ABSTRACT
售检票系统
BACKGROUND:Membrane ion channels of different types often do not function independently but cross-talk to each ,activation of one channel would inhibit or potentiate the other.The cross-talk between ion channels plays an important role in neuronal excitability and processing of neural signals.The N-methyl-D-aspartate receptor(NMDA receptor)and theγ-aminobutyric acid type A receptor(GABA A receptor)are the major excitatory and inhibitory receptors in the central nervous system,respectively.However,the cross-talk between the two channels has not been thoroughly studied.In particular,the characteristics of the cross-talk between the two channels in the central auditory system are not known.The inferior colliculus(IC)is a major information integration center of the central auditory pathways in mammals,in which the NMDA and GABA A receptors are abundantly expressed.The purpose of the present study was to investigate the funcational interaction between NMDA and GABA A receptors in the IC.
METHODS:The cross-talk between NMDA and GABA A receptors in cultured IC neurons from new born Wistar rats was investigated with whole-cell patch-clamp techniques.The membrane current induced by aspartate(I Asp)and that by GABA (I GABA)were recorded through a voltage-clamp mode with the membrane potential of the recorded cell held at-60mV.Drugs used in the present study were a
pplied with a rapid application technique termed“Y-tube”method.
RESULTS:(1)The current induced by Asp at100 μM was suppressed by the pre-perfusion of GABA at100 μM,indicating cross-inhibition of NMDA receptors by activation of GABA A receptors.The cross-inhibtion could be blocked by application of bicuculline(10μM),a selective GABA A receptor antagonist;(2)the current induced by GABA at100 μM was not suppressed by the pre-perfusion of100μM Asp,but that induced by GABA at3 μM was,indicating concentration-dependent cross-inhibition of GABA A receptors by activation of NMDA receptors.The cross-inhibtion could be blocked by application of D-2-amino-5-phosphonovalerate (100μM),a selective NMDA receptor antagonist;(3)The inhibition of I GABA by Asp was not affected by blockage of voltage-dependent Ca2+channels(VDCCs)with CdCl2;however,the inhibition was blocked when the Ca2+in the external solution was substituted by Ba2+or when10mM BAPTA-a membrane-permeable Ca2+chelator空气源热泵技术
was added into the pipette solution,suggesting that the inhibition was mediatd by the Ca2+influx specifically through NMDA receptor channels,rather than VDCCs;(4) KN-62,a potent inhibitor of Ca2+/calmodulin-dependent protein kinaseⅡ(CaMKⅡ), reduced the inhibition of I GABA by Asp,indicating the involvement of CaMKⅡin the cross-inhibition.
CONCLUSIONS:There exists a functional interaction between NMDA and GABA A receptors in cultured neurons of the IC in rats.The cross-inhibition between the two types of channels is mediated by Ca2+entry through NMDA receptor channels,rather than VGCCs,and activation of CaMKⅡis involved in the downstream of the signaling pathway.The cross-talk between NMDA and GABA A receptors suggests complexity of information processing in the central auditory system.
莹石球Key words:Inferior colliculus;N-methyl-D-aspartate receptor;γ-Aminobutyric acid type A receptor;Whole-cell patch-clamp;Cross-talk
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stc2052第一章绪论
1.1.1引言
谷氨酸（Glu）和γ-氨基丁酸（GABA）分别是下丘中最重要的兴奋性和抑制性的神经递质，而N-甲基-D-门冬氨酸（NMDA）型谷氨酸受体和GABA A受体分别是其在下丘中对应的主要的受体类型(Adams et al.,1979;Ma et al.,2002a; Ma et al.,2002b)。NMDA受体和GABA A受体均属于配体门控离子通道型受体，但它们的结构和功能却是不同的。NMDA受体主要介导兴奋性突触传递中的慢成分(Ma et al.,2002b)。在发育早期，GABA A受体在突触传递中起到的是兴奋性的作用(Owens et al.,1996)，但是在成年动物的神经系统中，则起到抑制性的作用。在听觉系统中，这两种受体都参与了听觉信息加工的过程(Faingold et al.,1989; Vater et al.,1992)。本章节就将对NMDA受体和GABA A受体的结构、功能的研究和最新的研究进展做简要的综述。
1.1.2GABA A受体概述
GABA A受体是中枢神经系统（CNS）中重要的抑制性离子通道受体，它们广泛而非均匀的分布在哺乳动物脑内。GABA A受体是配体门控离子通道型受体，主要通透氯离子，介导氯离子电流，其主要的拮抗剂有竞争性的拮抗剂荷包牡丹碱（bicuculline），和非竞争性的的拮抗剂印防己毒素（PTX）。每个GABA A受体分子是由5个亚单位组成的，目前发现的亚单位一共有6类19种，这些亚单位构成了中枢神经系统中亚型众多的GABA A受体。GABA A受体介导的抑制性对于神经环路的稳定性起着非常重要的作用。 氧化锆全瓷
1.1.
2.1GABA A受体组成和分布
GABA A受体在脑内广泛存在，与其他配体门控离子通道一样，GABA A受体是一个五聚体结构（Fig.1.1）(Lester et al.,2004)，在哺乳动物的神经系统中已经成功克隆的有19个亚基，它们包括：α1～α6,β 1～β4, γ1～γ4,ρ1～ρ3,δ和ε 亚基(Sieghart et al.,1999;Wilke et al.,1997)。GABA A受体的各亚基族之间氨基酸序列的同源程度大约为30-40%，而相同亚基族的不同个体之间的同源程度大约为60-80%，虽然在理论上，这些亚基种类众多，且可以有多种不同的组合方式，
但是事实上并不是所有的亚基彼此之间都可以有效的组合成功能性的受体。在大多数的GABA A受体中，只含有一种α亚基，只有在少数的受体中才存在两种不同的α亚基。大多数受体中也只含有一种γ
亚基，只有少数是含两种γ亚基并存的(Fritschy et al.,1995;Fritschy et al.,1997)。一般最常见的受体亚型是由2个α亚基，2个β亚基以及一个γ亚基所组成的，即形成α1 β 2 γ2的组合(Farrar et al.,1999; Tretter et al.,1997)。在哺乳动物的脑内，有60%的GABA A受体是这种组成形式，数量占其次的分别是α2β 3 γ2和α3β n γ2，而其他的所占比例都不超过总量的5%。当体外表达这三种亚基时，其组成比例为α： β ： γ =2:2:1(McKernan et al., 1996)。GABA A受体的亚基组成，不仅反映了受体的特点，也反映了受体在细胞表面的分布和动态的调控(Farrant et al.,2005)。一些特殊的亚基组成，对应着受体功能的多样性，比如，只有当γ亚基与α，β亚基共同表达的时候，对GABA A受体的增强剂效应才能被观察到。除了通透氯离子，GABA A受体还通透HCO3-，因为后者的平衡电位要大于细胞的静息电位，且GABA A受体对其通透率要远远小于对氯离子的通透率，所以当激活GABA A受体的时候，就导致了氯离子的净内流(Bormann et al.,1987)。
Fig.1.1GABA A受体的结构示意图。A,B.一个GABA A受体由α1β2γ2亚基组成，图中黑箭头所指即为位于α1和β2亚基衔接处的GABA的结合位点；C.GABA A受体的胞外结构域和跨膜结构域示意图（ECD:extracellular domain,TM:transmembrane）(Kash et al.,2004)。
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