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1.
Lynch BA Matagne A Brännström A von Euler A Jansson M Hauzenberger E Söderhäll JA 《Biochemical and biophysical research communications》2008,375(4):491-495
The synaptic vesicle protein 2A (SV2A), the brain-binding site of the anti-epileptic drug levetiracetam (LEV), has been characterized by Protein Tomography™. We identified two major conformations of SV2A in mouse brain tissue: first, a compact, funnel-structure with a pore-like opening towards the cytoplasm; second, a more open, V-shaped structure with a cleft-like opening towards the intravesicular space. The large differences between these conformations suggest a high degree of flexibility and support a valve-like mechanism consistent with the postulated transporter role of SV2A. These two conformations are represented both in samples treated with LEV, and in saline-treated samples, which indicates that LEV binding does not cause a large-scale conformational change of SV2A, or lock a specific conformational state of the protein. This study provides the first direct structural data on SV2A, and supports a transporter function suggested by sequence homology to MFS class of transporter proteins. 相似文献
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《Peptides》2017
The intracerebroventicular (i.c.v.) administration of glucagon-like peptide-2 (GLP-2) to rodents was shown to have antidepressant-like effects in imipramine-resistant depression-model mice. In order to utilize GLP-2 as a clinical treatment tool for depression, we herein focused on the intranasal delivery that is non-invasive approach, because the i.c.v. administration is invasive and impractical. In the present study, we prepared a GLP-2 derivative containing cell penetrating peptides (CPPs) and a penetration accelerating sequence (PAS) (PAS-CPPs-GLP-2) for the intranasal (i.n.) administration. PAS-CPPs-GLP-2 (i.n.) exhibited antidepressant-like effects in the forced-swim test (FST) and tail suspension test (TST) in naïve mice as well as adrenocorticotropic hormone (ACTH) treated-mice. However, PAS-CPPs-GLP-2 (i.v.) and the GLP-2 derivative containing CPPs without a PAS (CPPs-GLP-2) (i.n.) did not affect the immobility time in the mouse FST. Moreover, fluorescein isothiocyanate (FITC)-labeled PAS-CPPs-GLP-2 (i.n.), but not FITC-labeled CPPs-GLP-2 (i.n.) was distributed through the mouse brain after the FST session. These results suggest that PAS-CPPs-GLP-2 is effective for i.n. delivery to the brain, and may be useful in the clinical treatment of major depression. 相似文献
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杏仁核注射红藻氨酸诱导的边缘叶发作癫痫大鼠海马 Bad去磷酸化和Bcl-2表达上调 总被引:3,自引:1,他引:3
应用红藻氨酸(kainic acid,KA)诱导的人鼠边缘叶发作癫痫模型,检测Bad(Bcl-2-associated death protein)、14-3-3、磷酸化Bad、Bcl-XL和Bcl-2在癫痫人鼠海码神经元的表达。单侧杏仁核内注射KA诱导癫痫发作,持续记录脑电和局部脑血流(regional cerebral blood flow,r-CBF),发作1h后静脉注射30mg/kg安定终止发作,然后分别用cresyl violet染色和TUNEL染色观察海马神经元存活和凋亡的变化;用免疫荧光、Western blot和免疫沉淀俭测海马Bad、14-3-3、磷酸化Bad、Bcl-XL和Bcl-2的表达。结果表明,发作终止8h时出现TUNEL阳忡细胞,24h时达高峰;发作诱导Bad去磷酸化,去磷酸化的Bad与分了伴侣蛋F114-3-3解离,然后Bad与Bcl-XL结合:磷酸化Bad表达减少而Bcl-2表达增加;发作前后r-CBF无明显变化。以上结果提示,癫痫发作诱导Bad的去磷酸化和Bcl-2表达上调,Bad的上磷酸化可能具有损伤作用,而Bcl-2的表达上调则对癫痫神经元损伤具有保护作用,但与脑缺血无关。 相似文献
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Pinto-Duarte A Coelho JE Cunha RA Ribeiro JA Sebastião AM 《Journal of neurochemistry》2005,93(3):595-604
Adenosine, a neuromodulator of the CNS, activates inhibitory-A1 receptors and facilitatory-A2A receptors; its synaptic levels are controlled by the activity of bi-directional equilibrative nucleoside transporters. To study the relationship between the extracellular formation/inactivation of adenosine and the activation of adenosine receptors, we investigated how A1 and A2A receptor activation modifies adenosine transport in hippocampal synaptosomes. The A2A receptor agonist, CGS 21680 (30 nm), facilitated adenosine uptake through a PKC-dependent mechanism, but A1 receptor activation had no effect. CGS 21680 (30 nm) also increased depolarization-induced release of adenosine. Both effects were prevented by A2A receptor blockade. A2A receptor-mediated enhancement of adenosine transport system is important for formatting adenosine neuromodulation according to the stimulation frequency, as: (1) A1 receptor antagonist, DPCPX (250 nm), facilitated the evoked release of [(3)H]acetylcholine under low-frequency stimulation (2 Hz) from CA3 hippocampal slices, but had no effect under high-frequency stimulation (50 Hz); (2) either nucleoside transporter or A2A receptor blockade revealed the facilitatory effect of DPCPX (250 nm) on [3H]acetylcholine evoked-release triggered by high-frequency stimulation. These results indicate that A2A receptor activation facilitates the activity of nucleoside transporters, which have a preponderant role in modulating the extracellular adenosine levels available to activate A1 receptors. 相似文献
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Adenosine A2A receptors are required for glutamate mGluR5‐ and dopamine D1 receptor‐evoked ERK1/2 phosphorylation in rat hippocampus: involvement of NMDA receptor 下载免费PDF全文
Paraskevi Krania Eleni Dimou Maria Bantouna Stylianos Kouvaros Eirini Tsiamaki Costas Papatheodoropoulos Konstantinos Sarantis Fevronia Angelatou 《Journal of neurochemistry》2018,145(3):217-231
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为了探究A33核心启动子结肠癌特异性及SV40增强子对其转录水平的影响,该研究通过构建A33核心启动子和带SV40增强子的A33核心启动子(eA33)的荧光素酶报告基因载体pGL3-A33和pGL3-eA33,与内参照pRL-SV40质粒共转染至不同的细胞系中,利用双荧光素酶检测系统检测分析了A33和eA33启动子在不同细胞系中的转录活性。结果显示,A33核心启动子在结肠癌细胞系中具有转录活性低,但结肠癌特异性好的特点,而在其他类型癌细胞中基本没有活性。同时发现,eA33在各类癌细胞中的转录水平与A33相比,均呈大幅度提高,有显著性差异(P<0.01),但SV40增强子能显著增强A33启动子转录活性的同时减低了其结肠癌特异性。这为靶向癌症基因—病毒治疗策略在结肠癌的生物治疗应用中寻找结肠癌特异性的启动子奠定了研究基础。 相似文献
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Tebano MT Martire A Potenza RL Grò C Pepponi R Armida M Domenici MR Schwarzschild MA Chen JF Popoli P 《Journal of neurochemistry》2008,104(1):279-286
Brain-derived neurotrophic factor (BDNF), a member of neurotrophin family, enhances synaptic transmission and regulates neuronal proliferation and survival. Both BDNF and its tyrosine kinase receptors (TrkB) are highly expressed in the hippocampus, where an interaction with adenosine A2A receptors (A2A Rs) has been recently reported. In the present paper, we evaluated the role of A2A Rs in mediating functional effects of BDNF in hippocampus using A2A R knock-out (KO) mice. In hippocampal slices from WT mice, application of BDNF (10 ng/mL) increased the slope of excitatory post-synaptic field potentials (fEPSPs), an index of synaptic facilitation. This increase of fEPSP slope was abolished by the selective A2A antagonist ZM 241385. Similarly, genetic deletion of the A2A Rs abolished BDNF-induced increase of the fEPSP slope in slices from A2A R KO mice The reduced functional ability of BDNF in A2A R KO mice was correlated with the reduction in hippocampal BDNF levels. In agreement, the pharmacological blockade of A2 Rs by systemic ZM 241385 significantly reduced BDNF levels in the hippocampus of normal mice. These results indicate that the tonic activation of A2A Rs is required for BDNF-induced potentiation of synaptic transmission and for sustaining a normal BDNF tone in the hippocampus. 相似文献
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Tebano MT Martire A Rebola N Pepponi R Domenici MR Grò MC Schwarzschild MA Chen JF Cunha RA Popoli P 《Journal of neurochemistry》2005,95(4):1188-1200
Hippocampal metabotropic glutamate 5 receptors (mGlu5Rs) regulate both physiological and pathological responses to glutamate. Because mGlu5R activation enhances NMDA-mediated effects, and given the role played by NMDA receptors in synaptic plasticity and excitotoxicity, modulating mGlu5R may influence both the physiological and the pathological effects elicited by NMDA receptor stimulation. We evaluated whether adenosine A2A receptors (A(2A)Rs) modulated mGlu5R-dependent effects in the hippocampus, as they do in the striatum. Co-application of the A(2A)R agonist CGS 21680 with the mGlu5R agonist (RS)-2-chloro-s-hydroxyphenylglycine(CHPG) synergistically reduced field excitatory postsynaptic potentials in the CA1 area of rat hippocampal slices. Endogenous tone at A(2A)Rs seemed to be required to enable mGlu5R-mediated effects, as the ability of CHPG to potentiate NMDA effects was antagonized by the selective A(2A)R antagonist ZM 241385 in rat hippocampal slices and cultured hippocampal neurons, and abolished in the hippocampus of A(2A)R knockout mice. Evidence for the interaction between A(2A)Rs and mGlu5Rs was further strengthened by demonstrating their co-localization in hippocampal synapses. This is the first evidence showing that hippocampal A(2A)Rs and mGlu5Rs are co-located and act synergistically, and that A(2A)Rs play a permissive role in mGlu5R receptor-mediated potentiation of NMDA effects in the hippocampus. 相似文献
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Garry G Sedgwick Daniel G Hayward Barbara Di Fiore Mercedes Pardo Lu Yu Jonathon Pines Jakob Nilsson 《The EMBO journal》2013,32(2):303-314
The Anaphase Promoting Complex/Cyclosome (APC/C) in complex with its co‐activator Cdc20 is responsible for targeting proteins for ubiquitin‐mediated degradation during mitosis. The activity of APC/C–Cdc20 is inhibited during prometaphase by the Spindle Assembly Checkpoint (SAC) yet certain substrates escape this inhibition. Nek2A degradation during prometaphase depends on direct binding of Nek2A to the APC/C via a C‐terminal MR dipeptide but whether this motif alone is sufficient is not clear. Here, we identify Kif18A as a novel APC/C–Cdc20 substrate and show that Kif18A degradation depends on a C‐terminal LR motif. However in contrast to Nek2A, Kif18A is not degraded until anaphase showing that additional mechanisms contribute to Nek2A degradation. We find that dimerization via the leucine zipper, in combination with the MR motif, is required for stable Nek2A binding to and ubiquitination by the APC/C. Nek2A and the mitotic checkpoint complex (MCC) have an overlap in APC/C subunit requirements for binding and we propose that Nek2A binds with high affinity to apo‐APC/C and is degraded by the pool of Cdc20 that avoids inhibition by the SAC. 相似文献
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近年来,随着全球气候变化和人为影响加剧,半干旱草地生态系统的碳循环受到剧烈影响。半干旱草原区域CO_2模拟研究主要集中于已有观测资料的地区,然而,观测资料缺乏的草原区CO_2通量模拟却鲜少有人研究。因此选择缺通量资料的呼伦贝尔草原地区为主要研究对象,并将VPRM模型应用于缺资料地区,模拟了该区域内2016年的NEE时空分布。结果表明:(1)在特旱年的气候条件下2016年全年都表现为微弱的碳源(全年NEE值为47.27 gC/m~2),且其变化趋势与降水和气温在年内变化趋势相近。(2)空间上,根据趋势来看NEE在空间分布由草原区向草甸区、森林区逐渐降低。基于植被分布情况,不同植被类型的区域碳排放顺序为:克氏针茅草原和大针茅草原羊草草原杂草草甸草原(以线叶菊等为主)。(3)干旱胁迫是该地区表现为碳源的主要原因之一,而且降水与NEE表现出极显著的二次函数关系(R~2=0.938,P0.001),说明了干旱气候条件下,随着月降水量的增加,草原生态系统出现碳源向碳汇转移的趋势。(4)地上生物量(AGB)与GPP和Reco表现出了极显著的正相关关系(R~2分别为0.89和0.9,P0.01),与NEE表现出了极显著的负相关关系(R~2=0.68,P0.01),说明了草原的地上生物量增加能有效地降低二氧化碳排放。 相似文献
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Background, aim, and scope Traditional life cycle impact assessment methodologies have used aggregated characterization factors, neglecting spatial and
temporal variations in regional impacts like photochemical oxidant formation. This increases the uncertainty of the LCA results
and diminishes the ease of decision-making. This study compares four common impact assessment methods, CML2001, Eco-indicator
99, TRACI, and EDIP2003, on their underlying models, spatial and temporal resolution, and the level at which photochemical
oxidant impacts are calculated. A new characterization model is proposed that incorporates spatial and temporal differentiation.
Materials and methods A photochemical air quality modeling system (CAMx-MM5-SMOKE) is used to simulate the process of formation, transformation,
transport, and removal of photochemical pollutants. Monthly characterization factors for individual US states are calculated
at three levels along the cause–effect chain, namely, fate level, human and ecosystem exposure level, and human effect level.
Results and discussion The results indicate that a spatial variability of one order of magnitude and a temporal variability of two orders of magnitude
exist in both the fate level and human exposure and effect level characterization factors for NOx. The summer time characterization factors for NOx are higher than the winter time factors. However, for anthropogenic VOC, the summer time factors are lower than the winter
time in almost half of the states. This is due to the higher emission rates of biogenic VOCs in the summer. The ecosystem
exposure factors for NOx and VOC do not follow a regular pattern and show a spatial variation of about three orders of magnitude. They do not show
strong correlation with the human exposure factors. Sensitivity analysis has shown that the effect of meteorology and emission
inputs is limited to a factor of three, which is several times smaller than the variation seen in the factors.
Conclusions Uncertainties are introduced in the characterization of photochemical precursors due to a failure to consider the spatial
and temporal variations. Seasonal variations in photochemical activity influence the characterization factors more than the
location of emissions. The human and ecosystem exposures occur through different mechanisms, and impacts calculated at the
fate level based only on ozone concentration are not a good indicator for ecosystem impacts.
Recommendations and perspectives Spatial and temporal differentiation account for fate and transport of the pollutant, and the exposure of and effect on the
sensitive human population or ecosystem. Adequate resolution for seasonal and regional processes, like photochemical oxidant
formation, is important to reduce the uncertainty in impact assessment and improve decision-making power. An emphasis on incorporating
some form of spatial and temporal information within standard LCI databases and using adequately resolved characterization
factors will greatly increase the fidelity of a standard LCA.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
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Fujii S Kato H Ito K Itoh S Yamazaki Y Sasaki H Kuroda Y 《Cellular and molecular neurobiology》2000,20(3):331-350
1. Using simultaneous recordings of the field EPSP and the population spike in the CA1 neurons of guinea pig hippocampal slices, we confirmed that delivery of a high-frequency stimulation (tetanus: 100 pulses at 100 Hz) produced robust long-term potentiation of synaptic efficacy (LTP) in two independent components, a synaptic component that increases field excitatory postsynaptic potentials (EPSPs) and a component that results in a larger population spike amplitude for a given EPSP size (E-S potentiation).2. In the same cells, reversal of LTP (depotentiation; DP) in the field EPSP and in the E-S component is achieved by delivering low-frequency afferent stimulation (LFS:1 Hz, 1000 pulses) 20 min after the tetanus.3. When the tetanus or LFS was applied to CA1 inputs in the presence of an adenosine A1 receptor antagonist, 8-cyclopentyltheophylline (1 M), the field EPSP was enhances in LTP and attenuated in DP, while the E-S relationship was not significantly affected in either LTP or DP.4. When similar experiments were performed using an A2 receptor antagonist, CP-66713 (10 M), the field EPSP was blocked in LTP but facilitated in DP, while E-S potentiation was enhanced during both LTP and DP.5. The results show that endogenous adenosine, acting via A1 or A2 receptors, modulates both the synaptic and the E-S components of the induction and reversal of LTP. Based on the results, we discuss the key issue of the contribution of these receptors to the dynamics of neuronal plasticity modification in hippocampal CA1 neurons. 相似文献
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We examined brain phospholipase A2 (PLA2) activity and the expression of enzymes metabolizing arachidonic acid (AA) in cytosolic PLA2 knockout () mice to see if other brain PLA2 can compensate for the absence of cPLA2 alpha and if cPLA2 couples with specific downstream enzymes in the eicosanoid biosynthetic pathway. We found that the rate of formation of prostaglandin E2 (PGE2), an index of net cyclooxygenase (COX) activity, was decreased by 62% in the compared with the control mouse brain. The decrease was accompanied by a 50-60% decrease in mRNA and protein levels of COX-2, but no change in these levels in COX-1 or in PGE synthase. Brain 5-lipoxygenase (5-LO) and cytochrome P450 epoxygenase (cyp2C11) protein levels were also unaltered. Total and Ca2+-dependent PLA2 activities did not differ significantly between and control mice, and protein levels of type VI iPLA2 and type V sPLA2, normalized to actin, were unchanged. These results show that type V sPLA2 and type VI iPLA2 do not compensate for the loss of brain cPLA2 alpha, and that this loss has significant downstream effects on COX-2 expression and PGE2 formation, sparing other AA oxidative enzymes. This suggests that cPLA2 is critical for COX-2-derived eicosanoid production in mouse brain. 相似文献
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Axin2‐mTurquoise2: A novel reporter mouse model for the detection of canonical Wnt signalling 下载免费PDF全文
Jolanda. J. D. de Roo Cor Breukel Amiet R. Chhatta Margot M. Linssen Sandra A. Vloemans Daniela Salvatori Harald M. M. Mikkers Sjef J. Verbeek Frank J.T. Staal 《Genesis (New York, N.Y. : 2000)》2017,55(10)
The canonical Wnt signalling pathway has been implicated in organogenesis and self‐renewal of essentially all stem cell systems. In vivo reporter systems are crucial to assess the role of Wnt signalling in the biology and pathology of stem cell systems. We set out to develop a Turquoise (TQ) fluorescent protein based Wnt reporter. We used a CRISPR‐Cas9 approach to insert a TQ fluorescent protein encoding gene into the general Wnt target gene Axin2, thereby establishing a Wnt reporter mouse similar to previously generated Wnt reporter mice but with the mTurquoise2 gene instead of E. coli‐β‐galactosidase (LacZ). The use of mTurquoise2 is especially important in organ systems in which cells need to a be alive for further experimentation such as in vitro activation or transplantation studies. We here report successful generation of Axin2‐TQ mice and show that cells from these mice faithfully respond to Wnt signals. High Wnt signals were detected in the intestinal crypts, a classical Wnt signalling site in vivo, and by flow cytometry in the thymus. These mice are an improved tool to further elucidate the role of Wnt signalling in vivo. 相似文献
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MeCP2(Methyl CpG binding protein 2)基因突变可导致Rett综合征(Rett syndrome, RTT)。目前已报道的MeCP2敲除小鼠表型与RTT病人症状存在显著差异。为探索MeCP2在脑发育中的作用及其导致RTT的机制,本研究利用CRISPR/Cas9技术构建了MeCP2基因敲除大鼠模型。通过构建靶向敲除MeCP2基因的载体,体外将Cas9 mRNA和sgRNA显微注射到SD大鼠受精卵中,在MeCP2基因exon2中造成移码突变,从而获得MeCP2基因敲除大鼠。利用测序和Western blotting方法鉴定MeCP2敲除大鼠,并对其表型和行为学特征进行分析,发现MeCP2敲除大鼠体重降低,存在焦虑倾向和认知缺陷。本研究成功构建了MeCP2基因敲除大鼠模型,其表型类似人类RTT患者的症状,为后续MeCP2功能研究提供了更好的动物模型。 相似文献