单细胞基因表达分析技术在神经科学研究中的应用

单细胞的分子生物学是神经科学中较新的领域,研究对象包括单细胞DNA、RNA、蛋白质和线粒体DNA。单细胞基因表达分析技术具有传统技术难以相比的优势,正成为神经科学研究的重要工具。本文将介绍单细胞基因表达分析技术的操作流程、技术和方法的特点,概述其在神经科学研究中的应用,并展望其应用前景。     

神经递质受体和离子通道的移植

本文介绍了当前国际上在神经科学研究领域中的一个较新动向:分子生物学及分子遗传学的技术用于神经递质受体及离子通道的研究。着重介绍了南非爪蟾卵母细胞翻译系统在这一研究中的作用及特点,以及应用该方法在神经生物学的研究。这一方法的应用可能从基因水平认识各种神经递质受体、离子通道的结构与功能。     

中国科学院生物物理研究所视觉信息加工开放研究实验室研究课题申请指南

中国科学院视觉信息加工开放研究实验室专门从事视觉信息加工研究,这是当前神经科学的主要研究内容之一。 本开放研究实验室接受国内外神经科学家开展视觉信息加工研究的申请,一经批准即予以资助,并安排在本实验室工作。本实验室着重开展下列内容的研究工作:     

光片显微成像在斑马鱼模型中的生命科学研究的最新进展

光片显微成像是一种新兴的影像技术。相比于其他光学成像技术,光片显微成像技术具有成像深度大、对比度高、成像速度快、低光漂白和光毒性、高时空分辨率等特性。这些优点都是生命科学研究所急需的。近几年,研究者应用光片显微成像技术在斑马鱼模型基础上取得了很多进展。本综述主要介绍了光片显微成像技术在胚胎生物学和神经科学方面相关的研究成果。     

单细胞转录组测序技术新进展及其在造血系统研究中的应用

单细胞测序技术凭借其能全面反映细胞群体异质性这一优势,近年来发展迅速.其中,单细胞转录组测序技术提供了在分子水平上对细胞作分类或表征的替代方法,在发育生物学、神经科学、血液学、免疫及癌症等研究领域均展示出了广泛的应用前景.本文总结了近年来单细胞转录组测序技术的主要发展趋势,并列举了该技术在造血系统中的应用.     

分享科学，创造梦——美国第43届神经科学年会参会感

美国第43届神经科学年会于2013年11月9－13日在美国圣迭戈召开。本文根据作者的参会经历,对此次会议的概况、规模,以及内容作一简介,以期让读者了解神经科学这一研究领域的前沿以及美国神经科学年会的概况。     

《自然》:脑神经元连接同步定位首获成功

来自伦敦大学威康信托基金会神经科学学院的研究人员研发了一种新型技术,首次可以同时定位大脑中神经元之间的连接,以及功能,这对于未来研发计算机模拟大脑具有重要意义。这一研究成果公布在Nature杂志上。领导这一研究的是伦敦大学神经科学学院ThomasD.Mrsic-Flogel博士,其研究组主要兴趣在于大脑是如何编码信息,如何从一个或多个神     

中国科学院生物物理研究所视觉信息加工开放研究实验室研究课题申请指南

中国科学院生物物理研究所视觉信息加工开放研究实验室专门从事视觉信息加工研究,这是当前神经科学的主要研究内容之一。本开放实验室接受国内外神经科学家开展视觉信息加工研究的申请,一经批准即予以资助,并安排在本实验室工作。本实验室着重开展下列内容的研究工作:1.视觉通路和神经回路的研究,包括传递和加工视觉信息,如深度、形状、运动、颜色等的神经通路和回路。2.神经递质、调质、受体和离子通道在视觉各层     

《自然-神经科学》：研究揭开偏头痛患者怕光之谜

携带黑视蛋白的细胞能够解释为什么光线会给偏头痛患者带来痛苦。研究人员在本周的《自然-神经科学》杂志网络版上报告了这一研究成果。     

背景在动物图形识别中的作用

70年代以来,从神经科学中独立出一新型的分枝——神经行为学(Neu-roethology)。本文以背景对动物图形识别的影响为例,介绍近年来用神经行为学的观点和手段(如行为实验,神经生理学技术和神经元模型等)在这方面所取得的新进展。     

微透析校正的相关问题和方法

微透析技术是研究生物动态变化的一种新型的活体生物采样技术,近年来由于实验方法的不断改进,微透析技术已广泛应用于在体的定量研究。在进行生物细胞外液的定量研究中,微透析探针的校正是十分必要的。本从微透析的回收率、影响因素及校正方法等方面简要介绍了微透析校正的相关问题。     

Effects of tissue trauma on the characteristics of microdialysis zero-net-flux method sampling neurotransmitters

Microdialysis has been used for studying neurochemistry in brain regions that respond to afferent inputs or administered drugs. As the knowledge derived from and concerning microdialysis grows, so do the concerns over its invasiveness and, hence, the credibility of resulting data. Recent experimental and theoretical studies impugned the validity of the microdialysis zero-net-flux (ZNF) method in measuring brain extracellular neurotransmitters, suggesting that the tissue trauma resulting from probe implantation seriously compromises its worth. This paper developed a theoretical model to study the influences of two categories of tissue trauma on microdialysis ZNF operation: (1) morphological alterations in tissue extracellular structure and (2) physiological impairment of neurotransmitter release and uptake processes. Model results show that alterations of tissue extracellular structure negligibly affect the accuracy of the ZNF method in determining the basal level of extracellular neurotransmitter but do affect the fundamental characteristics of microdialysis: the extraction efficiency and relative recovery. An inhibited or damaged neurotransmitter uptake process always decreases the efficiency of microdialysis extraction, but rise of the relative recovery of neurotransmitters with the same uptake inhibition/damage occurs only when there is far more damage to the neurotransmitter release than to the uptake process in the tissue. A criterion for this rising trend of microdialysis relative recovery is discussed in terms of trauma parameters and neurotransmitter uptake inhibition.     

In vivo microdialysis--a new approach to the analysis of neurotransmitters in the brain

Microdialysis is a new technique to monitor levels of chemical compounds in the extracellular space over time. It involves the implantation of a microdialysis probe into the brain tissue. The probe is similar to a push-pull probe but the perfusate is contained inside a semi-permeable membrane located at the tip of the probe. Substances in the extracellular fluid will diffuse into the perfusate while substances included in the perfusate will diffuse into the tissue. This principle opens up a wealth of possibilities to monitor chemical events within the brain and to study the working mechanism of various drugs. The perfusate may be analysed by a number of different techniques. In this paper we give a short summary of various HPLC techniques that have proven particularly useful.     

The use of microdialysis for studying the regional effects of pharmacological manipulation on extracellular levels of amino acids--some methodological aspects.

The purpose of this study was to examine and validate the use of microdialysis for sampling and pharmacologically manipulating extracellular amino acids in the brain. Repeated use of microdialysis probes in acute intracerebral experiments did not significantly alter the relative recovery in vitro for the amino acids quantitated (GABA, aspartate, glutamate, glycine, taurine, and alanine). Regional differences in basal levels of some of the amino acids were detected in dialysates collected from the dorsomedial hypothalamus, striatum, and frontal cortex. The percent in vitro recoveries for the amino acids from the probes used in the three regions were not significantly different suggesting that the regional differences in basal levels of amino acids were functionally derived and not a consequence of variations in probe recovery. Perfusion with nipecotic acid, an inhibitor of GABA uptake, resulted in selective elevations in extracellular GABA in the three regions studied. Conversely, perfusion with high-potassium, a depolarizing agent, resulted in significant elevations in not only extracellular GABA but also aspartate, glutamate, and taurine. Thus, microdialysis is a method which can be employed to assess and to pharmacologically manipulate extracellular amino acids in the rat brain.     

Evidence on extracellular dopamine level in rat striatum: implications for the validity of quantitative microdialysis

Microdialysis zero-net-flux (ZNF) method is commonly used to monitor drug-induced changes in neurotransmitter baseline and release/uptake processes. Recent studies in this field suggest that microdialysis ZNF method seriously underestimates the resting concentration of extracellular dopamine in the rat neostriatum because probe implantation preferentially damages nearby dopamine release sites and that dopamine uptake inhibition increases the relative recovery of dopamine by microdialysis. This study assessed the validity of these claims by examining current data on extracellular dopamine levels at rest and after drug application obtained by voltammetry, a technique thought to induce less tissue disruption than microdialysis. To obtain the extracellular baseline value for dopamine from the evoked overflow data, we modified the existing dopamine kinetic model to suit both the resting and stimulated circumstances. It was found that dopamine uptake inhibition did in fact decrease the microdialysis relative recovery of dopamine, implying that the average basal extracellular dopamine level is within the range of 7-20 nm in rat striatum. This study concludes that the microdialysis ZNF method indeed underestimates the extracellular dopamine concentration, although not by as much as had been thought. Chronic microdialysis damages both neurotransmitter release and uptake, but it does so in a somewhat relative and proportional way for both processes. Thus the validity of the microdialysis ZNF method is not seriously undermined.     

Microdialysis of dopamine interpreted with quantitative model incorporating probe implantation trauma

Although microdialysis is widely used to sample endogenous and exogenous substances in vivo, interpretation of the results obtained by this technique remains controversial. The goal of the present study was to examine recent criticism of microdialysis in the specific case of dopamine (DA) measurements in the brain extracellular microenvironment. The apparent steady-state basal extracellular concentration and extraction fraction of DA were determined in anesthetized rat striatum by the concentration difference (no-net-flux) microdialysis technique. A rate constant for extracellular clearance of DA calculated from the extraction fraction was smaller than the previously determined estimate by fast-scan cyclic voltammetry for cellular uptake of DA. Because the relatively small size of the voltammetric microsensor produces little tissue damage, the discrepancy between the uptake rate constants may be a consequence of trauma from microdialysis probe implantation. The trauma layer has previously been identified by histology and proposed to distort measurements of extracellular DA levels by the no-net-flux method. To address this issue, an existing quantitative mathematical model for microdialysis was modified to incorporate a traumatized tissue layer interposed between the probe and surrounding normal tissue. The tissue layers are hypothesized to differ in their rates of neurotransmitter release and uptake. A post-implantation traumatized layer with reduced uptake and no release can reconcile the discrepancy between DA uptake measured by microdialysis and voltammetry. The model predicts that this trauma layer would cause the DA extraction fraction obtained from microdialysis in vivo calibration techniques, such as no-net-flux, to differ from the DA relative recovery and lead to an underestimation of the DA extracellular concentration in the surrounding normal tissue.     

Intracerebral Microdialysis: Electrophysiological Evidence of a Critical Pitfall

Abstract: Using microdialysis probes incorporating an electrode for the recording of extracellular field potentials, we have found that microdialysis markedly inhibited the propagation of spreading depression. This effect was independent of the microdialysis flow rate and did not result from tissue injury following probe implantation. Increasing the K⁺ concentration in the perfused artificial CSF dose-dependently restored the propagation of spreading depression and revealed a large, synchronous transient increase in extracellular glutamate. These findings clearly illustrate that microdialysis can influence the experimental or pathological conditions under study, by buffering transient changes in the extracellular fluid composition. Epileptic seizures and ischaemia are two important conditions that may be prone to such a detrimental interaction.     

Coupled Effects of Mass Transfer and Uptake Kinetics on In Vivo Microdialysis of Dopamine

Abstract: Voltammetric microelectrodes and microdialysis probes were used simultaneously to monitor extracellular dopamine in rat striatum during electrical stimulation of the medial forebrain bundle. Microelectrodes were placed far away (1 mm) from, immediately adjacent to, and at the outlet of microdialysis probes. In drug-naive rats, electrical stimulation (45 Hz, 25 s) evoked a robust response at microelectrodes far away from the probes, but there was no response at microelectrodes adjacent to and at the outlet of the probes. After nomifensine administration (20 mg/kg i.p.), stimulation evoked robust responses at all three microelectrode placements. These results demonstrate first that evoked release in tissue adjacent to microdialysis probes is suppressed in comparison with evoked release in tissue far away from the probes and second that equilibration of the dopamine concentration in the extracellular fluid adjacent to and far away from the probes is prevented by the high-affinity dopamine transporter. Hence, models of microdialysis, which assume the properties of tissue to be spatially uniform, require modification to account for the distance that separates viable sites of evoked dopamine release from the probe. We introduce new mass transfer resistance parameters that qualitatively explain the observed effects of uptake inhibition on stimulation responses recorded with microdialysis and voltammetry.     

Microdialysis measures of functional increases in ACh release in the hippocampus with and without inclusion of acetylcholinesterase inhibitors in the perfusate

Because brain extracellular acetylcholine (ACh) levels are near detection limits in microdialysis samples, an acetylcholinesterase (AChE) inhibitor such as neostigmine is often added to microdialysis perfusates to increase ACh levels in the dialysate, a practice that raises concerns that the inhibitor might alter the results. Two experiments compared functional differences in ACh release with and without neostigmine. In the first experiment, 30-60% increases in extracellular ACh concentrations in the hippocampus were evident during food-rewarded T-maze training with 20-500 nm neostigmine in the perfusate but no increases were seen without neostigmine. In the second experiment, 78% increases in ACh release in the hippocampus were seen after injections of the GABA(A) receptor antagonist, bicuculline, into medial septum only if neostigmine (50 nm) was included in the perfusate. These findings suggest that, in the hippocampus, endogenous brain AChEs are very efficient at removing extracellular ACh, obscuring differences in ACh release in these experiments. Therefore, inclusion of AChE inhibitors in the microdialysis perfusate may be necessary under some conditions for observations of functional changes in release of ACh in the hippocampus.     

Citrulline extracellular level in the nucleus accumbens during feeding behaviour

In Sprague-Dawley rats, by means of in vivo microdialysis combined with HPLC analysis it was shown that a consumption of a novel food did not produce any changes in extracellular levels ofcitrulline (an NO-co-product) in the medial n. accumbens. In contrast, the rejection of the novel food caused a rise of the extracellular citrulline level in this brain area which can be completely prevented by intra-accumbal infusion of 0.5 mM 7-nitroindazple, a neuronal NO-synthase inhibitor. The data obtained reveal for the first time that new food rejection (but not its consumption) is characterized by neuronal NO-synthase activation and, very likely, NO production in the medial nucleus accumbens.