共查询到20条相似文献,搜索用时 31 毫秒
1.
Mohammad H. Jalilian Tehrani Brian J. Baumgartner Eugene M. Barnes Jr. 《Brain research》1997,776(1-2)
Clathrin-coated vesicles are thought to be a vehicle for the sequestration of GABAA receptors. For coated vesicles from bovine cerebrum, we examined the binding properties of [3H]muscimol, a GABAA-specific agonist, [3H]flunitrazepam, a benzodiazepine agonist, and [35S]t-butylbiocyclophosphorthionate (TBPS), a ligand for GABAA receptor channels. Under standard conditions, the binding level of [3H]muscimol, [3H]flunitrazepam, and [35S]TBPS to coated vesicles represented 12.3±1.8%, 7.9±1%, and 10.2±1.8%, respectively, of that in crude synaptic membranes. Coated vesicles showed a single [3H]flunitrazepam binding site with a KD value (12 nM) which was 9-fold that for synaptic membranes. The allosteric coupling between binding sites was measured by the addition of GABA to [3H]flunitrazepam and [35S]TBPS binding assays. For [3H]flunitrazepam binding to synaptic membranes, GABA gave an EC50=2.0 μM and at saturation (100 μM) an enhancement of 122%. This stimulation was completely blocked by the GABA antagonist SR95531. In contrast, neither GABA nor SR95531 had a significant effect on [3H]flunitrazepam binding to CCVs, indicating that the allosteric interaction between GABA and benzodiazepine binding sites is abolished. Likewise, GABA displaced nearly all of the [35S]TBPS binding to synaptic membranes but had no effect on binding to coated vesicles, indicating that coupling between the GABA binding sites and chloride channel is also impaired. Thus GABAA receptors appear to be uncoupled during normal intracellular trafficking via coated vesicles. The presence of major GABAA receptor subunits on these particles was verified by quantitative immunoblotting. Relative to the levels in synaptic membranes, CCVs contained 110±14% and 29.5±3.8%, respectively, of the immunoreactivity for GABAA receptor β2 and α1 subunits. Thus, in comparison to GABAA receptors on synaptic membranes, those on CCVs have a reduced α1/β2-subunit ratio. It may be suggested that a selective decline in the content of α1 subunits in coated vesicles could in part account for GABAA receptor uncoupling. 相似文献
2.
3.
The acute effects of γ-vinyl-GABA (GVG) on GABAergic inhibition were investigated in the hippocampal slice preparation using the paired-pulse test of inhibition during extracellular recordings. Superfusion of GVG (100–500 μM) for 60 min resulted in a concentration-dependent decrease in GABAergic inhibition. Slices superfused with higher concentrations of GVG (0.5–1 mM) were hyperexcitable as demonstrated by the appearance of multiple spikes. Binding studies showed that GVG (1 mM) had no effect on the binding of [3H]flunitrazepam or [3H]TBOB and displaced no more than 15% of specific [3H]GABA binding, which indicates that GVG-induced disinhibition is not mediated through an action at the GABAA receptor complex. Consistent with this suggestion is the finding that GVG (500 μM) had little effect on the inhibition of the orthodromically evoked CA1 population spike produced by the GABAA receptor agonist muscimol (10 μM), whereas this inhibition was considerably attenuated by the GABAA receptor antagonist, bicuculline methiodide (5 μM). The results of this study suggest that the acute actions of GVG on the GABAergic neurotransmitter system are not involved in its anticonvulsant effect. 相似文献
4.
Damijana M. Juri Marija
arman-Kr
an 《International journal of developmental neuroscience》2001,19(7):5
In astrocytes, nerve growth factor (NGF) synthesis and secretion is stimulated by the cytokine interleukin-1β (IL-1β). In the present study, the role of IL-1 receptor binding sites in the regulation of NGF release was evaluated by determining the pharmacological properties of astroglially localized IL-1 receptors, and, by comparing the effects of both the agonists (IL-1α and IL-1β) and the antagonist (IL-1ra)—members of the IL-1 family on NGF secretion from rat neonatal cortical astrocytes in primary culture. Using receptor-binding studies, binding of [125I] IL-1β to cultured astrocytes was saturable and of high affinity. Mean values for the KD and Bmax were calculated to be 60.7±7.4 pM and 2.5±0.1 fmol mg-1 protein, respectively. The binding was rapid and readily reversible. IL-1 receptor agonists IL-1α (Ki of 341.1 pM) and IL-1β (Ki 59.9 pM), as well as the antagonist IL-1ra (Ki 257.6 pM), displaced specific [125I] IL-1β binding from cultured astrocytes in a monophasic manner. Anti-IL-1RI antibody completely blocked specific [125I] IL-1β binding while anti-IL-1RII antibody had no inhibitory effect. Exposure of cultured astrocytes to IL-1α and IL-1β revealed the functional difference between the agonists in influencing NGF release. In contrast to IL-1β (10 U/ml), which caused a 3-fold increase in NGF secretion compared to control cells, IL-1α by itself had no stimulatory action on NGF release. The simultaneous application of IL-1α and IL-1β elicited no additive response. IL-1ra had no effect on basal NGF release but dose-dependently inhibited the stimulatory response induced by IL-1β. We concluded that IL-1β-induced NGF secretion from cultured rat cortical astrocytes is mediated by functional type I IL-1 receptors, whereas IL-1α and IL-1ra, in spite of their affinity for IL-1RI, have no effect on NGF secretion from these cells. Type II IL-1R is not present on rat neonatal cortical astrocytes. 相似文献
5.
S. Adjeround M.C. Tonon M. Lamacz E. Leneveu M.E. Stoeckel M.L. Tappaz L. Cazin J.M. Danger C. Bernard H. Vaudry 《Brain research bulletin》1986,17(5)
Measurement of glutamate decarboxylase (GAD) activity in the intermediate lobe of the frog pituitary and brain showed that neurointermediate lobe extracts represented 12% of the GAD activity detected in the whole brain. No significant activity was measured in distal lobe extracts. Immunocytochemical studies revealed GAD-containing fibers among the parenchyma! cells of the pars intermedia. The localization of GAD-like material in the intermediate lobe of the frog pituitary suggested a possible role of γ-aminobutyric acid (GABA) in the regulation of melanotropic cell secretion. Administration of GAB A (10−6 to 10−4 M), to perifused neurointermediate lobes caused a brief stimulation of alpha-melanocyte stimulating hormone (α-MSH) release followed by an inhibition. Picrotoxin (10−4 M), a Cl− channel blocker, abolished only the stimulatory effect of GAB A (10−4 M), whereas bicuculline (10−4 M), a specific antagonist of GABAA receptors, totally inhibited the effects of GABA (both stimulatory and inhibitory phases). Bicuculline induced by itself a slight stimulation of α-MSH release, suggesting that GABA-ergic nerve fibers present in the intermediate lobe are functionally active in vitro. The GABAA agonist muscimol (10−7 to 10−4 M) mimicked the biphasic effect of GABA on α-MSH release. Administration of baclofen, a specific GABABB agonist (10−7 to 10−4 M) induced a dose-dependent inhibition of α-MSH secretion. In contrast to GABA or muscimol, baclofen did not cause any stimulatory effect whatever the dose. Taken together these result suggested that GABAA and GABab receptors were present on frog melanotrophs. Since bicuculline totally inhibited GABA effects (stimulation and inhibition) on α-MSH release, it appears however that the effect of GABA is mainly achieved through activation of G AB AA receptors. 相似文献
6.
We have studied the effects of phenoxybenzamine, an irreversible α-adrenoceptor antagonist on the binding of the α-adrenoceptor ligands [3H]prazosin and [3H]clonidine to rabbit brain membranes. Where possible changes in binding were related to changes in central α-adrenoceptor function. Phenoxybenzamine showed a similar α1/α2-adrenoceptor selectivity in the brain to that previously reported in the periphery. Much higher doses were required to reduce specific clonidine binding and to interfere with the hypotensive response to intracisternal clonidine than to reduce specific prazosin binding. Recovery of binding site number of both α1- and α2-adrenoceptor selective ligands was slower than in peripheral tissues (heart and spleen). Recovery was log linear and the half time (t1/2) for recovery of the maximum number of specific prazosin and clonidine binding sites in forebrain was 10.8 ± 2.6days and 6.1 ± 0.1days and in hindbrain 13.3 ± 3.1days and 4.6 ± 1.8days, respectively.t1/2 for recovery of the in vivo hypotensive response to intracisternal clonidine was 2.7 ± 1.0days. Recovery of this response was attenuated by treatment with the inhibitor of protein synthesis, 5-fluorouracil. This suggests that recovery after phenoxybenzamine in brain, as in the periphery, may depend at least in part on synthesis of new receptor protein. The recovery of brain adrenoceptor number after phenoxybenzamine may be an index of receptor turnover and is much slower in brain than in heart and spleen. 相似文献
7.
To support a postulated neurotransmitter character of γ-aminobutyric acid (GABA) in the vertebrate vestibule, [3H]GABA binding was measured in a crude membrane preparation of chick inner ear ampullary cristae. In the absence of divalent cations bound [3H]GABA was displaced by unlabeled GABA, muscimol or bicuculline, but it was not displaced by (±)-baclofen. A single population of [3H]GABA binding sites with an equilibrium constant of 19.4 nM and a maximum binding capacity of 0.58 pmol/mg protein was found. These results suggest the possible existence of a synaptic GABAA receptor in the chick inner ear membranes and sustain the neurotransmitter role of GABA in the chick vestibule. 相似文献
8.
By using an animal model of parkinsonism, we examined the expression of GABAA receptor (R) and metabotropic glutamate receptor (mGluR) 5 in the basal ganglia after transplantation with dopamine-rich tissue. The adult rats were unilaterally lesioned by the injection of 6-hydroxydopamine to their left medial forebrain bundles. At 5–10 weeks following the dopaminergic denervation, the levels of GABAAR in the left caudate-putamen and globus pallidus were about 20 and 16% lower than that of the right intact (control) sides, as shown by [3H]flunitrazepam binding autoradiography on the brain sections. However, the receptor density increased to around 132 and 130% of control levels in the entopeduncular nucleus and substantia nigra pars reticulata of the lesioned sides. Furthermore, in situ hybridization analysis exhibited parallel trends of changes in the levels of the GABAAR α1 and α2 subunit and mGluR5 mRNAs in the neurons of the brain regions with that of the proteins detected by the binding assay. A number of the rats 5 weeks postlesion were transplanted with the ventral mesencephalon of the embryonic rat into their left striata. Five weeks later, the changes in the [3H]flunitrazepam binding seemed to be recovered by approximately 50–63% on the grafted sides of the areas. Moreover, the transplantation appeared to produce a nearly complete reversal of the lesion-induced alterations in the levels of the mRNAs. Thus, the data indicate the mechanism of gene regulation for the modified expression of the receptors and could implicate the participation of the receptors in the pathogenesis of Parkinson's disease. 相似文献
9.
In this study, we investigated the modulatory effect of 5α-pregnan-3α-ol-20-one, a neurosteroid, on the binding characteristics of [
]flunitrazepam (2 nM), [
]muscimol (5 nM), and 4 nM [
]t-butylbicyclophosphorothionate (TBPS) in cerebral cortex, cerebellum, and hippocampus of control, ethanol-dependent, and ethanol-withdrawn rats. 5α-Pregnan-3α-ol-20-one potentiated the binding of [
]flunitrazepam and [
]muscimol in all the rat brain regions investigated in this study. There was a significant increase in the maximal potentiation of [
]flunitrazepam as well as [
]muscimol binding (Emax) in the ethanol-dependent rat cerebellum as compared to control group (p<0.025). Furthermore, 5α-pregnan-3α-ol-20-one elicited a biphasic response, i.e., it potentiated the binding of [
]TBPS at lower concentrations (100 nM) and inhibited the binding at higher concentrations (>100 nM). There was a significant higher inhibition of [
]TBPS binding (−Emax) by 5α-pregnan-3α-ol-20-one in the hippocampus of ethanol-dependent as well as ethanol-withdrawn rats (p<0.025). These observations suggest that the neurosteroid binding site associated with the γ-aminobutyric acidA (GABAA) receptors in cerebellum and hippocampus plays an important role during ethanol-dependence and ethanol-withdrawal, and some of the changes following ethanol dependence and its withdrawal may be mediated through the neurosteroid binding site. 相似文献
10.
Thomas S. King Daniel Potter Inn Soo Kang Catherine Norris Eileen Chen Robert S. Schenken Martin A. Javors 《Brain research》1999,824(1)
Immortalized GT1–7 neurons were used to characterize the effect of muscimol, a GABAA receptor agonist, to enhance pulsatile gonadotropin-releasing hormone (GnRH) release. GT1–7 neurons were grown on Cytodex-3 beads and placed in special superfusion microchambers. The cells were superfused at a rate of 6.2 ml·h−1 with Media 199 (pH 7.35) using a commercially available perfusion system. After a pre-muscimol period of 120 min, the cells were exposed for 5 min to 0.35, 1, 5 or 10 μM muscimol or 5 μM muscimol+20 μM of the GABAA receptor antagonist, bicuculline. Following removal of the muscimol (and bicuculline, in the case of the latter experiment), the superfusion was continued for another 115 min. Sample fractions were collected at 5 min intervals throughout the perfusion. Basal GnRH release from the GT1–7 neurons was pulsatile with an average interpulse interval of 45.4±0.5 min and an average pulse amplitude of 191.5±22.6 pg·min·ml−1. Our results also demonstrated that the GABAA receptor agonist, muscimol, enhances pulsatile GnRH release from GT1–7 neurons in culture. The response to muscimol was saturable and concentration-dependent with an EC50 of 0.47 μM. The effects of 5 μM muscimol to increase GnRH pulsatility were blocked by co-exposure to the GABAA receptor antagonist, bicuculline. The average GnRH interpulse intervals were 41.7±1.8 min, 32.5±2.9 min, 30.6±0.7 min and 25.5±0.4 min in the period following exposure to 0.35, 1, 5 and 10 μM of muscimol, respectively (post-muscimol period). GnRH pulse amplitude (mean-area for each pulse) was increased during exposure to muscimol but not during the pre- or post-muscimol periods. The GABAA receptor antagonist, bicuculline, itself had no effect on pulsatile GnRH release. These results are consistent with previously published reports suggesting that activation of the GABAA receptor stimulates hypothalamic GnRH release in embryonic and neonatal animals. 相似文献
11.
Multiple subtypes of GABAA receptors are expressed in the rat central nervous system (CNS). To determine the distribution and proportion of α5 subunit containing receptors, quantitative autoradiographic analyses were performed with both [3H]L-655,708 and [3H]Ro15-1788, an α5 selective and a non selective benzodiazepine binding site ligand, respectively. High densities of [3H]L-655,708 binding sites were observed in hippocampus and olfactory bulb, where α5 receptors accounted for 20–35% of total [3H]Ro15-1788 binding sites. Low levels of [3H]L-655,708 sites were associated with the cortex as well as amygdala, thalamic, hypothalamic and midbrain nuclei. These observations indicate that although [3H]L-655,708 binding sites have an overall low expression in rat CNS, they may contribute significantly to GABAergic inhibition in specific brain regions. 相似文献
12.
Xiao Xiao Thomas J. McCown Juan Li George R. Breese A. Leslie Morrow R. Jude Samulski 《Brain research》1997,756(1-2)
In the inferior colliculus, adeno-associated virus (AAV) vectors are capable of gene transfer and stable, long-term expression, but it remained to be shown if this in vivo gene transfer could alter focal seizure sensitivity in the inferior colliculus. Because GABA receptors directly modulate inferior collicular seizures, AAV vectors were constructed with a cytomegalovirus (CMV) promoter and a truncated, human GABAA α1 cDNA in both the sense and antisense orientations. Seven days after collicular microinjection of the sense vectors (1 μl; 3×109 particles/μl), neurons exhibited GABAA α-like immunoreactivity in amounts far exceeding endogenous concentrations. Unilateral or bilateral sense vector infusion had no effect on inferior collicular seizure parameters or on [3H]zolpidem binding. In contrast, bilateral infusion of the antisense AAV-GABAA α1 vector (1 μl; 3×108 particles/μl) caused a 137% increase in the seizure duration. Moreover, unilateral antisense vector infusion produced a localized, 48% decrease in [3H]zolpidem binding. Thus, in the inferior colliculus, antisense AAV-CMV vectors can reduce a specific receptor subunit protein and change receptor function that directly influences in vivo seizure sensitivity. 相似文献
13.
The potential heterogeneity in the allosteric coupling between GABA and ω1 binding sites within the native GABAA receptor complex has been evaluated in the rat by measuring the enhancement by GABA of [3H]zolpidem binding to ω1 site in membranes from three rat brain structures (neocortex, cerebellum and hippocampus) and brain sections. The maximal stimulatory effect of GABA was significantly higher (265 ± 47%) in cortical membranes than in cerebellar (165 ± 48%) or in hippocampal (118 ± 17%) membranes. These differences are not due either to the presence of different amounts of residual GABA in the membrane preparations or to the labeling, in presence of GABA, of binding sites other than ω1 since: (1) the pharmacological properties of the [3H]zolpidem binding sites were similar in the three regions; (2) the degree of allosteric enhancement was unrelated to the relative proportion of ω1 sites in each structure; and (3) GABA did not increase the Bmax for [3H]zolpidem. Regional differences in the effect of 100 μM GABA on [3H]zolpidem binding were also observed by quantitative autoradiography. Regions where the strongest (3–4-fold) effects of GABA in [3H]zolpidem binding were observed included the substantia nigra, lateral geniculate body, olfactory tubercule and red nucleus. A large increase in [3H]zolpidem binding was also demonstrated in the cingulate and frontoparietal cortices with higher effects in deep (4.2-fold) rather than in superficial layers (3.3-fold). Heterogeneous subregional increases in [3H]zolpidem binding in the presence of GABA were quantified within the cerebellum, hippocampus and superior colliculus. In the cerebellum the effect of this neurotransmitter was larger in the molecular (3.8-fold) than in the granular (2.2-fold) layer. In the hippocampus the effect of GABA was also heterogeneous with larger increases in CA1 and CA2 fields (3.5-fold) than in CA3 field (2.2-fold) and dentate gyrus (2.5-fold). Finally in the deep layers of the superior colliculus GABA stimulation of [3H]zolpidem binding was greater than the superficial layer. In the other structures examined the GABA-induced increase in [3H]zolpidem binding was less than 3-fold. The smallest stimulations were quantified in the entorhinal cortex (2.1-fold), amygdala (2.4-fold) and nucleus accumbens (1.7-fold). These results suggest that [3H]zolpidem sites are associated to, at least, two GABAA receptor subtypes that can be differentiated by their allosteric interaction between GABA and [3H]zolpidem sites. 相似文献
14.
Mariano Sebastin Viapiano Alba Marta Mitridate de Novara Sara Fiszer de Plazas Carlos Eduardo Bozzini 《Brain research》2001,894(1)
The central nervous system is severely affected by hypoxic conditions, which produce alterations in neural cytoarchitecture and neurotransmission, resulting in a variety of neuropathological conditions such as convulsive states, neurobehavioral impairment and motor CNS alterations. Some of the neuropathologies observed in hypobaric hypoxia, corresponding to high altitude conditions, have been correlated with a loss of balance between excitatory and inhibitory neurotransmission, produced by alterations in glutamatergic and GABAergic receptors. In the present work, we have studied the effect of chronic hypobaric hypoxia (506 hPa, 18 h/day×21 days) applied to adult male mice on GABAA receptors from cerebral cortex, to determine whether hypoxic exposure may irreversibly affect central inhibitory neurotransmission. Saturation curves for [3H]GABA specifically bound to GABAA receptors in isolated synaptic membranes showed a 30% decrease in maximal binding capacity after hypoxic exposure (Bmax control, 4.70±0.19, hypoxic, 3.33±0.10 pmol/mg protein), with no effect on GABA binding sites affinity (Kd control: 159.3±13.3 nM, hypoxic: 164.2±15.1 nM). Decreased Bmax values were observed up to the 10th post-hypoxic day, returning to control values by the 15th post-hypoxic day. Pharmacological properties of GABAA receptor were also affected by hypoxic exposure, with a 45 to 51% increase in the maximal effect by positive allosteric modulators (pentobarbital and 5α-pregnan-3α-ol-20-one). We conclude that long-term hypoxia produces a significant but reversible reduction on GABA binding to GABAA receptor sites in cerebral cortex, which may reflect an adaptive response to this sustained pathophysiological state. 相似文献
15.
Catecholamine receptor binding sites were measured in discrete hypothalamic nuclei or regions as well as in certain extrahypothalamic areas of the adult male rat. For each assay, discrete areas were microdissected from frozen tissue sections and pooled from several animals. Specific high affinity binding sites were assessed at fixed ligand concentrations for [3H]p-aminoclonidine (PAC) and [3H](2-C 2′,6′-(CH3O)2 phenoxyethylamino)-methylbenzodioxan (WB-4101) for α-adrenergic receptor sites, for [3H]dihydroalprenolol (DHA) for β-adrenergic receptor sites, and for [3H]2-amino-6, 7-dihydroxy-1,2,3,4-tetrahydronaphtalene (ADTN) and [3H]spiroperidol in the presence of cinanserin for dopaminergic receptor sites.Regional variations in [3H]WB-4101 binding were relatively small in magnitude, with most hypothalamic and extrahypothalamic areas possessing between 60 and 90% of the binding in frontal cortex. [3H]PAC binding showed a wider range of binding density across brain areas than did [3H]WB-4101, but, in general, variations in [3H]PAC binding paralleled those in [3H]WB-4101 binding. In hypothalamus, binding was characterized as being predominantly to α1-receptors in the of [3H]WB-4101 and to α2-receptors in the case of [3H]PAC. The medial hypothalamic areas exhibited a somewhat higher density of these α-adrenergic sites than did the lateral hypothalamus (perifornical hypothalamus and medial forebrain bundle). Also, the ratio of [3H]PAC to [3H]WB-4101 binding differed in different hypothalamic areas, ranging from 1.5:1 to 4:1. The median eminence was exceptional in that it contained appreciable [3H]PAC but no significant [3H]WB-4101 binding sites at the radioligand concentrations tested. Binding of [3H]DHA to β-adrenergic receptors varied over approximately a 3-fold range in the different hypothalamic areas, with binding highest in the medial forebrain bundle and the medial preoptic area, and lowest in the periventricular, dorsomedial and posterior hypothalamic nuclei, the median eminence and the zona incerta. The ratio of β-adrenergic to α-adrenergic binding sites was generally lower in the medial than in the lateral hypothalamic areas and higher in the extrahypothalamic areas examined than in the hypothalamus. With regard to [3H]spiroperidol and [3H]ADTN binding to dopaminergic sites, the striatum, nucleus accumbens and olfactory tubercle showed a greater density of [3H]spiroperidol than of [3H]ADTN sites, in contrast to the hypothalamus where [3H]ADTN binding was more predominant. Within the hypothalamus, [3H]ADTN binding was relatively uniform, while [3H]spiroperidol binding was quite high in four hypothalamic areas (lateral perifornical area, medial forebrain bundle, paraventricular and dorsomedial nuclei), intermediate in the median eminence and arcuate nucleus, and low or not detectable in all other hypothalamic areas. 相似文献
16.
David C. U'Prichard Terry D. Reisine Stephen T. Mason Hans C. Fibiger Henry I. Yamamura 《Brain research》1980,187(1)
Bilateral lesion of the ascending noradrenergic fibers in the dorsal bundle of adult Wistar rats with 4 μg 6-hydroxydopamine caused extensive depletion of norepinephrine in all forebrain areas, but led to a 54% increase in norepinephrine levels in the cerebellum. β-Adrenergic receptor binding of [3Hdihydroalprenolol was significantly increased in all forebrain areas depleted of norepinephrine except hypothalamus. The increase in [3Hdihydroalprenolol binding was due to 62% and 34% increases in the number of β-receptor sites in the frontal cerebral cortex and hippocampus respectively. Binding of [3HWB-4101 toα1-adrenergic receptors after dorsal bundle lesion was augmented generally to a lesser extent than β-receptor binding, with significantly increased numbers of sites only in the frontal cortex (74%), thalamus (20%) and septum. Bothα1-andβ-receptor binding sites were reduced in number by 25–28% in the cerebellum of dorsal bundle-lesioned rats, whereas intraventricular administration of 6-hydroxydopamine to adult rats, which depletes norepinephrine in the cerebellum by 96%, increased cerebellarα1-andβ-receptor binding by 33–40%. Binding of [3Hclonidine to forebrainα2-adrenergic receptors was significantly elevated in the frontal cortex, but reduced in the amygdala and septum, after dorsal bundle lesion. 相似文献
17.
High density neuronal cultures from rat E18 hippocampus and cortex have been characterised with respect to cholinergic binding sites. No specific binding of [3H]nicotine or [3H]cyttine to live cells in situ was detected, although the limit for detection was estimated to be 30 fmol/mg protein. Muscarinic binding sites labelled with [3H]QNB were present at a density of 0.75 pmol/mg protein. [125I]α-Bungarotoxin (αBgt) bound to hippocampal cultures with a Bmax of 128 fmol/mg protein and a Kd of 0.6 nM; cortical cultures expressed five times fewer [125I]α-Bgt binding sites. Fluorescence cytochemistry with rhodamine-α-Bgt indicated that 95% of hippocampal neurons were labelled, compared with only 36% of cortical neurons. Average densities of 4 × 104 and 2 × 104 binding sites/cell were calculated for hippocampal and cortical cultures, respectively. Double labelling experiments with mAb307 (which recognises the rat α7 nicotinic receptor subunit) and rhodamine-α-Bgt gave coincident labelling patterns, supporting the correlation between the α7 subunit and Bgt-sensitive neuronal nicotinic receptor. Treatment of hippocampal cultures with 10 μM nicotine for 14 days elicited a 40% increase in the numbers of [125I]α-Bgt binding sites, mimicking the up-regulation observed in vivo studies. Primary cultures offer a useful in in vitro system for investigating the expression and regulation of brain α-Bgt-sensitive receptors. 相似文献
18.
Employing the Nystatin-perforated whole-cell patch-clamp recording technique, the modulatory effects of serotonin (5-HT) on γ-aminobutyric acid (GABA)-activated whole-cell currents were investigated in neurons acutely dissociated from the superficial laminae (laminae I and II) of the rat spinal dorsal horn. The results showed: (1) GABA acted on GABAA receptors and elicited inward Cl− currents (IGABA) at a holding potential (VH) of −40 mV; (2) 5-HT potentiated GABA-induced Cl− current without affecting the reversal potential of IGABA and the apparent affinity of GABA to its receptor; (3) α-methyl-5-HT, a selective agonist of 5-HT2 receptor, mimicked the potentiation effect of 5-HT on IGABA, whereas ketanserine, an antagonist of 5-HT2 receptor, blocked the potentiation effect of 5-HT; (4) Chelerythrine, an inhibitor of protein kinase C, reduced the potentiation effect of 5-HT on IGABA. The present results indicate: (1) The potentiation of 5-HT on IGABA is mediated by 5-HT2 receptor and through a protein kinase-dependent transduction pathway; (2) The interactions between 5-HT and GABA might play an important role in the modulation of nociceptive information transmission at spinal cord level. 相似文献
19.
Vladimir Znamensky Joyce A. Echo Nicole Lamonte Garrison Christian Andr Ragnauth Richard J. Bodnar 《Brain research》2001,906(1-2)
Food intake is significantly increased by administration of μ-selective opioid agonists into the nucleus accumbens, particularly its shell region. Pretreatment with either opioid (μ, δ1, δ2 or κ1) or dopaminergic (D1) receptor antagonists in the nucleus accumbens shell reduce μ opioid agonist-induced feeding. Selective GABAA (muscimol) and GABAB (baclofen) agonists administered into the nucleus accumbens shell each stimulate feeding which is respectively and selectively blocked by GABAA (bicuculline) and GABAB (saclofen) antagonists. The present study investigated whether feeding elicited by the μ-selective opioid agonist, [
-Ala2,NMe4,Gly-ol5]-enkephalin in the nucleus accumbens shell was decreased by intra-accumbens pretreatment with an equimolar dose range of either GABAA or GABAB antagonists, and further, whether general opioid or selective GABA antagonists decreased feeding elicited by GABAA or GABAB agonists in the nucleus accumbens shell. Feeding elicited by the μ-selective opioid agonist was dose-dependently increased following intra-accumbens pretreatment with GABAA (bicuculline) antagonism; this enhancement was significantly blocked by pretreatment with general or μ-selective opioid antagonists. In contrast, μ opioid agonist-induced feeding elicited from the nucleus accumbens shell was dose-dependently decreased by GABAB (saclofen) antagonism. Neither bicuculline nor saclofen in the nucleus accumbens shell altered baseline food intake. Whereas muscimol-induced feeding elicited from the nucleus accumbens shell was reduced by bicuculline and naltrexone, but not saclofen pretreatment, baclofen-induced feeding elicited from the nucleus accumbens shell was reduced by saclofen, but not by bicuculline or naltrexone. These data indicate that GABAA and GABAB receptor subtype antagonists differentially affect feeding elicited by μ opioid receptor agonists within the nucleus accumbens shell in rats. 相似文献
20.
Ewa Malatynska Cortland Miller Nancy Schindler Alicia Cecil Angela Knapp Glenda Crites Howell Rogers 《Brain research》1999,851(1-2)
WSS-1 cells expressing (α1γ2)GABAA receptors show an augmented 36Cl− response to GABA in the presence of amitriptyline that is increased by flumazenil, unlike augmentation by diazepam which is blocked by flumazenil. This amitriptyline effect is opposite to the inhibition of GABA-stimulated 36Cl influx manifested in membrane vesicles prepared from drug-naive rats or submissive rats (a model of depression) but is similar to that seen in tissue from amitriptyline-treated rats or dominant rats. The results suggest a novel mechanism of antidepressant drug action having a delayed onset. 相似文献