These results suggest that the applicability of medical marijuana would be broadened by concurrent inhibition of COX-2. PMID:24267894

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.

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.

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.

Myosin IIb-dependent Regulation of Actin Dynamics Is Required for N-Methyl-D-aspartate Receptor Trafficking during Synaptic Plasticity.

PubMed

Bu, Yunfei; Wang, Ning; Wang, Shaoli; Sheng, Tao; Tian, Tian; Chen, Linlin; Pan, Weiwei; Zhu, Minsheng; Luo, Jianhong; Lu, Wei

2015-10-16

N-Methyl-d-aspartate receptor (NMDAR) synaptic incorporation changes the number of NMDARs at synapses and is thus critical to various NMDAR-dependent brain functions. To date, the molecules involved in NMDAR trafficking and the underlying mechanisms are poorly understood. Here, we report that myosin IIb is an essential molecule in NMDAR synaptic incorporation during PKC- or θ burst stimulation-induced synaptic plasticity. Moreover, we demonstrate that myosin light chain kinase (MLCK)-dependent actin reorganization contributes to NMDAR trafficking. The findings from additional mutual occlusion experiments demonstrate that PKC and MLCK share a common signaling pathway in NMDAR-mediated synaptic regulation. Because myosin IIb is the primary substrate of MLCK and can regulate actin dynamics during synaptic plasticity, we propose that the MLCK- and myosin IIb-dependent regulation of actin dynamics is required for NMDAR trafficking during synaptic plasticity. This study provides important insights into a mechanical framework for understanding NMDAR trafficking associated with synaptic plasticity. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

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.

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.

Superficial NK1 expressing spinal dorsal horn neurones modulate inhibitory neurotransmission mediated by spinal GABA(A) receptors.

PubMed

Rahman, Wahida; Sikandar, Shafaq; Sikander, Shafaq; Suzuki, Rie; Hunt, Stephen P; Dickenson, Anthony H

2007-06-04

Lamina 1 projection neurones which express the NK1 receptor (NK1R+) drive a descending serotonergic pathway from the brainstem that enhances spinal dorsal horn neuronal activity via the facilitatory spinal 5-HT3 receptor. Selective destruction of these cells via lumbar injection of substance P-saporin (SP-SAP) attenuates pain behaviours, including mechanical and thermal hypersensitivity, which are mirrored by deficits in the evoked responses of lamina V-VI wide dynamic range (WDR) neurones to noxious stimuli. To assess whether removing the origin of this facilitatory spino-bulbo-spinal loop results in alterations in GABAergic spinal inhibitory systems, the effects of spinal bicuculline, a selective GABA(A) receptor antagonist, on the evoked neuronal responses to electrical (Abeta-, Adelta-, C-fibre, post-discharge and Input) and mechanical (brush, prod and von Frey (vF) 8 and 26 g) stimuli were measured in SAP and SP-SAP groups. In the SAP control group, bicuculline produced a significant dose related facilitation of the electrically evoked Adelta-, C-fibre, post-discharge and input neuronal responses. The evoked mechanical (prod, vF8 g and 26 g) responses were also significantly increased. Brush evoked neuronal responses in these animals were enhanced but did not reach significance. This facilitatory effect of bicuculline, however, was lost in the SP-SAP treated group. The generation of intrinsic GABAergic transmission in the spinal cord appears dependent on NK1 bearing neurons, yet despite the loss of GABAergic inhibitory controls after SP-SAP treatment, the net effect is a decrease in spinal cord excitability. Thus activation of these cells predominantly drives facilitation.

LTP-triggered cholesterol redistribution activates Cdc42 and drives AMPA receptor synaptic delivery

PubMed Central

Brachet, Anna; Norwood, Stephanie; Brouwers, Jos F.; Palomer, Ernest; Helms, J. Bernd

2015-01-01

Neurotransmitter receptor trafficking during synaptic plasticity requires the concerted action of multiple signaling pathways and the protein transport machinery. However, little is known about the contribution of lipid metabolism during these processes. In this paper, we addressed the question of the role of cholesterol in synaptic changes during long-term potentiation (LTP). We found that N-methyl-d-aspartate–type glutamate receptor (NMDAR) activation during LTP induction leads to a rapid and sustained loss or redistribution of intracellular cholesterol in the neuron. A reduction in cholesterol, in turn, leads to the activation of Cdc42 and the mobilization of GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid–type glutamate receptors (AMPARs) from Rab11-recycling endosomes into the synaptic membrane, leading to synaptic potentiation. This process is accompanied by an increase of NMDAR function and an enhancement of LTP. These results imply that cholesterol acts as a sensor of NMDAR activation and as a trigger of downstream signaling to engage small GTPase (guanosine triphosphatase) activation and AMPAR synaptic delivery during LTP. PMID:25753037

Non-synaptic receptors and transporters involved in brain functions and targets of drug treatment

PubMed Central

Vizi, ES; Fekete, A; Karoly, R; Mike, A

2010-01-01

Beyond direct synaptic communication, neurons are able to talk to each other without making synapses. They are able to send chemical messages by means of diffusion to target cells via the extracellular space, provided that the target neurons are equipped with high-affinity receptors. While synaptic transmission is responsible for the ‘what’ of brain function, the ‘how’ of brain function (mood, attention, level of arousal, general excitability, etc.) is mainly controlled non-synaptically using the extracellular space as communication channel. It is principally the ‘how’ that can be modulated by medicine. In this paper, we discuss different forms of non-synaptic transmission, localized spillover of synaptic transmitters, local presynaptic modulation and tonic influence of ambient transmitter levels on the activity of vast neuronal populations. We consider different aspects of non-synaptic transmission, such as synaptic–extrasynaptic receptor trafficking, neuron–glia communication and retrograde signalling. We review structural and functional aspects of non-synaptic transmission, including (i) anatomical arrangement of non-synaptic release sites, receptors and transporters, (ii) intravesicular, intra- and extracellular concentrations of neurotransmitters, as well as the spatiotemporal pattern of transmitter diffusion. We propose that an effective general strategy for efficient pharmacological intervention could include the identification of specific non-synaptic targets and the subsequent development of selective pharmacological tools to influence them. PMID:20136842

γ-glutamyl transpeptidase 1 specifically suppresses green-light avoidance via GABAA receptors in Drosophila.

PubMed

Liu, Jiangqu; Gong, Zhefeng; Liu, Li

2014-08-01

Drosophila larvae innately show light avoidance behavior. Compared with robust blue-light avoidance, larvae exhibit relatively weaker green-light responses. In our previous screening for genes involved in larval light avoidance, compared with control w(1118) larvae, larvae with γ-glutamyl transpeptidase 1 (Ggt-1) knockdown or Ggt-1 mutation were found to exhibit higher percentage of green-light avoidance which was mediated by Rhodopsin6 (Rh6) photoreceptors. However, their responses to blue light did not change significantly. By adjusting the expression level of Ggt-1 in different tissues, we found that Ggt-1 in malpighian tubules was both necessary and sufficient for green-light avoidance. Our results showed that glutamate levels were lower in Ggt-1 null mutants compared with controls. Feeding Ggt-1 null mutants glutamate can normalize green-light avoidance, indicating that high glutamate concentrations suppressed larval green-light avoidance. However, rather than directly, glutamate affected green-light avoidance indirectly through GABA, the level of which was also lower in Ggt-1 mutants compared with controls. Mutants in glutamate decarboxylase 1, which encodes GABA synthase, and knockdown lines of the GABAA receptor, both exhibit elevated levels of green-light avoidance. Thus, our results elucidate the neurobiological mechanisms mediating green-light avoidance, which was inhibited in wild-type larvae. © 2014 International Society for Neurochemistry.

Properties of synaptic transmission from the reticular formation dorsal to the facial nucleus to trigeminal motoneurons during early postnatal development in rats.

PubMed

Gemba-Nishimura, A; Inoue, T; Nakamura, S; Nakayama, K; Mochizuki, A; Shintani, S; Yoshimura, S

2010-03-31

We previously reported that electrical stimulation of the reticular formation dorsal to the facial nucleus (RdVII) elicited excitatory masseter responses at short latencies and that RdVII neurons were antidromically activated by stimulation of the trigeminal motor nucleus (MoV), suggesting that excitatory premotor neurons targeting the MoV are likely located in the RdVII. We thus examined the properties of synaptic transmission from the RdVII to jaw-closing and jaw-opening motoneurons in horizontal brainstem preparations from developing rats using voltage-sensitive dye, patch-clamp recordings and laser photostimulation. Electrical stimulation of the RdVII evoked optical responses in the MoV. Combined bath application of the non-N-methyl-d-aspartate (non-NMDA) receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), and the NMDA receptor antagonist DL-2-amino-5-phosphonopentanoic acid (APV) reduced these optical responses, and addition of the glycine receptor antagonist strychnine and the GABA(A) receptor antagonist bicuculline further reduced the remaining responses. Electrical stimulation of the RdVII evoked postsynaptic currents (PSCs) in all 19 masseter motoneurons tested in postnatal day (P)1-4 rats, and application of CNQX and the NMDA receptor antagonist (+/-)-3(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) reduced the PSC amplitudes by more than 50%. In the presence of CNQX and CPP, the GABA(A) receptor antagonist SR95531 further reduced PSC amplitude, and addition of strychnine abolished the remaining PSCs. Photostimulation of the RdVII with caged glutamate also evoked PSCs in masseter motoneurons of P3-4 rats. In P8-11 rats, electrical stimulation of the RdVII also evoked PSCs in all 14 masseter motoneurons tested, and the effects of the antagonists on the PSCs were similar to those in P1-4 rats. On the other hand, RdVII stimulation evoked PSCs in only three of 16 digastric motoneurons tested. These results suggest that both neonatal and

GABA type a receptor trafficking and the architecture of synaptic inhibition.

PubMed

Lorenz-Guertin, Joshua M; Jacob, Tija C

2018-03-01

Ubiquitous expression of GABA type A receptors (GABA A R) in the central nervous system establishes their central role in coordinating most aspects of neural function and development. Dysregulation of GABAergic neurotransmission manifests in a number of human health disorders and conditions that in certain cases can be alleviated by drugs targeting these receptors. Precise changes in the quantity or activity of GABA A Rs localized at the cell surface and at GABAergic postsynaptic sites directly impact the strength of inhibition. The molecular mechanisms constituting receptor trafficking to and from these compartments therefore dictate the efficacy of GABA A R function. Here we review the current understanding of how GABA A Rs traffic through biogenesis, plasma membrane transport, and degradation. Emphasis is placed on discussing novel GABAergic synaptic proteins, receptor and scaffolding post-translational modifications, activity-dependent changes in GABA A R confinement, and neuropeptide and neurosteroid mediated changes. We further highlight modern techniques currently advancing the knowledge of GABA A R trafficking and clinically relevant neurodevelopmental diseases connected to GABAergic dysfunction. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 78: 238-270, 2018. © 2017 Wiley Periodicals, Inc.

Peripheral Interventions Enhancing Brain Glutamate Homeostasis Relieve Amyloid β- and TNFα- Mediated Synaptic Plasticity Disruption in the Rat Hippocampus.

PubMed

Zhang, Dainan; Mably, Alexandra J; Walsh, Dominic M; Rowan, Michael J

2017-07-01

Dysregulation of glutamate homeostasis in the interstitial fluid of the brain is strongly implicated in causing synaptic dysfunction in many neurological and psychiatric illnesses. In the case of Alzheimer's disease (AD), amyloid β (Aβ)-mediated disruption of synaptic plasticity and memory can be alleviated by interventions that directly remove glutamate or block certain glutamate receptors. An alternative strategy is to facilitate the removal of excess glutamate from the nervous system by activating peripheral glutamate clearance systems. One such blood-based system, glutamate oxaloacetate transaminase (GOT), is activated by oxaloacetate, which acts as a co-substrate. We report here that synthetic and AD brain-derived Aβ-mediated inhibition of synaptic long-term potentiation in the hippocampus is alleviated by oxaloacetate. Moreover the effect of oxaloacetate was GOT-dependent. The disruptive effects of a general inhibitor of excitatory amino acid transport or TNFα, a pro-inflammatory mediator of Aβ action, were also reversed by oxaloacetate. Furthermore, another intervention that increases peripheral glutamate clearance, peritoneal dialysis, mimicked the beneficial effect of oxaloacetate. These findings lend support to the promotion of the peripheral clearance of glutamate as a means to alleviate synaptic dysfunction that is caused by impaired glutamate homeostasis in the brain. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons

PubMed Central

Oswald, Manfred J.; Schulz, Jan M.; Kelsch, Wolfgang; Oorschot, Dorothy E.; Reynolds, John N. J.

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg2+-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg2+-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission. PMID:25914618

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons.

PubMed

Oswald, Manfred J; Schulz, Jan M; Kelsch, Wolfgang; Oorschot, Dorothy E; Reynolds, John N J

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg(2+)-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg(2+)-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission.