D-型丝氨酸、丝氨酸消旋酶及D-型氨基酸氧化酶在小鼠出生后个体发育中的分布变化及相互关系

目的：探讨小鼠个体发育过程中皮层、纹状体、小脑、心脏、肺、肝脏、脾脏、肾脏和骨骼肌中D-型丝氨酸,丝氨酸消旋酶(SR)及D-型氨基酸氧化酶(DAO)的分布特征及相互关系。方法：RT-PCR法检测SR和DAO mRNA水平;Western blot检测SR蛋白水平;比色法检测DAO酶活力;高效液相法检测游离D-型丝氨酸含量．结果：小鼠出生后早期个体发育过程中皮层、纹状体和小脑内游离D-型丝氨酸含量急剧增加并伴有SR平行增加．在检测组织中,仅小脑和肾脏可检测到DAO酶活力,且随着小脑和肾脏内DAO酶活力的增加,D-型丝氨酸含量急剧下降,并在出生后第三周末降至成年微量水平,但此时DAO酶活力却仍不断增加．结论：哺乳动物体内D-型丝氨酸主要由SR催化生成,但在皮层和纹状体等D-型丝氨酸浓度很高,但缺乏DAO的组织中D-型丝氨酸的降解尚有其它机制参与．小脑和肾脏中的DA0除参与D-型氨基酸代谢外,可能还具有其它的生理功能。     

腹腔注射L-丝氨酸后大鼠梗死侧脑皮质L-丝氨酸与D-丝氨酸含量的测定

目的:制作大鼠永久性脑梗死模型,腹腔注射L-丝氨酸,测定梗死侧大脑皮质L-丝氨酸与D-丝氨酸含量的变化,为应用L-丝氨酸辅助治疗脑梗死提供方法学上的依据。方法:利用邻苯二甲醛及N-乙酰半胱氨酸柱前衍生,高效液相色谱荧光检测法测定脑组织中L-丝氨酸与D-丝氨酸含量。结果:流动相A为82%0.1 mol·L-1磷酸盐缓冲液,流动相B为18%甲醇条件下,L-丝氨酸与D-丝氨酸得到很好的分离;腹腔注射L-丝氨酸1 h后,梗死侧脑皮质L-丝氨酸含量迅速增加,2 h即达高峰[(6.85±0.30)μmol·g-1],为正常脑组织的4倍以上,12 h时仍是正常的2倍以上水平[(3.56±0.22)vs(1.52±0.04)μmol·g-1];D-丝氨酸水平升高缓慢,升高幅度低,6 h时达高峰,为正常的2倍多[(0.75±0.05)vs(0.32±0.01)μmol·g-1]。结论:腹腔注射L-丝氨酸能快速升高梗死侧大脑皮质L-丝氨酸水平,对D-丝氨酸影响相对较小。     

柱前手性衍生化-HPLC法测定人血浆和尿液中D-谷氨酸和D-丝氨酸的浓度

目的:建立手性分离D-谷氨酸和D-丝氨酸的方法,测定人血浆和尿液中D-谷氨酸和D-丝氨酸的浓度。方法:12对极性氨基酸标准液混合后,加入衍生化试剂邻苯二甲醛、N-乙酰-L-半胱氨酸衍生化后,采用高效液相色谱法进样测定。色谱柱为大连依利特HypersilC18,流动相为甲醇-50mmol·L-1醋酸铵缓冲液(pH6.0),梯度洗脱,流速为1.0mL·min-1,柱温为室温,进样量为20μL,荧光检测波长为350nm(激发波长)、450nm(发射波长)。结果:D-谷氨酸、D-丝氨酸检测浓度分别在0.5～20、0.5～80nmol·L-1范围内线性关系良好,平均方法回收率分别为96.4%～103.6%、97.1%～100.5%,日内、日间RSD均<8%。测得正常人血浆中D-谷氨酸和D-丝氨酸平均含量分别为(2.43±0.37)、(1.07±0.11)nmol·mL-1,尿样中未能检测到2种D型氨基酸。结论:本研究建立的手性分离测定D-谷氨酸和D-丝氨酸的分析方法可行,可用于测定人血浆和尿液中二者的浓度。     

胰岛素降低海马谷氨酸和D-丝氨酸含量改善糖尿病大鼠空间学习记忆能力

目的观察胰岛素对糖尿病(DM)大鼠空间学习记忆及海马组织中谷氨酸和D-丝氨酸含量的影响。方法采用尾静脉注射链脲佐菌素(STZ)制备大鼠DM模型。注射STZ第3天建模成功后,大鼠sc给予胰岛素2 U·kg-1,每天1次,持续82 d。定期检测各组大鼠体质量及空腹血糖。第81天进行Morris水迷宫实验,检测大鼠学习记忆能力;实验结束后取海马组织,观察形态变化,并测定谷氨酸和D-丝氨酸含量。结果与正常对照组比较,DM模型组大鼠体质量明显减轻(P<0.01),血糖明显升高(P<0.01),逃避潜伏期明显延长,原平台象限游泳时间显著减少(P<0.01),海马组织中谷氨酸及D-丝氨酸的含量均显著升高(P<0.01)。胰岛素治疗组体质量增加,血糖含量恢复到正常水平。与DM模型组相比,胰岛素治疗组大鼠逃避潜伏期显著缩短(P<0.01),原平台象限游泳时间占总时间百分比显著增加(P<0.01);海马组织中谷氨酸和D-丝氨酸的含量也分别由DM模型组的(1.550±0.054)和(0.084±0.05)mg·g-1下降为胰岛素治疗组的(1.137±0.023)和(0.068±0.004)mg·g-1。结论胰岛素可以改善糖尿病大鼠空间学习记忆能力,这可能与其降低海马组织中谷氨酸和D-丝氨酸含量有关。     

NMDA受体与精神分裂症的研究进展

N-甲基-D-天门冬氨酸受体（NMDA）是中枢神经系统中重要的兴奋性氨基酸受体,在细胞凋亡、学习记忆等生理过程中起着重要的作用。精神分裂症是一种病因未明的以阳性症状、阴性症状、认知功能障碍等为主要临床表现的精神疾病。有研究表明NMDA受体可影响神经元细胞的增殖分化及学习记忆能力,但具体机制还不清楚。因此作用于NMDA受体的抗精神分裂症药物可能是未来研究热点。     

小胶质细胞的发育及在中枢神经系统的功能研究进展

小胶质细胞是一类广泛分布于中枢神经系统(CNS)的免疫细胞,其数量占整个神经胶质细胞数量的5%~20%。小胶质细胞最早由Hortega通过碳酸银法所区分鉴定出来,认为小胶质细胞来源于中胚层,在胚胎发育后期形成血管时侵入脑内。最新研究表明,小胶质细胞来源于卵黄囊原始巨噬细胞。小胶质细胞作为CNS常驻免疫细胞,属于单核巨噬细胞系,是CNS抵御病原入侵的一道重要的免疫防线。静息状态下,小胶质细胞对神经系统起到监视维持内稳态的作用;其活化后,一方面起到吞噬细胞碎片的作用,另一方面在不同病理条件下分泌不同的细胞因子起到神经保护与神经毒性双重作用。本文对小胶质细胞的发育及在CNS中的作用予以综述。     

几种胶质细胞在新生儿脑损伤中作用及其机制的研究进展

新生儿脑损伤严重威胁着儿童生命安全与生活质量,可遗留不同程度的神经系统后遗症,目前尚无完全有效的治疗方法来杜绝这些后遗症的发生。脑组织损伤后,胶质细胞更具活力,在维持大脑稳态中发挥重要作用,因此研究胶质细胞在新生儿脑损伤中的作用具有重要意义。本文就胶质细胞在新生儿脑损伤中作用及其机制的最新研究进展进行综述,以期为新生儿脑损伤的治疗提供新思路。     

丝氨酸蛋白酶抑制剂的研究进展

目的对丝氨酸蛋白酶抑制剂的结构、功能等以及在黄粉虫中参与多种丝氨酸蛋白酶(serine protease,SP)级联的活性调控机制的研究进展进行综述。方法参考国外29篇文献,简述其结构、功能、与其相关的疾病及其研究的模式生物进行总结。结果丝氨酸蛋白酶抑制剂(serpin)是蛋白酶抑制因子中最大也是最重要的超家族,具有一定的特殊结构,serpin使用自杀底物原理与靶酶形成稳定的共价复合物,导致靶酶失活。该类型蛋白超家族成员遗传性结构或异常分泌将导致许多疾病的发生。在黄粉虫中丝氨酸蛋白酶抑制剂参与多种丝氨酸蛋白酶(serine protease,SP)级联的活性调控。结论目前对其生物学特性以及结构与功能关系的研究较多,对于临床应用及其模式生物的研究有待于更深入的研究。     

氟吡啶对脑损伤保护作用的研究

氟吡啶是中枢性非阿片类镇痛药,具有镇痛及其它多各作用。近期研究表明该药亦具有神经元保护作用。本文对氟吡啶作用机制的研究进展作综合评述。     

Notch信号通路与中枢神经系统疾病的研究进展

Notch信号通路在神经元和神经胶质细胞正常生长成熟和中枢神经系统疾病的发生发展中有重要地位。小胶质细胞是中枢神经系统中重要的神经胶质细胞,不但参与大脑的正常发育,而且与炎症性疾病的发生发展密切相关。直到最近才被公认Notch信号通路是小胶质成熟和活化的重要角色。本文综述了一些Notch信号通路在中枢神经系统疾病中的研究进展。     

Converging Evidence on D-Amino Acid Oxidase–Dependent Enhancement of Hippocampal Firing Activity and Passive Avoidance Learning in Rats

BackgroundN-methyl-D-aspartate (NMDA) receptor activation requires the binding of a co-agonist on the glycine-binding site. D-serine is the main endogenous co-agonist of NMDA receptors, and its availability significantly depends on the activity of the metabolic enzyme D-amino acid oxidase (DAAO). Inhibition of DAAO increases the brain levels of D-serine and modulates a variety of physiological functions, including cognitive behavior.MethodsHere, we examined the effects of a novel 4-hydroxypyridazin-3(2H)-one derivative DAAO inhibitor, Compound 30 (CPD30), on passive avoidance learning and on neuronal firing activity in rats.ResultsD-serine administration was applied as reference, which increased cognitive performance and enhanced hippocampal firing activity and responsiveness to NMDA after both local and systemic application. Similarly to D-serine, CPD30 (0.1 mg/kg) effectively reversed MK-801–induced memory impairment in the passive avoidance test. Furthermore, local iontophoretic application of CPD30 in the vicinity of hippocampal pyramidal neurons significantly increased firing rate and enhanced their responses to locally applied NMDA. CPD30 also enhanced hippocampal firing activity after systemic administration. In 0.1- to 1.0-mg/kg doses, CPD30 increased spontaneous and NMDA-evoked firing activity of the neurons. Effects of CPD30 on NMDA responsiveness emerged faster (at 10 minutes post-injection) when a 1.0-mg/kg dose was applied compared with the onset of the effects of 0.1 mg/kg CPD30 (at 30 minutes post-injection).ConclusionsThe present results confirm that the inhibition of DAAO enzyme is an effective strategy for cognitive enhancement. Our findings further facilitate the understanding of the cellular mechanisms underlying the behavioral effects of DAAO inhibition in the mammalian brain.     

Altered Acquisition and Extinction of Amphetamine-Paired Context Conditioning in Genetic Mouse Models of Altered NMDA Receptor Function

Repeated intermittent exposure to amphetamine (AMPH) results in the development of persistent behavioral and neurological changes. When drug exposure is paired with a specific environment, contextual cues can control conditioned responses, context-specific sensitization, and alterations in dendritic morphology in the nucleus accumbens (NAc). Intact N-methyl-D-aspartate (NMDA) glutamate receptor signaling is thought to be required for associative learning. The acquisition of context-specific behavioral sensitization to AMPH and extinction of conditioned hyperactivity have been investigated in two genetically modified mouse strains: the serine racemase homozygous knockout (SR−/−) and glycine transporter 1 heterozygous mutant (GlyT1−/+). These strains have reciprocally altered NMDA receptor co-agonists, D-serine and glycine, levels that result in decreased (SR−/−) or increased (GlyT1−/+) NMDA receptor signaling. AMPH-induced changes in dendritic morphology in the NAc were also examined. SR−/− mice showed reduced expression of context-specific sensitization and conditioned hyperactivity. However, the conditioned hyperactivity in these mice is completely resistant to extinction. Extinction reversed AMPH-induced increased in NAc spine density in wild-type but not SR−/− mice. GlyT1 −/+ mice showed a more rapid acquisition of sensitization, but no alteration in the extinction of conditioned hyperactivity. The SR−/− data demonstrate that a genetic model of NMDA receptor hypofunction displays a reduced ability to extinguish conditioned responses to drug-associated stimuli. Findings also demonstrate that the morphological changes in the NAc encode conditioned responses that are sensitive to extinction and reduced NMDA receptor activity. NMDA receptor hypofunction may contribute to the comorbidity of substance abuse in schizophrenia.     

近年来戒毒药物的研究进展

目的对近年来戒毒药物的研究进展作一综述。方法查阅近年来国内外公开发表的有关研究论文 ,按戒毒药物的作用靶点分类汇总。结果与结论目前戒毒药物的作用靶点主要包括阿片受体、α2 受体、DA2 受体、M受体、NMDA受体以及钙离子通道等 ,与此相对应的戒毒药物均具有较好的疗效。寻找有效的成瘾药物治疗的作用靶点以及在分子水平上研制新的戒毒药物是目前此类工作的研究方向     

Dose escalating safety study of CNS 5161 HCl, a new neuronal glutamate receptor antagonist (NMDA) for the treatment of neuropathic pain

AIMS: The purpose of the current study was to establish the safety and maximal tolerated dose of CNS 5161 HCl. METHODS: Forty patients with chronic neuropathic pain (23 male, 17 female) were treated with escalating dosages of CNS 5161. All adverse events to study drug, blood pressure, heart rate, ECG, drug level and clinical laboratory values were monitored. Actual pain was measured on a 100-mm visual analogue scale (VAS) and ordinal verbal pain scores. RESULTS: The most commonly occurring nervous system disorder was headache, which was found more often during placebo than during CNS 5161 HCl treatment. Visual disturbances were experienced by 16.7% of patients receiving 250 microg and by 33.3% receiving 500 microg CNS 5161 HCl, but not during placebo treatment. An increase in blood pressure was observed in 8.3% of patients receiving 250 microg and in 50% of patients receiving 500 microg CNS 5161 HCl, compared with 15.4% during placebo treatment. The study was abandoned after two patients entered the 750 microg cohort due to a sustained systolic blood pressure response. Although this study was underpowered for the confirmation of efficacy, some indications of greater pain relief after 500 microg CNS 5161 compared with placebo could be observed (change in VAS between baseline and 12 h 10 +/- 22 mm vs. 2 +/- 19 mm; P = 0.11). CONCLUSIONS: CNS 5161 HCl was reasonably well tolerated up to 500 microg. The most common adverse events were hypertension, headache and mild visual disorders.     

脑脊液中D-Ser对齿状回颗粒细胞层突触传递和可塑性相关蛋白的作用研究

目的研究了脑脊液中D-丝氨酸(D-Ser)对齿状回颗粒细胞层突触传递和突触可塑性相关蛋白的作用。方法利用手术种植给药瘘管方法,在手术后第3天开始,隔天予以对照IgG和抗D-Ser IgG,给药3 d。之后采用胞外记录方法,刺激电极置于海马穿通纤维,记录齿状回颗粒细胞层LTP的诱导的变化,采用分子生物学方法检测皮层和海马突触可塑性相关蛋白表达的变化。结果抗D-Ser IgG连续中枢注射后,齿状回颗粒细胞层群峰电位的幅度明显下降,LTP的诱导受到明显阻断,同时海马和皮层部位突触可塑性相关蛋白的表达也发生了明显的变化。GAP-43在皮层和海马的表达分别升高约52%,58%。而MAP2的表达皮层降低约32%,海马降低约59%。皮层synapsinⅠ表达降低约45%,海马synapsinⅠ降低约57%。结论海马部位突触可塑性相关蛋白的表达对脑脊液中D-Ser浓度的变化表现出更高的敏感性,脑脊液中D-Ser具有重要的中枢调节作用,为神经精神相关疾病患者的临床治疗新药物和新方法的发现提供了理论支持。     

Effects of MK-801 on the expression of serine racemase and d-amino acid oxidase mRNAs and on the D-serine levels in rat brain

We have investigated the acute effects of the increasing doses of non-competitive N-methyl-d-aspartate receptor antagonist MK-801 (0.2-1.6 mg/kg) on the expression of serine racemase and d-amino acid oxidase (DAO) mRNAs in several brain areas of rats. We have also evaluated the effects of the chronic administration of MK-801 (0.4 mg/kg) on the gene expression of serine racemase and DAO and on the d-serine concentrations. A dose-dependent augmentation of the expression of serine racemase mRNA was seen in most brain areas at both 1 and 4 h after the administration. In contrast, a drastic decline in the expression of DAO mRNA was observed in most brain areas 1 h after the MK-801 administration, whereas a dose-dependent elevation in the expression of DAO mRNA was observed in most brain areas 4 h after the administration. The chronic MK-801 administration produced a significant increase in the expression of serine racemase mRNA in almost all brain areas, whereas no significant changes were found in the level of DAO mRNA in most brain areas. In addition, the chronic administration caused a slight but significant elevation in the concentrations of d-serine in the cortex and striatum. These present findings indicate that increasing the serine racemase mRNA and no changes in the DAO mRNA after the chronic administration could contribute to the elevation of the d-serine level in the forebrain, and that serine racemase and DAO could play an important role in the regulation of N-methyl-d-aspartate receptors via the d-serine metabolism.