老年学习记忆减退大鼠脑突触体膜流动性改变

选用Morris水迷宫将老年大鼠分为学习记忆正常和学习记忆减退两部分,采用荧光偏振技术,对青年、老年记忆正常和老年记忆减退鼠脑分离实触体膜流动性进行测定,并检测神经节苷脂GM1对膜流动性的影响．结果表明老年记忆减退鼠新皮质、海马结构突触体膜荧光各向异性明显增加,即膜流动性显著降低,GM1对膜流动性有明显改善作用．相关分析表明新皮质、海马结构实触膜流动性与老年学习记忆减退密切相关,GM1的积极作用为临床治疗提供实验依据．     

老年学习记忆减退的神经行为学基础

用改良Ｍｏｒｒｉｓ水迷宫将老年大鼠分为老年学习记忆减退组和老年学习记忆正常组,并与青年组对照,检测其空间参考记忆和空间工作记忆,并对海马结构内胶质纤维酸性蛋白（ＧＦＡＰ）阳性胶质细胞和ＧＡＢＡ能中间神经元在光镜和电镜水平进行定量分析。结果显示老年学习记忆减退大鼠的空间参考记忆能力明显下降,并且其行为模式发生了改变,空间工作记忆改变不明显;齿状回分子层的病理改变非常明显,表现为ＧＡＢＡ能神经元丢失和     

钙神经素与学习和记忆

钙神经素 (calcineurin ,CN)是一种钙依赖的蛋白磷酸酶 ,其催化亚基的基因编码具严格保守性。近年来研究证明其在学习和记忆中有重要作用 ,参与了大脑神经元突触效应的去增强、多种不同机制的长时程抑制 (LTD)、长时程增强 (LTP)、认知记忆、短期记忆向长期记忆的转换、脑老化等过程。深入研究CN参与学习和记忆的机制及其与记忆减退性疾病的关系 ,具有重要理论与实践意义     

长时程增强效应初始阶段有突触前蛋白簇数目的快速增加

突触传递的长时程增强效应 (ＬＴＰ)和长时程抑制效应 (ＬＴＤ)反映神经元间突触传递效能的变化 ,目前认为这是学习和记忆的基础 ,而谷氨酸受体在ＬＴＰ和ＬＴＤ的诱导中起关键作用。海马是与学习和记忆功能密切相关的脑区 ,Ａｎｔｏｎｏｖａ等近来研究发现 ,体外培养的海马神经元在ＬＴＰ初始阶段有突触后谷氨酸受体 (ＧｌｕＲ1)簇数目的增加 ,而且在突触前神经元有突触前蛋白簇突触素数目以及突触素与谷氨酸受体共存位点数目的快速、持久的增加。进一步实验证明ＬＴＰ初始阶段并没有新蛋白的合成 ,突触前和突触后神经元蛋白质数目的快速增…     

大鼠纹状体边缘区与Meynert氏基底核的突触联系及其与学习记忆功能的关系——WGA-HRP束路追踪法、电镜法和行为实验法研究

纹状体边缘区（ＭｒＤ）是我们先后在大鼠、猫和猴脑内新发现的一个区域。它是位于纹状体尾内侧、环绕着苍白球头外侧的一层梭形细胞。ＭｒＤ的细胞构筑、免疫组化特性和纤维联系形式不同于纹状体其它区。ＭｒＤ的离心投射终止在苍白球尾侧,接近Ｍｅｙｎｅｒｔ基底核（ＮＢＭ）附近。损毁双侧ＭｒＤ后,动物的学习记忆功能减弱。ＮＢＭ已知与动物智能有关。本研究用纤维溃变和束路追踪电镜法结合行为实验方法,旨在了解ＭｒＤ与ＮＢＭ之间有无突触联系,以及此种联系与动物学习记忆的关系。用纤维溃变和束路追踪电镜法研究表明,由ＭｒＤ发出的纤维终末与ＮＢＭ的胆碱能神经元胞体间存在着突触联系。损伤ＭｒＤ造成ＭｒＤ和ＮＢＭ形成突触联系的终末溃变后,动物的学习记忆能力降低。研究结果表明边缘区与Ｍｅｙｎｅｒｔ基底核间存在着突触联系,而这种联系很可能是ＭｒＤ的学习记忆功能的结构基础之一。     

突触的可塑性与学习,记忆机制

位于哺乳动物海马、小脑皮层的不同类型的可塑性突触,分别具有突触传递的长时程强化(LTP)或抑制(LTD)现象,它们可能是某些经典条件反射形成的基础。以LTD型突触为记忆装置的小脑局部神经网络,具有典型的适应控制能力。突触可塑性的另一类表现是突触前纤维长芽,有证据表明,伴随大脑—红核系统条件反射的建立,在红核神经元胞体附近有新的突触形成,这可能是长期记忆的基础。     

衰老性记忆障碍与脑突触体内游离[Ca~(2 )]_i的相关性

采用行为观察和生化检测相结合的方法 ,在过去工作的基础上 ,研究了 12月龄和 18月龄小鼠学习记忆能力的变化和 18月龄小鼠四个脑区 (海马、大脑皮层、四叠体和小脑 )突触体内 [Ca2 ]i 的水平 ,同时还比较了老年记忆保持良好组与记忆障碍组小鼠的脑钙水平。结果表明 ,随着年龄的增长 ,小鼠的学习记忆能力显著下降 ,上述脑区 (除大脑皮层外 )突触内 [Ca2 ]i 均明显升高 ,其中老年记忆障碍小鼠脑钙水平升高最为显著。提示 ,小鼠衰老性记忆障碍可能与其脑突触体内 [Ca2 ]i 的超载有关。     

脑内胆碱能M受体和肾上腺素能α_2受体与老年记忆衰退的关系

很多资料表明中枢胆碱能系统的变化在老年记忆减退中起重要作用。有人发现海马与大脑皮层中胆碱能M受体的明显减少可能是正常老年记忆衰退的指标。但是越来越多的学者认为记忆功能的机理不只是胆碱能系统的作用,而是多种神经递质系统与神经活性物质共同协调和制约的结果,如有人观察到大脑皮层和海马肾上腺素能神经元的退变与记忆衰退有密切的关系。本文试图同时测定老年大鼠的大脑皮层、海马和下丘脑内M受体以及α_2肾上腺素能受体     

突触素、神经肽Y(NPY)在糖尿病模型大鼠脑组织细胞中的表达研究

对突触素(synaptophysin)、神经肽Y(NPY)在链尿佐菌素诱导的糖尿病模型大鼠额叶皮质和海马组织细胞中的表达进行研究,并利用UTHSCSA Image Tools 3.0进行图象分析,同时对其与学习记忆的关系进行探讨.选取成年Sprague-Dawley雄性大鼠20只,体重200-300g,随机分为两组.实验组用链尿佐菌素诱导产生糖尿病模型,并以血糖测定和尿糖水平测定进行筛选,另一组为空白对照组.继续饲养4周后,各组大鼠先进行Y型迷宫测试其学习记忆能力,然后取出脑组织,制做连续冰冻切片,对大脑额叶皮质和海马组织进行突触素、神经肽Y酶标免疫组织化学染色,观察这些蛋白在糖尿病大鼠和正常大鼠脑中表达的差异.结果发现糖尿病大鼠在Y型迷宫测试中,错误次数明显增多,糖尿病大鼠额叶皮质和海马神经元数目较正常对照组明显减少,神经细胞内突触素和神经肽Y的表达均较正常对照组明显下降.我们的研究显示突触素和神经肽Y在糖尿病大鼠大脑额叶皮质和海马组织内表达的减少可能与糖尿病组神经细胞突触数目及突触的可塑性下降、学习和记忆能力障碍有关.这可能是造成糖尿病性痴呆的一个因素.     

海马神经元新连接结构的初步原子力显微观察

目的:利用原子力显微技术(AFM)观察原代培养的海马神经元超微结构及其相互间的连接结构。方法:选择生长良好的海马神经元,用戊二醛固定30min后,固定于AFM基底上进行扫描和观测。结果:正常海马神经元表面光滑,起伏均匀、规律,突起及细胞之间的超微结构清晰,神经元胞体间存在膜性连接及长程非突触性突起连接。结论:神经元间存在直接膜性连接和长程非突触性突起连接结构。     

Higher environmental temperature-induced change in synaptosomal acetylcholinesterase activity of brain regions

Expcsure of adult male albino rats to higher environmental temperature (HET) at 35° for 2–12 hr or at 45° for 1–2 hr increases hypothalamic synaptosomal acetylcholinesterase (AChE) activity. Synaptosomal AChE activity in cerebral cortex of rats exposed to 35° for 12 hr and in cerebral cortex and pons-medulla of rats exposed to 45° for 1–2 hr are also activated. AChE activity of synaptosomes prepared from normal rat brain regions incubated in-vitro at 39° or 41° for 0.5 hr increases significantly in cerebral cortex and hypothalamus. The activation of AChE in ponsmedulla is also observed when this brain region is incubated at 41° for 0.5 hr. Increase of (a) the duration of incubation at 41° and (b) the incubation temperature to 43° under in-vitro condition decreases the synaptosomal AChE activity. Lioneweaver-Burk plots indicate that (a) in-vivo and invitro HET-induced increases of brain regional synaptosomal AChE activity are coupled with an increase ofV _max without any change inK _m (b) very high temperature (43° under in-vitro condition) causes a decrease inV _max with an increase inK _m of AChE activity irrespective of brain regions. Arrhenius plots show that there is a decrease in transition temperature in hypothalamus of rats exposed to either 35° or 45°; whereas such a decrease in transition temperature of the pons-medulla and cerebral cortex regions are observed only after exposure to 45°. These results suggests that heat exposure increases the lipid fluidity of synaptosomal membrane depending on the brain region which may expose the catalytic site of the enzyme (AChE) and hence activate the synaptosomal membrane bound AChE activity in brain regions. Further the in-vitro higher temperature (43°C)-induced inhibition of synaptosomal AChE activity irrespective of brain regions may be the cause iof partial proteolysis/disaggregation of AChE oligomers and/or solubilization of this membrane-bound enzyme.To whom to address reprint requests:     

Reduced mitochondrial buffering of voltage-gated calcium influx in aged rat basal forebrain neurons

Alterations of neuronal Ca(2+) homeostatic mechanisms could be responsible for many of the cognitive deficits associated with aging in mammals. Mitochondrial participation in Ca(2+) signaling is now recognized as a prominent feature in neuronal physiology. We combined voltage-clamp electrophysiology with Ca(2+)-sensitive ratiometric microfluorimetry and laser scanning confocal microscopy to investigate the participation in Ca(2+) buffering of in situ mitochondria in acutely dissociated basal forebrain neurons from young and aged F344 rats. By pharmacologically blocking mitochondrial Ca(2+) uptake, we determined that mitochondria were not involved in rapid buffering of small Ca(2+) influx through voltage-gated Ca(2+) channels (VGCCs) in the somatic compartment. For larger Ca(2+) influx, aged mitochondria showed a significant buffering deficit. Evidence obtained with the potentiometric indicator, JC-1, suggests a significantly reduced mitochondrial membrane potential in aged neurons. These results support the interpretation that there is a fundamental difference in the way young and aged neurons buffer Ca(2+), and a corresponding difference in the quality of the Ca(2+) signal experienced by young and aged neurons for different intensities of cytoplasmic Ca(2+) influx.     

Acetylcholinesterase activity and type C synapses in the hypoglossal, facial and spinal-cord motor nuclei of rats. An electron-microscope study

Using the electron-microscope technique of Lewis and Shute, we studied the localization of the acetylcholinesterase (AChE) activity in the hypoglossal, facial and spinal-cord motor nuclei of rats. The technique used selectively detects synapses with subsynaptic cisterns (type C synapses) as well as heavy deposits of reaction products in the rough endoplasmic reticulum, in fragments of the nuclear envelope, in some Golgi zones and on parts of the pericaryal plasma membrane, the axolemma and the dendritic membrane. In C synapses, AChE activity was located in the synaptic cleft and on the membrane of presynaptic boutons. Some C synapses exhibited distinct synaptic specialization in the form of multiple 'active zones'. These zones were characterized by dense presynaptic projections, short dilations of the synaptic cleft, and postsynaptic densities localized between the postsynaptic membrane and the outer membrane of the subsynaptic cistern. Within the postsynaptic densities, rows of rod- or channel-like structures were observed. The subsynaptic cisterns were continuous with the positive rough endoplasmic reticulum. The results are discussed in terms of the possible role of C synapses in the regulation of AChE synthesis in postsynaptic cholinergic neurons and/or in the regulation of AChE release into the extracellular space as well as in the establishment of new synaptic contacts.     

Acetylcholinesterase activity and type C synapses in the hypoglossal,facial and spinal-cord motor nuclei of rats

Summary Using the electron-microscope technique of Lewis and Shute, we studied the localization of the acetylcholinesterase (AChE) activity in the hypoglossal, facial and spinal-cord motor nuclei of rats. The technique used selectively detects synapses with subsynaptic cisterns (type C synapses) as well as heavy deposits of reaction products in the rough endoplasmic reticulum, in fragments of the nuclear envelope, in some Golgi zones and on parts of the pericaryal plasma membrane, the axolemma and the dendritic membrane. In C synapses, AChE activity was located in the synaptie cleft and on the membrane of presynaptic boutons. Some C synapses exhibited distinct synaptic specialization in the form of multiple active zones. These zones were characterized by dense presynaptic projections, short dilations of the synaptic cleft, and postsynaptic densities localized between the postsynaptic membrane and the outer membrane of the subsynaptic cistern. Within the postsynaptic densities, rows of rod- or channel-like structures were observed. The subsynaptic cisterns were continuous with the positive rough endoplasmic reticulum. The results are discussed in terms of the possible role of C synapses in the regulation of AChE synthesis in postsynaptic cholinergic neurons and/or in the regulation of AChE release into the extracellular space as well as in the establishment of new synaptic contacts.In honour of Prof. P. van Duijn     

Uptake and distribution in rat brain of organic and inorganic selenium

Polyunsaturated fatty acids (PUFAs) occur in relatively high amounts in phospholipids of the synapses. PUFAs may thus determine the fluidity of the synaptosomal membrane and, hereby, they may regulate the neuronal transmission. It was therefore tempting to suggest a system in the brain, that inhibits autooxidation of PUFAs. In order to trace such a protection system, Wistar rats were equally loaded with 4500 kBq of 75-Se either as selenite or as L-Se-methionine. By means of gradient ultracentrifugation, particulate fractions of the brains were isolated, and the radioactivity as well as the glutathione-transferase and -peroxidase activities were estimated. The distribution of the two selenium components among the particulate fractions was different. Thus, selenite gave higher radioactivity in myelin, then followed by the light synaptosomal and the vesicular fraction. L-Se-methionine was more equally incorporated in all particulate fractions, although highest activity was found in the mitochondrial fraction. Myelin and synaptic vesicles were devoid of transferase activity. On the other hand, the synaptosomal fraction showed highest specific transferase activity. The glutathione peroxidase activity was highest in the myelin fraction, followed by the vesicular and the synaptosomal fractions. The data obtained thus support the idea that the PUFAs of the synaptic compartment are protected against peroxidation, at least in part, by the selenium containing glutathione peroxidase.     

The Lipid Peroxidation Product, 4-Hydroxy-2-trans-Nonenal, Alters the Conformation of Cortical Synaptosomal Membrane Proteins

Abstract: Alzheimer's disease (AD) is widely held to be a disorder associated with oxidative stress due, in part, to the membrane action of amyloid β-peptide (Aβ). Aβ-associated free radicals cause lipid peroxidation, a major product of which is 4-hydroxy-2- trans -nonenal (HNE). We determined whether HNE would alter the conformation of synaptosomal membrane proteins, which might be related to the known neurotoxicity of Aβ and HNE. Electron paramagnetic resonance spectroscopy, using a protein-specific spin label, MAL-6(2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl), was used to probe conformational changes in gerbil cortical synaptosomal membrane proteins, and a lipid-specific stearic acid label, 5-nitroxide stearate, was used to probe for HNE-induced alterations in the fluidity of the bilayer domain of these membranes. Synaptosomal membranes, incubated with low concentrations of HNE, exhibited changes in protein conformation and bilayer order and motion (fluidity). The changes in protein conformation were found to be concentration- and time-dependent. Significant protein conformational changes were observed at physiologically relevant concentrations of 1–10 µ M HNE, reminiscent of similar changes in synaptosomal membrane proteins from senile plaque- and Aβ-rich AD hippocampal and inferior parietal brain regions. HNE-induced modifications in the physical state of gerbil synaptosomal membrane proteins were prevented completely by using excess glutathione ethyl ester, known to protect neurons from HNE-caused neurotoxicity. Membrane fluidity was found to increase at higher concentrations of HNE (50 µ M ). The results obtained are discussed with relevance to the hypothesis of Aβ-induced free radical-mediated lipid peroxidation, leading to subsequent HNE-induced alterations in the structure and function of key membrane proteins with consequent neurotoxicity in AD brain.     

Effect of preganglionic sympathectomy on the cholinesterase activity in the superior cervical ganglia of rats and hamsters

Summary By employing biochemical assay and histochemical enzyme techniques the effect of preganglionic sympathectomy on the cholinesterase (ChE) activity in the superior cervical ganglia of rats and hamsters was investigated. Biochemical assays indicate that the ChE activity in the superior cervical ganglia of adult rats and hamsters is 57.19 and 28.63 respectively (expressed in u moles acetylcholine hydrolyzed per min per g of tissue); two weeks after preganglionic denervation, about 50% and 60% of ChE activity are lost respectively. Histochemical enzyme examination reveals that in the rat superior cervical ganglion, the majority of the neurons are adrenergic with weak to moderate acetylcholinesterase (AChE) reaction and the minority of the neurons are cholinergic with strong AChE activity, while only one type of adrenergic neurons exhibits a weak AChE activity in the hamster superior cervical ganglion. The AChE activity is localized in the perinuclear area, in the cisternae of the rough surfaced endoplasmic reticulum, in the Golgi complex and on the plasma membrane of the hamster's neurons; it is mainly localized in the cisternae of the rough surfaced endoplasmic reticulum of the rat's neurons. AChE reaction product is also detected on the axolemmal membranes of the preganglionic nerve fibers in the sympathetic ganglia of rats and hamsters.After preganglionic sympathectomy, the AChE activity in the adrenergic neurons and in the preganglionic unmyelinated nerve fibers is markedly reduced, whereas the cholinergic neurons and preganglionic myelinated nerve fibers remain unchanged. On the basis of these results two conclusions have been reached: (1) The fact that strong AChE activity localized in the cholinergic neurons and preganglionic myelinated fibers is not influenced by denervation, suggests that these structures are able to produce AChE. (2) The reduction of AChE activity in the rat and hamster superior cervical ganglia two weeks after preganglionic denervation, observed by histochemical examination, can be correlated with a concomitant measurable reduction determined by biochemical assays.Supported in part by a grant from the National Science Council, Republic of China. The author wishes to express his gratitude to the Department of Pharmacology, College of Medicine, National Taiwan University, for the use of its equipment for biochemical assays     

Changes in the calcium transport systems of rat brain synaptosomes and their possible role in the pathophysiology of aging

Studies were undertaken on the age-associated peculiarities of the Ca2+ transport systems of the rat brain synaptosomes. It has been found that 45Ca2+ uptake reduced with ageing. The above reduction was not linked with the changes in the permeability of potential-dependent synaptosomal membrane Ca2+ depending upon the membrane potential. The distribution of calcium across the mitochondrial membrane changed with ageing, shifting towards higher extramitochondrial calcium levels in old rats, both in isolated and in synaptosomal mitochondria. While studying calcium efflux from mitochondria, it was found that, at equivalent calcium loads, the calcium efflux rates were slower in old rats as compared to adult animals. As observed, both resting [Ca2+]i and that obtained after K-depolarization drastically increased in old animals. The possible pathogenic mechanisms in neuronal injury, conditioned by this increase, are discussed.