Muscimol Binding Research Articles

Anxiolytic effects of Julibroside C1 isolated from Albizzia julibrissin in mice

Julibroside C1 is a saponin-containing compound isolated from Albizzia julibrissin Durazz. In this study, we investigated the putative anxiolytic effects of Julibroside C1 using the elevated plus maze (EPM) in mice. Julibroside C1 at doses of 0.5 and 1 mg/kg significantly increased the time spent in the open arms and the number of entries into the open arms of the EPM compared to the control group. Moreover, the anxiolytic-like effects of Julibroside C1 (0.5 mg/kg) were blocked by WAY-100635 (5-HT1A receptor antagonist), bicuculline (GABA(A) receptor antagonist), and flumazenil (antagonist of the GABA(A) receptor benzodiazepine site). However, Julibroside C1 did not change locomotor activity or induce myorelaxant effects. We used quantitative receptor autoradiography to investigate the effects of Julibroside C1 on alterations in mouse brain receptors. After acute treatment with Julibroside C1 (0.5 mg/kg), [(3)H]-8-OH-DPAT binding was significantly decreased in the CA1 region of the hippocampus and [(3)H]-flunitrazepam binding was decreased remarkably in the cingulate cortex region. However, [(3)H]-muscimol binding did not show a significant change in any brain region. Taken together, our findings suggest that Julibroside C1 shows anxiolytic-like effects, which might be mediated by the 5-HT1A and GABA(A)-benzodiazepine receptor systems.

DHA prevents altered 5-HT1A, 5-HT2A, CB1 and GABAA receptor binding densities in the brain of male rats fed a high-saturated-fat diet

Low levels of docosahexaenoic acid (DHA) have been linked to a number of mental illnesses such as memory loss, depression and schizophrenia. While supplementation of DHA is beneficial in improving memory and cognition, the influence of dietary fats on the neurotransmitters and receptors involved in cognitive function is still not known. The aim of this study was to investigate serotonin receptor (5-HT1A and 5-HT2A), cannabinoid receptor (CB1) and gamma-aminobutyric acid type A (GABAA) receptor binding densities in the brain of male rats fed a high-saturated-fat (HF) diet, as well as the effect of DHA supplementation on HF diet. Alterations of these receptors in the post-mortem rat brain were detected by [3H]-WAY-100635, [3H]-ketanserin, [3H]-CP-55,940 and [3H]-muscimol binding autoradiography, respectively. In the hippocampus, the 5-HT1A, CB1 and GABAA receptor binding densities significantly increased in response to an HF diet, while in the hypothalamus, 5-HT1A and CB1 binding densities significantly increased in HF-fed rats. Importantly, DHA supplementation prevented the HF-induced increase of receptors binding density in the hippocampus and hypothalamus. Furthermore, DHA supplementation attenuated 5-HT2A receptor binding density in the caudate putamen, anterior cingulate cortex and medial mammillary nucleus, which was also increased in HF group. This study showed that an HF diet increased 5-HT1A, 5-HT2A, CB1 and GABAA receptor binding densities in the brain regions involved in cognitive function and that dietary DHA can attenuate such alterations. These findings provide insight into the mechanism by which DHA supplementation ameliorates reduced cognitive function associated with an HF diet.

Increases in [3H]Muscimol and [3H]Flumazenil Binding in the Dorsolateral Prefrontal Cortex in Schizophrenia Are Linked to α4 and γ2S mRNA Levels Respectively

BackgroundGABAA receptors (GABAAR) are composed of several subunits that determine sensitivity to drugs, synaptic localisation and function. Recent studies suggest that agonists targeting selective GABAAR subunits may have therapeutic value against the cognitive impairments observed in schizophrenia. In this study, we determined whether GABAAR binding deficits exist in the dorsolateral prefrontal cortex (DLPFC) of people with schizophrenia and tested if changes in GABAAR binding are related to the changes in subunit mRNAs. The GABA orthosteric and the benzodiazepine allosteric binding sites were assessed autoradiographically using [3H]Muscimol and [3H]Flumazenil, respectively, in a large cohort of individuals with schizophrenia (n = 37) and their matched controls (n = 37). We measured, using qPCR, mRNA of β (β1, β2, β3), γ (γ1, γ2, γ2S for short and γ2L for long isoform, γ3) and δ subunits and used our previous measurements of GABAAR α subunit mRNAs in order to relate mRNAs and binding through correlation and regression analysis.ResultsSignificant increases in both [3H]Muscimol (p = 0.016) and [3H]Flumazenil (p = 0.012) binding were found in the DLPFC of schizophrenia patients. Expression levels of mRNA subunits measured did not show any significant difference in schizophrenia compared to controls. Regression analysis revealed that in schizophrenia, the [3H]Muscimol binding variance was most related to α4 mRNA levels and the [3H]Flumazenil binding variance was most related to γ2S subunit mRNA levels. [3H]Muscimol and [3H]Flumazenil binding were not affected by the lifetime anti-psychotics dose (chlorpromazine equivalent).ConclusionsWe report parallel increases in orthosteric and allosteric GABAAR binding sites in the DLPFC in schizophrenia that may be related to a “shift” in subunit composition towards α4 and γ2S respectively, which may compromise normal GABAergic modulation and function. Our results may have implications for the development of treatment strategies that target specific GABAAR receptor subunits.

In vitro neuropharmacological evaluation of penitrem-induced tremorgenic syndromes: Importance of the GABAergic system

The effects of the fungal neurotoxin penitrem A on the GABAergic and glutamatergic systems in rat brain were evaluated. Penitrem A inhibited binding of the GABA A-receptor ligand [ 3H]TBOB to rat forebrain and cerebellar membrane preparations with IC 50 (half maximal inhibitory concentration) values of 11 and 9 μM, respectively. Furthermore, penitrem A caused a concentration-dependent increase of [ 3H]flunitrazepam and [ 3H]muscimol binding in rat forebrain, but not in cerebellar preparations. The stimulation of [ 3H]flunitrazepam binding by penitrem A was abolished by the addition of GABA. In cerebellar preparations, a different pharmacological profile was found, with penitrem A allosterically inhibiting [ 3H]TBOB binding by interacting with a bicuculline-sensitive site. Moreover, penitrem A inhibited the high affinity uptake of GABA and glutamate into cerebellar synaptosomes with IC 50 values of 20 and 47 μM, respectively. The toxin showed no effect on NMDA or AMPA glutamate receptor binding. In conclusion, our results suggest that penitrem A exerts region-specific effects in the brain, leading to positive modulation of GABA A-receptor function in forebrain. Conversely, penitrem A may act as a bicuculline-like convulsant in cerebellum.

Autoradiographic analysis of GABAA receptor binding in the neural anxiety network of postpartum and non-postpartum laboratory rats

Postpartum female rats exhibit a suppression of anxiety-related behaviors when compared to diestrous virgin females, pregnant females, and males. This blunted anxiety promotes optimal maternal care and involves elevated GABA neurotransmission, possibly including greater density of GABAA and benzodiazepine receptors in the postpartum brain. We here examined autoradiographic binding of [3H]muscimol to measure the total population of GABAA receptors and [3H]flunitrazepam to assess density of benzodiazepine sites in the medial prefrontal cortex, bed nucleus of the stria terminalis, amygdala, hippocampus, and periaqueductal gray of female rats sacrificed on day 7 postpartum, day 10 of pregnancy, or as diestrous virgins. A group of sexually naïve male rats was also included. We found that [3H]muscimol binding did not differ among groups in any site but that diestrous virgin females had greater [3H]flunitrazepam binding in the CA1 and dentate gyrus of the hippocampus compared to mid-pregnant females and males. Notably, postpartum and diestrous virgin females did not significantly differ in binding of either ligand in any site examined. This is the first study to evaluate the densities of GABAA and benzodiazepine binding sites simultaneously across three female reproductive states and sex with a focus on brain sites influencing anxiety-related behaviors. The results suggest that changes in other GABAA receptor characteristics such as subunit composition, or increased presynaptic GABA release during interactions with offspring, must instead play a greater role in the postpartum suppression of anxiety in laboratory rats.

Modulation of the GABAA Receptor by Barbiturates and Pregnane Steroids: Differential Effects of the Influence of Assay Temperature

The effect of temperature on the modulation of the GABAA receptor by barbiturates and steroids has been investigated in-vitro using a radioreceptor binding assay. Displaceable [3H]muscimol binding to a crude membrane preparation from rat cerebral cortex was enhanced by the endogenous steroid metabolite, 5 beta-pregnan-3 alpha-ol-20-one, by the synthetic steroid, alphaxalone, and by pentobarbitone in a dose-dependent manner. Hydrocortisone and corticosterone had no significant effect on [3H]muscimol binding. Analysis of binding data using a curve-fitting program ('Ligand') showed that both pentobarbitone (1 mM) and 5 beta-pregnan-3 alpha-ol-20-one (10 microM) increased the apparent number of high affinity binding sites in the membrane but had no effect on the affinity of [3H]muscimol binding (Kd approx. 11 nM). Increasing the assay temperature from 0 degrees C to 35 degrees C decreased [3H]muscimol binding and decreased the enhancement of binding by pentobarbitone but had no effect on 5 beta-pregnan-3 alpha-ol-20-one enhancement of binding. 5 alpha-Pregnan-3 alpha-ol-20-one increased the apparent rate of association of [3H]muscimol binding to its receptor whereas pentobarbitone had no effect. These different effects on the apparent association rate and the different responses to temperature, suggest that the barbiturate and steroid may interact with the GABAA receptor through different binding sites.

Effect of single and repeated convulsions on glutamate decarboxylase (GAD) activity and [3H]muscimol binding in the rat brain

Effect of single and repeated convulsions on glutamate decarboxylase (GAD) activity and [3H]muscimol binding in the rat brain

The cerebellar GABA AR α6-R100Q polymorphism alters ligand binding in outbred Sprague–Dawley rats in a similar manner as in selectively bred AT and ANT rats

The alcohol-tolerant AT and alcohol-nontolerant ANT rat lines have been selectively bred for innate sensitivity to ethanol-induced motor impairment. The cerebellar GABA A receptor (GABA AR) α6 subunit alleles α6-100R and α6-100Q are segregated in the AT and ANT rats, respectively. This α6 polymorphism might explain various differences in pharmacological properties and density of GABA ARs between the rat lines. In the present study, we have used nonselected outbred Sprague–Dawley rats homozygous for the α6-100RR (RR) and α6-100QQ (QQ) genotypes to show that these RR and QQ rats display similar differences between genotypes as AT and ANT rat lines. The genotypes differed in their affinity for [ 3H]Ro 15-4513 and classic benzodiazepines (BZs) to cerebellar “diazepam-insensitive” (DZ-IS) binding sites, in density of cerebellar [ 3H]muscimol binding and in the antagonizing effect of furosemide on GABA-induced inhibition of [ 3H]EBOB binding. The results suggest the involvement of α6-R100Q polymorphism in these line differences and in the differences previously found between AT and ANT rats. In addition, the α6-R100Q polymorphism induces striking differences in [ 3H]Ro 15-4513 binding kinetics to recombinant α6β3γ2s receptors and cerebellar DZ-IS sites. Association of [ 3H]Ro 15-4513 binding was ∼10-fold faster and dissociation was ∼3–4-fold faster in DZ-IS α6βγ2 receptors containing the α6-100Q allele, with a resulting change of ∼2.5-fold in equilibrium dissociation constant ( K D). The results indicate that in addition to the central role of the homologous α6-100R/Q (α1-101H) residue in BZ binding and efficacy, this critical BZ binding site residue has a major impact on BZ binding kinetics.

Decreased glutamic acid decarboxylase mRNA expression in prefrontal cortex in Parkinson's disease

Parkinson's disease (PD) patients typically suffer from motor disorders but mild to severe cognitive deficits can also be present. Neuropathology of PD primarily involves loss of dopaminergic neurons in the substantia nigra, pars compacta, although more widespread pathology from the brainstem to the cerebral cortex occurs at different stages of the disease. Cognitive deficits in PD are thought to involve the cerebral cortex, and imaging studies have identified the dorsolateral prefrontal cortex (DLPFC) as a possible site for some of the symptoms. GABAergic neurons in the cerebral cortex play a key role in the modulation of pyramidal neurons and alterations in muscimol binding to GABA A receptors have been reported in Brodmann area 9 (BA9) of the prefrontal cortex in PD patients (Nishino et al., 1988). In order to further assess the likelihood that GABAergic activity is altered in the prefrontal cortex in PD, gene expression of the 67 kilodalton isoform of the GABA-synthesizing enzyme, glutamic acid decarboxylase (GAD67 encoded by the GAD1 gene), was examined in BA9 of post-mortem brains from 19 patients and 20 controls using isotopic in situ hybridization histochemistry. GAD67 mRNA labeling was examined and quantified on X-ray films and emulsion radioautographs. We show that GAD67 mRNA labeling is significantly lower in PD compared to control cases. Analysis of emulsion radioautographs indicates that GAD67 mRNA labeling is decreased in individual neurons and is not paralleled by a decrease in the number of GAD67 mRNA-labeled neurons. Analysis of expression data from a microarray study performed in 29 control and 33 PD samples from BA9 confirms that GAD67 expression is decreased in PD. Another finding from the microarray study is a negative relationship between GAD67 mRNA expression and age at death. Altogether, the results support the possibility that GABAergic neurotransmission is impaired in the DLPFC in PD, an effect that may be involved in some of the behavioral deficits associated with the disease.

Melatonin Increases Bladder Capacity via GABAergic System and Decreases Urine Volume in Rats

Impaired melatonin production is involved in disruption of the normal circadian pattern of sleep, which leads to nocturia in older adults. Melatonin improves nocturia. We hypothesized that melatonin could alleviate urinary frequency by suppressing the brain micturition center. We investigated the central contribution of melatonin to bladder function and urine volume. Cystometry was done in conscious female Sprague-Dawley rats (Japan SLC, Hamamatsu, Japan). We examined the effect of melatonin alone (4.3 x 10(-1) to 4.3 x 10 pmol intracerebroventricularly) or with the gamma-aminobutyric acid(A) antagonist bicuculline (5.0 x 10(-5) mg/kg intravenously) on bladder function. The influence of melatonin (4.3 x 10 pmol intracerebroventricularly) on urine volume was investigated in water loaded rats. Blood samples were collected to determine antidiuretic hormone, atrial natriuretic peptide and brain natriuretic peptide 4 hours after melatonin administration. Melatonin significantly increased bladder capacity dose dependently by 27.0%, 40.8% and 63.5% at 4.3 x 10(-1), 4.3 and 4.3 x 10 pmol, respectively, but had no significant effect on bladder contraction pressure. Bicuculline inhibited the melatonin induced increases in bladder capacity. Melatonin decreased urine volume in water loaded rats but plasma antidiuretic hormone, atrial natriuretic peptide and bladder contraction pressure were not changed. Results suggest that melatonin increases bladder capacity via gamma-aminobutyric acid(A) receptor in the brain and decreases urine volume. Thus, melatonin could be beneficial for nocturia via a central nervous system effect.

Effects of benzodiazepine treatment on cortical GABA A and muscarinic receptors: Studies in schizophrenia and rats

Changes in cortical γ-aminobutyric acid A (GABA A) receptors and muscarinic receptors have been reported in schizophrenia, a disorder treated with antipsychotic drugs and benzodiazepines. As there is a reported functional relationship between the GABAergic and cholinergic systems in the human central nervous system we have investigated whether there are changes in the GABA A and muscarinic receptors in the cortex of subjects from APD-treated subjects with schizophrenia and whether changes were different in subjects who had also received benzodiazepine treatment. We failed to show any strong correlations between changes in GABA A and muscarinic receptors in the CNS of subjects with schizophrenia. We showed that subjects with schizophrenia treated with benzodiazepines had lower levels of muscarinic receptors; which was not the case in rats treated with APDs, benzodiazepines or a combination of both drugs. Further, the benzodiazepine binding site, but not the muscimol binding site, was decreased in the parietal cortex of subjects with schizophrenia independent of benzodiazepine status at death. These data would therefore support our previously stated hypotheses that changes in the cortical cholinergic and GABAergic systems are involved in the pathophysiology of schizophrenia.

Stoichiometry of Expressed α4β2δ γ-Aminobutyric Acid Type A Receptors Depends on the Ratio of Subunit cDNA Transfected

The gamma-aminobutyric acid type A receptor (GABA(A)R) is the target of many depressants, including benzodiazepines, anesthetics, and alcohol. Although the highly prevalent alphabetagamma GABA(A)R subtype mediates the majority of fast synaptic inhibition in the brain, receptors containing delta subunits also play a key role, mediating tonic inhibition and the actions of endogenous neurosteroids and alcohol. However, the fundamental properties of delta-containing GABA(A)Rs, such as subunit stoichiometry, are not well established. To determine subunit stoichiometry of expressed delta-containing GABA(A)Rs, we inserted the alpha-bungarotoxin binding site tag in the alpha(4), beta(2), and delta subunit N termini. An enhanced green fluorescent protein tag was also inserted into the beta(2) subunit to shift its molecular weight, allowing us to separate subunits using SDS-PAGE. Tagged alpha(4)beta(2)delta GABA(A)Rs were expressed in HEK293T cells using various ratios of subunit cDNA, and receptor subunit stoichiometry was determined by quantitating fluorescent alpha-bungarotoxin bound to each subunit on Western blots of surface immunopurified tagged GABA(A)Rs. The results demonstrate that the subunit stoichiometry of alpha(4)beta(2)delta GABA(A)Rs is regulated by the ratio of subunit cDNAs transfected. Increasing the ratio of delta subunit cDNA transfected increased delta subunit incorporation into surface receptors with a concomitant decrease in beta(2) subunit incorporation. Because receptor subunit stoichiometry can directly influence GABA(A)R pharmacological and functional properties, considering how the transfection protocols used affect subunit stoichiometry is essential when studying heterologously expressed alpha(4)beta(2)delta GABA(A)Rs. Successful bungarotoxin binding site tagging of GABA(A)R subunits is a novel tool with which to accurately quantitate subunit stoichiometry and will be useful for monitoring GABA(A)R trafficking in live cells.

Decreased GABAA and GABAB receptor functional activity in cannabinoid CB1 receptor knockout mice

The interaction between brain GABAergic and endocannabinoid systems was evaluated by examining the quantitative and functional status of GABAergic receptors in cannabinoid CB(1) receptor knockout (CB(1)(-/-)) mice. To this aim, GABA(A) ([(3)H]-Muscimol binding assay), GABA(B) (baclofen-stimulated [(35)S]-GTPγS binding assay), GABA(A)α(1), GABA(A)α(2) and GABA(A)γ(2) receptors gene expression (real-time reverse transcriptase polymerase chain reaction [PCR]) were carried out in CB(1)(-/-) and wild-type mice (CB(1)(+/+)). [(3)H]-Muscimol binding assays revealed significant reduction in the density of GABA(A) receptors in CA2 (30%) and DG (28%) of the hippocampus, thalamus (40%), cingulate (28%) and motor cortex (35%) of CB(1)(-/-) mice. Functional activity of metabotropic GABA(B) receptors was measured by evaluating the ability of GABA(B) agonist baclofen to stimulate [(35)S]-GTPγS binding. The results showed significant reduced [(35)S]-GTPγS binding in CA1 (61%), CA3 (51%) and DG (60%) of CB(1)(-/-) mice compared with CB(1)(+/+) mice. Real-time reverse transcriptase PCR was carried out for evaluating gene expression of α(1), α(2) and γ(2) subunits of GABA(A) receptor in the amygdala. The results showed significant reduced GABA(A)α(1) (50%) and GABA(A)α(2) (40%) receptor subunits gene expression in the amygdala of CB(1)(-/-) mice. No difference was observed in GABA(A)γ(2) receptor subunit gene expression. This study provides strong evidence of the involvement of CB(1) receptors in the control of GABAergic responses mediated by GABA(A) and GABA(B) receptors, and suggests a possible role of the endocannabinoid system in the regulation of anxiety-related disorders.

Subchronic phencyclidine in rats: alterations in locomotor activity, maze performance, and GABAA receptor binding

Phencyclidine (PCP), an antagonist at the N-methyl-D-aspartate subtype of ionotropic glutamatergic receptors, decreases gamma-aminobutyric acid (GABA)ergic inhibition, suggesting that changes in GABAergic receptor function underlie behavioral and cognitive deficits resulting from repeated administration of PCP. To test this hypothesis, male Sprague-Dawley rats treated with PCP (4.5 mg/kg, intraperitoneal, twice a day for 7 consecutive days) or saline were tested in behavioral and cognitive tasks 7 days after injections. The PCP group showed increased amphetamine (1.5 mg/kg)-stimulated locomotor activity, and exhibited a greater number of errors in the double Y-maze memory task, when compared with controls. Subchronic PCP treatment increased [H]muscimol-binding sites and decreased affinity for [H]muscimol binding in frontal cortex, hippocampus, and striatum in comparison with controls. There were no changes in the expression of glutamic acid decarboxylase or the GABA membrane transporter protein. These data show that subchronic PCP administration induces an impaired performance of a previously learned task and an enhanced response to amphetamine in the rat. The observed changes in GABAA receptors in the rat brain are consistent with the notion that alterations in GABAergic receptor function contribute to the behavioral and cognitive impairments associated with repeated exposure to PCP.

In vitro and whole animal evidence that methylmercury disrupts GABAergic systems in discrete brain regions in captive mink

The effects of mercury (Hg) on key components of the GABAergic system were evaluated in discrete brain regions of captive juvenile male American mink ( Neovison vison) using in vitro and in vivo (whole animal) experimental approaches. In vitro studies on cortical brain tissues revealed that inorganic Hg (HgCl 2; IC50 = 0.5 ± 0.2 µM) and methyl Hg (MeHgCl; IC50 = 1.6 ± 0.2 µM) inhibited glutamic acid decarboxylase (GAD; EC 4.1.1.15) activity. There were no Hg-related effects on [ 3H]-muscimol binding to GABA(A) receptors (IC50s > 100 µM). HgCl 2 (IC50 = 0.8 ± 0.3 µM) but not MeHgCl (IC50 > 100 µM) inhibited GABA-transaminase (GABA-T; EC 2.6.1.19) activity. In a whole animal study, neurochemical indicators of GABAergic function were measured in brain regions (occipital cortex, cerebellum, brain stem, and basal ganglia) of captive mink fed relevant levels of MeHgCl (0 to 2 µg/g feed, ppm) daily for 89 d. No effects on GAD activity were measured. Concentration-dependent decreases in [ 3H]-muscimol binding to GABA(A) receptors and GABA-T activity were found in several brain regions, with reductions as great as 94% (for GABA(A) receptor levels) and 71% (for GABA-T activity) measured in the brain stem and basal ganglia. These results show that chronic exposure to environmentally relevant levels of MeHg disrupts GABAergic signaling. Given that GABA is the main inhibitory neurotransmitter in the mammalian nervous system, prolonged disruptions of its function may underlie the sub-clinical impacts of MeHg at relevant levels to animal health.

Prototypic GABAA Receptor Agonist Muscimol Acts Preferentially Through Forebrain High-Affinity Binding Sites

Muscimol has been regarded as a universal agonist for all gamma-aminobutyric acid type A receptor (GABA(A)-R) subtypes. However, brain regional distribution of muscimol's high-affinity binding sites greatly differs from those of other binding sites of the GABA(A)-R. To test whether behavioral effects of muscimol correlated with the density of high-affinity [(3)H]muscimol binding, we examined several GABA(A)-R subunit gene-modified mouse lines: alpha1, alpha4, or delta-knockouts (KO), alpha4+delta-double KO, and Thy1.2 promoter-driven alpha6 transgenic mice (Thy1alpha6). We determined the high-affinity [(3)H]muscimol binding in brain sections by quantitative autoradiography and sedative/ataxic effects induced in vivo by muscimol using a constant speed rotarod. alpha4-KO mice had reduced [(3)H]muscimol binding in the caudate-putamen, thalamus, and hippocampus, and were less sensitive to the behavioral impairment by muscimol. Similarly, delta-KO mice also had reduced binding to forebrain regions and a lower behavioral sensitivity to muscimol than their wild-type controls. In contrast, alpha1-KO mice had unaltered behavioral sensitivity to muscimol and unaltered [(3)H]muscimol binding, even though previous studies have demonstrated dramatically reduced binding to various other GABA(A)-R sites in these mice. Finally, Thy1alpha6 mice exhibited increased behavioral sensitivity to muscimol, and to another direct GABA-site agonist gaboxadol, and increased [(3)H]muscimol binding in the cerebral cortex and hippocampus. Thus, the differences in sedative and motor-impairing actions of muscimol in various mouse models correlated with the level of forebrain high-affinity [(3)H]muscimol binding. These data suggest that a small special population of GABA(A)-Rs, most likely extrasynaptic non-alpha1-containing receptors, strongly contributes to the in vivo pharmacological effects of muscimol.

Time course of the effects of steroid hormone deprivation elicited by ovariectomy or ovariectomy plus adrenalectomy on the affinity and density of GABA binding sites in distinct rat brain areas.

The time course of the behaviour of GABAA receptors (affinity of muscimol for GABA binding sites and their number) in membranes obtained from five distinct rat brain areas was evaluated in dependence of the decline of the serum concentrations of estradiol and progesterone following hormone deprivation by either ovariectomy (OVX) or ovary- plus adrenalectomy (OVX-ADX) after 1, 2, 3, 5, 14 and 30 days. Diestrus rats served as control. The effect of OVX on the GABA binding sites was due to a decrease of affinity rather than alterations in their density. Conversely, the combined surgery of OVX plus ADX affected both the affinity and the density. Also, the OVX-linked changes of muscimol binding were observed only after five days as compared to control animals, while the initial OVX-ADX associated changes occurred already after one day. OVX-ADX produced significant decreases in the number of binding sites at the shortest times investigated, in frontal cortex (FC) at day one and two and in the medulla oblongata (MED) after three days, which disappeared in the subsequent times studied. Concomitant with the major decrease of steroid hormone serum levels one day after OVX-ADX the FC displayed an increase in affinity by 45%. From the time when steroid serum levels are low the alterations in affinity elicited by both OVX-ADX and OVX only take a similar course, reflected by the following decrease in affinity. In MED an additional adrenalectomy did not significantly modify the OVX linked decrease of affinity, except that the initial effect was already evident two days after OVX-ADX.(ABSTRACT TRUNCATED AT 250 WORDS)

Transient oligemia is associated with long-term changes in binding densities of cortical inhibitory GABA A receptors in the rat brain

Recently, we could demonstrate in rats that a short transient oligemic period of only 20-minute duration, induced by systemic hypotension, resulted in a transient decline of spatial memory capacities without any histological damage over a subsequent period of 6 months. In our present study, we checked for more subtle alterations within the highly vulnerable hippocampal CA1 subfield using quantification of neuronal cell density and semi-quantitative analysis of the ischemia-sensitive protein MAP2. Since hippocampal excitatory and inhibitory neurotransmitter receptors are crucially involved in spatial memory processes, quantitative in vitro receptor autoradiography was performed using [ 3H]MK-801, [ 3H]AMPA, and [ 3H]muscimol for labeling of NMDA, AMPA, and GABA A receptors, respectively. Ligand binding values were analyzed in the ischemia-sensitive hippocampal formation and in the parietal cortex. Transient oligemia did not cause any reduction of the neuronal cell density in the vulnerable CA1 subfield nor did it cause any significant changes of MAP2 protein expression in the dendritic layers of CA1. After oligemia, hippocampal binding densities of NMDA, AMPA, and GABA A receptors remained unchanged in all subfields investigated. Similarly, there was no difference in cortical ligand binding values of the excitatory NMDA and AMPA receptors comparing oligemic to sham-operated control rats. In the parietal cortex, however, transient oligemia caused a significant reduction of [ 3H]muscimol binding values compared to controls. These findings demonstrate long-term changes in receptor binding densities even after short oligemic periods possibly reflecting part of the molecular basis leading to impaired cognitive abilities.

Excitatory and inhibitory neurotransmission is chronically altered following perinatal NMDA receptor blockade

N-methyl- d-aspartate (NMDA) receptor blockade in rodents induces behavioural and neurochemical changes reminiscent of schizophrenia symptoms and pathology. To examine how NMDA receptor blockade affects glutamatergic and GABAergic pathways when administered during early brain development, [ 3H]MK-801 and [ 3H]muscimol binding to NMDA and GABA A receptors was examined at four time-points following injections of phencyclidine (PCP) or saline on postnatal days (PN)7, 9 and 11. [ 3H]MK-801 binding was significantly increased in PCP-treated rats in the thalamus from PN18 to PN96, in the prefrontal and anterior cingulate cortices at PN32, and in the hippocampus at PN96. In a similar manner, [ 3H]muscimol binding was increased in PCP-treated rats in the thalamus and hippocampus from PN18 to PN96, and in the prefrontal and anterior cingulate cortices at PN32. Glutamatergic and GABAergic transmission is therefore chronically altered by this treatment, which has relevance to disease processes that may be involved in schizophrenia.

Honokiol Modulates GABAA Receptors Subunit Specifically

Honokiol, a neolignan compound isolated from Magnolia species has been suggested to interact with GABAA receptors. Evidence comes from honokiol-induced enhanced [3H]muscimol binding [1] and anxiolytic action in behavioural studies [2]. The molecular mechanism and possible subunit-specific effects of honokiol on GABAA receptors are currently unknown.[...]