在饮水条件反射中大鼠海马CA3区突触效应的变化

我们的工作表明,大鼠在明暗辨别学习过程中海马齿状回有习得性长时程增强(Long-term potentiation,LTP)现象,又CA_3区在大鼠学习和记忆过程有重要作用。本实验观察大白鼠海马CA_3区锥体细胞在条件性饮水反应的建立、巩固和消退过程中其突触效应的变化规律,以进一步探讨习得性LTP的特性,及从突触水平探讨海马CA_3区在学习记忆功能中的作用。     

大鼠海马CA3区的习得性长时程突触增强

本实验应用慢性埋植电极技术以电生理学结合行为学的方法,观察大鼠条件性饮水反应的建立、消退和再建立过程中,其海马CA_3区突触效应的变化规律。以刺激内嗅区的穿通纤维(PP)诱发的单突触的群体锋电位(PS)及群体兴奋性突触后电位(EPSPs)为指标,经叠加处理分析,发现随着条件反应的建立,海马CA_3锥体细胞出现突触效应的长时程增强(LTP),它随行为反应的实验性消退而消退,而在随后再次建立条件反应时,又重新出现;且无论此LTP达最高水平还是它的完全消退均超前于条件性行为反应的水平。又在一个实验日训练作业结束时PS并未立即随之增大,在24h内它随时间而发展,但到第4小时已达最高水平,且条件反应率是与PS的水平相应的,对PS与EPSPs的斜率进行相关分析表明,PS的变化主要是突触传递功效的变化。上述结果表明,海马CA_3区随着行为训练有习得性LTP产生。从其发神变化特点及其与条件性行为的关系,提示此习得性LTP极其可能是本实验中学习和记忆的展经基础。     

幼年和老年大鼠学习中海马CA_3区长时程突触增强的年龄特征

研究发现幼年和老年大鼠在条件性饮水反应的建立、消退和再建立过程中,海马CA_3区有习得性长时程突触增强(LTP)的形成、消退和再形成现象。在它的形成和再形成以及每实验日训练作业后习得性LTP的发展上,幼年鼠明显快于老年鼠,而习得性LTP的消退,在两组间无明显差异。这既表明海马CA_3区的习得性LTP具年龄特征,也为论证习得性LTP可能是学习和记忆的神经基础之一提供了新的证据。     

电休克对海马 CA_3区习得性长时程突触增强的影响

每天训练作业结束后对动物进行一次电休克处理,较多的动物(4/6)虽用了比对照组约多一倍的训练次数,但仍未能产生长时程突触增强(LTP),相应地条件反应也未能建立;部分动物(2/6)的突触效应不受影响,能产生LTP,并相应地条件反应亦能建立,且 LTP 发展超前于条件性行为的发展。在条件反应巩固后给大鼠一次电休克,可使它的海马 CA_3区的习得性 LIP—时性地下降,条件反应率也相应地下降。经1—4h,LTP 完全恢复,条件反应率也相应地恢复到电休克前的水平。表明习得性 LTP 受影响,可使条件性行为随之相应改变。它为论证习得性 LTP 可能是记忆的神经基础之一提供了进一步的证据。     

噪声对幼年和老年大鼠海马CA_3区习得性长时程突触增强的影响

经强噪声重复暴露(96 dB,2h/d,25 d)后,幼年和老年大鼠在条件反应的建立和再建立过程中,其海马CA_3区习得性长时程突触增强(LTP)的发展均受到明显阻抑,相应地动物达到学会标准需更多的训练。但幼年鼠受阻抑的程度更为严重。对习得性LTP的消退则在幼年和老年大鼠均无明显影响。本研究从突触功能的可塑性方面揭示幼年功物特别容易受到强噪声重复暴露的危害,提示应重视环境噪声对人类婴幼儿脑功能的损害作用的研究。     

不同浓度降钙素基因相关肽对小鼠海马区长时程增强的影响

本研究旨在探讨不同浓度的降钙素基因相关肽(calcitonin gene-related peptide,CGRP)对小鼠海马长时程增强的影响。将30日龄C57BL/6J小鼠随机分为空白组、不同浓度(50,100,200 nmol/L)CGRP组,CGRP+CGRP阻断剂组,每组10只。外源性给予海马区脑片不同浓度的CGRP,用离体细胞外场电位记录检测其对小鼠海马突触传递及长时程增强的影响。结果显示,给予不同浓度CGRP对小鼠海马突触前递质的释放没有明显的影响,但100和200 nmol/L CGRP可易化海马长时程增强的诱发,表现为长时程增强幅度的增加,并且这一作用可被CGRP特异性阻断剂CGRP8-37阻断。以上结果提示,CGRP可通过特异性受体浓度依赖性促进小鼠海马长时程增强的诱发。     

NMDA受体在海马CA_3区习得性LTP保持中的作用

在大鼠明暗分辨学习的建立和巩固过程中,通过与记录电极一起慢性埋植于海马CA_3区的注药管微量注射NMDA受体的特异性拮抗剂2-amino-5-phosphonovaleric acid(APV),观察对海马CA_3区突触效应及与之相关的习得性行为的影响。结果如下:(1)在动物经训练PS峰幅值刚增大至最高水平后,即在习得性LTP刚好形成后,每实验单元先于CA_3区注射AFV 1μl(2mmol/L),然后在药物有效作用时间内再进行训练,则PS峰值不能随训练而保持在最高水平,相反经8个实验单元,PS峰值降至实验前水平;相应地动物的正确反应率不能随训练而巩固,反而下降至10％以下。(2)在动物习得性LTP已形成并经一单元训练PS保持在最高水平后,于每实验单元训练前注射APV 1μl(2mmol/L),PS峰值同样不会随训练而保持在最高水平,经14个实验单元注药和训练,PS峰值逐渐降至实验前水平,相应地动物行为的正确反应率也降至10％以下,习得性行为消退,不过其消退速度比前一情况的动物为慢,说明习得性LTP发展情况不同,APV的作用效率有差别。结果表明:NMDA受体在习得性LTP的巩固中起着重要作用。     

尾核内注入GABA、Picrotoxin对家兔食物性运动条件反射的影响

于家兔尾核头部分别注入γ-氨基丁酸(GABA)3mg/5μl、GABA 转氨酶的抑制剂氨氧乙酸(AOAA)10μg/5μl 及 GABA 受体阻断剂印防已毒素(Picrotoxin)0.5μg/5μl 后,可暂时抑制食物性条件反射的出现,但一般运动、摄食等机能无明显障碍。作为对照,在尾核头部注入生理盐水或士的宁不影响条件反射的出现,注射 Picrotoxin 等于内囊区及海马也不影响条件反射活动。实验结果表明,尾核头部 GABA 能突触传递与实现条件反射活动有关。     

幼年和老年人鼠学习中海马CA[3]区长时程突触增强的年龄特征

研究发现幼年和老年大鼠在条件性饮水反应的建立、消退和再建立过程中,海马CA[3]区有习得性长时程突触增强（LTP）的形成、消退和再形成现象。在它们的形成和再形成以及每实验日训练作业后习得性LTP的发展上,幼年鼠明显快于老年鼠,而习得性LTP的消退,在两组间无明显差异。这既表明海马CA[3]区的习得性LTP具年龄特征,也为论证习得性LTP可能是学习和记忆的神经基础之一提供了新的证据。     

GABA_B自调性受体调节长时增强效应

对长时增强效应(long—term potentiation,LTP)的理解应从细胞分子水平去观察脊椎动物的学习与记忆。一些资料证明,在海马CA_1区诱导的LTP需要有N-methyl-D-aspartate(NMDA)受体系统短暂的激活。在低频传递时,γ-氨基丁酸(GABA)可通过突触抑制来阻断NMDA受体系统的明显活动。这种阻断作用是使神经元超极化时,Mg~(2+)阻断了由NMDA受体所调控的离子通道而引起的。在高频传递时,由于突     

Reelin and ApoE receptors cooperate to enhance hippocampal synaptic plasticity and learning

Two apolipoprotein E (apoE) receptors, the very low density lipoprotein (VLDL) receptor and apoE receptor 2 (apoER2), are also receptors for Reelin, a signaling protein that regulates neuronal migration during brain development. In the adult brain, Reelin is expressed by GABA-ergic interneurons, suggesting a potential function as a modulator of neurotransmission. ApoE receptors have been indirectly implicated in memory and neurodegenerative disorders because their ligand, apoE, is genetically associated with Alzheimer disease. We have used knockout mice to investigate the role of Reelin and its receptors in cognition and synaptic plasticity. Mice lacking either the VLDL receptor or the apoER2 show contextual fear conditioning deficits. VLDL receptor-deficient mice also have a moderate defect in long term potentiation (LTP), and apoER2 knockouts have a pronounced one. The perfusion of mouse hippocampal slices with Reelin has no effect on baseline synaptic transmission but significantly enhances LTP in area CA1. This Reelin-dependent augmentation of LTP is abolished in VLDL receptor and apoER2 knockout mice. Our results reveal a role for Reelin in controlling synaptic plasticity in the adult brain and suggest that both of its receptors are necessary for Reelin-dependent enhancement of synaptic transmission in the hippocampus. Thus, the impairment of apoE receptor-dependent neuromodulation may contribute to cognitive impairment and synaptic loss in Alzheimer disease.     

An unexpected effect of capsaicin on spontaneous GABA-ergic IPSCS in hippocampal cell cultures

Vanilloid receptors 1 (VRs1) expressed in a subpopulation of sensory neurons and responsible for processing of chemical and thermal noxious stimuli were also shown to be expressed in several cerebral structures and to be involved in the regulation of glutamatergic synaptic transmission. In this study, we started to investigate the possibility that VRs1 are also involved in the regulation of GABA-ergic synaptic transmission. For this purpose, the effect of a VR1 agonist, capsaicin, on spontaneous GABA-ergic inhibitory postsynaptic currents (IPSCs) was studied in hippocampal cell cultures using a patch-clamp technique. It was found that capsaicin (10 μM) decreased both the frequency and amplitude of spontaneous IPSCs. This finding suggests the involvement of VRs1 in the regulation of neuronal firing in some GABA-ergic interneurons and in the modulation of the efficacy of GABA-ergic synaptic transmission. However, considering the direction of the effect (a decrease in the IPSC frequency) and lack of its desensitization, the involvement of other receptor(s) also cannot currently be ruled out. Neirofiziologiya/Neurophysiology, Vol. 38, No. 4, pp. 364–367, July–August, 2006.     

Homeostatic plasticity of GABA-ergic synaptic transmission in rat hippocampal cell cultures

It has been shown recently that prolonged blockade of neuronal firing activates several homeostatic mechanisms in neocortical networks, including alteration of glutamatergic and GABA-ergic synaptic transmission, and postsynaptic changes are involved in both cases. We studied whether such treatment also affects GABA-ergic synaptic transmission in hippocampal cell cultures. Using whole-cell voltage-clamp recording and local extracellular stimulation, we investigated evoked inhibitory postsynaptic currents (IPSC) in cultured rat hippocampal neurons grown with the sodium channel blocker tetrodotoxin (TTX) and under control conditions. We found that chronic TTX treatment significantly decreased the amplitude of evoked IPSC. This decrease was accompanied by an increase in the coefficient of variation of the above parameter, which is suggestive of a presynaptic mechanism. In contrast, no changes in the IPSC reversal potential or paired-pulse depression were observed in TTX-treated cultures. We conclude that alteration of GABA-ergic synaptic transmission contributes to the homeostatic plasticity in hippocampal neuronal networks, and this change is at least in part due to a presynaptic mechanism.Neirofiziologiya/Neurophysiology, Vol. 36, Nos. 5/6, pp. 432–437, September–December, 2004.This revised version was published online in April 2005 with a corrected cover date and copyright year.     

Applicability of Peak-Scaled Nonstationary Fluctuation Analysis to the Study of Inhibitory Synaptic Transmission in Hippocampal Cultures

At present, there are no direct methods to determine the number of synaptic receptor-related channels activated in the course of synaptic transmission (N) or a value of the single-channel conductance (γ). Peak-scaled nonstationary fluctuation analysis (PS NSFA) should be considered the most well-developed indirect approach used for estimating these parameters. Despite the relatively wide using of this approach for the analysis of various synaptic currents, some aspects of possible errors that can occur in the course of data acquisition or their subsequent processing have not been studied. We examined in detail the problem of applicability of PS NSFA in the study of spontaneous and evoked GABA-ergic inhibitory postsynaptic currents (IPSCs). IPSCs were recorded using a dual patch-clamp technique from hippocampal neurons growing in low-density cultures. Parameters of the recorded IPSCs and values for different components of GABA-ergic synaptic transmission reported earlier were used for simulations and PS-NSFA analysis. In Monte Carlo computer simulations of evoked IPSCs, the influence of series resistance, background noise, asynchronicity of transmitter release, GABA_A channel properties, dendritic attenuation, and instrumental filtering on γ estimates obtained by PS NSFA was examined. We concluded that the γ and, consequently, N values may be satisfactorily estimated by the suggested approach using spontaneous and evoked IPSCs recorded in inhibitory synaptic connections in hippocampal cultures within a wide range of experimental conditions. We also estimated the mean of the single-channel conductance of synaptic GABA_A receptors in neurons from primary hippocampal cultures and found that this value (29 ± 5 pS) agrees well with the high conductance of single synaptic GABA_A receptors observed in acute hippocampal slices. This indicates that dissociated cultures are an adequate model for studying the properties of synaptic GABA_A receptors. Neirofiziologiya/Neurophysiology, Vol. 37, No. 4, pp. 379–388, July–August, 2004.     

Nociception-induced spatial and temporal plasticity of synaptic connection and function in the hippocampal formation of rats: a multi-electrode array recording

Background

Pain is known to be processed by a complex neural network (neuromatrix) in the brain. It is hypothesized that under pathological state, persistent or chronic pain can affect various higher brain functions through ascending pathways, leading to co-morbidities or mental disability of pain. However, so far the influences of pathological pain on the higher brain functions are less clear and this may hinder the advances in pain therapy. In the current study, we studied spatiotemporal plasticity of synaptic connection and function in the hippocampal formation (HF) in response to persistent nociception.

Results

On the hippocampal slices of rats which had suffered from persistent nociception for 2 h by receiving subcutaneous bee venom (BV) or formalin injection into one hand paw, multisite recordings were performed by an 8 × 8 multi-electrode array probe. The waveform of the field excitatory postsynaptic potential (fEPSP), induced by perforant path electrical stimulation and pharmacologically identified as being activity-dependent and mediated by ionotropic glutamate receptors, was consistently positive-going in the dentate gyrus (DG), while that in the CA1 was negative-going in shape in naïve and saline control groups. For the spatial characteristics of synaptic plasticity, BV- or formalin-induced persistent pain significantly increased the number of detectable fEPSP in both DG and CA1 area, implicating enlargement of the synaptic connection size by the injury or acute inflammation. Moreover, the input-output function of synaptic efficacy was shown to be distinctly enhanced by the injury with the stimulus-response curve being moved leftward compared to the control. For the temporal plasticity, long-term potentiation produced by theta burst stimulation (TBS) conditioning was also remarkably enhanced by pain. Moreover, it is strikingly noted that the shape of fEPSP waveform was drastically deformed or split by a TBS conditioning under the condition of persistent nociception, while that in naïve or saline control state was not affected. All these changes in synaptic connection and function, confirmed by the 2-dimentional current source density imaging, were found to be highly correlated with peripheral persistent nociception since pre-blockade of nociceptive impulses could eliminate all of them. Finally, the initial pharmacological investigation showed that AMPA/KA glutamate receptors might play more important roles in mediation of pain-associated spatiotemporal plasticity than NMDA receptors.

Conclusion

Peripheral persistent nociception produces great impact upon the higher brain structures that lead to not only temporal plasticity, but also spatial plasticity of synaptic connection and function in the HF. The spatial plasticity of synaptic activities is more complex than the temporal plasticity, comprising of enlargement of synaptic connection size at network level, deformed fEPSP at local circuit level and, increased synaptic efficacy at cellular level. In addition, the multi-synaptic model established in the present investigation may open a new avenue for future studies of pain-related brain dysfunctions at the higher level of the neuromatrix.     

学习过程中MF－CA3与PP－CA3突触传递效应的变化

应用慢性电极埋植技术以电生理学结合行为学的方法,探查在学习过程中大鼠海马ＣＡ３区两种不同输入突触（ＭＦ－ＣＡ３突触和ＰＰ－ＣＡ３突触）的可塑性变化及其相互关系。结果如下：（１）在分辨反应的建立过程中,在ＣＡ３区由ＭＦ诱发（ＭＦ－ＣＡ３）的群体锋电位（ｐｏｐｕｌａｔｉｏｎｓｐｉｋｅ,ＰＳ）和由ＰＰ诱发（ＰＰ－ＣＡ３）的群体锋电位,两者的峰值同步增大,同步达最高水平,且ＰＳ峰值达最高水平先于行为反应达学会标准;（２）在自然消退过程中,两者的ＰＳ峰值也是同步恢复至训练前水平的。结果表明,在ＣＡ３区这两种输入突触的习得性ＬＴＰ的产生和消退都是同步的,提示了它们之间可能具有协同作用。     

Reactions of hippocampal neurons at different stages of rat avoidance conditioning

The electrophysiological responses of neurons were compared in hippocampal slices from rats acquired and not acquired the passive avoidance reaction after the same conditioning procedure. Associative conditioning was accompanied by a gradual increase in the amplitude of the population spike evoked in CA1 area by stimulation of Schaffer collaterals. However, after reaching the learning criterion, the population spike significantly decreased. These phenomena were observed only at low (not maximal) intensities of test stimuli. After reaching the learning criterion, the paired-pulse facilitation was significantly higher in the slices prepared from the well-learned animals as compared with other groups (those having not reached the learning criterion, passive and active control). The obtained evidence validates the hypothesis that the observed intergroup differences stem from modifications of synaptic efficacy and suggests that after behavioral acquisition, plasticity induced by associative learning was substituted by other mechanisms probably related with declarative memory formation.