B型γ—氨基丁酸受体研究进展

ＧＡＢＡＢ受体是近年来新发现的ＧＡＢＡ受体亚型。它的活化在突触后膜增加Ｋ＾＋电导,引起长时程晚抑制性突触后电位;在突触前膜则抑制Ｃａ＾２＋电导,使兴奋性或抑制性递质的释放减少,该受体活动对机体的镇痛、肌肉痉挛、癫痫的发作等生理和病理生理过程都有重要的影响。     

B型γ－氨基丁酸受体研究进展

ＧＡＢＡＢ受体是近年来新发现的ＧＡＢＡ受体亚型。它的活化在突触后膜增加Ｋ＋电导,引起长时程晚抑制性突触后电位;在突触前膜则抑制Ｃａ２＋电导,使兴奋性或抑制性递质的释放减少。该受体活动对机体的镇痛、肌肉痉挛、癫痫的发作等生理和病理生理过程都有重要的影响。     

神经活性甾体与GABAA受体

中枢内存在的神经活性甾体能快速改变神经元兴奋性,它们不是与甾体受体发生作用,而是与脑内主要的抑制性神经递质γ－氨基丁酸（ＧＡＢＡ）受体相互作用,调制基介导的突触抑制功能,引发中枢抑制或兴奋性效应的变化,大多数甾体能增强ＧＡＢＡ受体活化ＣＩ＾－通道的通透性,引发中枢抑制或兴奋性效应的变化。该效应与特定的ＧＡＢＡＡ受体亚基α３和γ２有关,有些甾体具与ＧＡＢＡＡ受体拮抗剂类似的惊厥效应,总之,神经活性甾     

AMPA受体失敏和快速兴奋性突触传递

ＡＭＰＡ受体是离子型谷氨酸受体中重要的一类亚型,在中枢神经系统内主要介导快速的兴奋性突触传递。近年来,ＡＭＰＡ受体独特的失敏特性逐渐被阐明,已经确定了一些特异调节ＡＭＰＡ受体失敏的化合物。大量的生理学和药理学证据表明,ＡＭＰＡ受体失敏在快速兴奋性突触传递中起着重要的作用,对单个突触的传递效率、神经元的整合功能和突触的可塑性均有影响。     

鸡视网膜谷氨酸受体和GABA受体在两栖类卵母细胞中的表达

本工作利用两栖类卵母细胞作为功能表达系统,对鸡视网膜中的谷氨酸受体和ＧＡＢＡ受体的类型和基本性质进行了研究。在注射鸡视网膜ｍＲＮＡ的卵母细胞上,谷氨酸受体有明显的表达。Ｌ－Ｇｌｕ及其类似物ＫＡ,ＡＭＰＡ,ＱＡ都毫无例外地能诱导卵母细胞产生快速平滑的去极化电流,而ＮＭＤＡ,Ｌ－ＡＰ４,ＡＣＰＤ以及天冬氨酸不能诱导明显的电流反应。并且ＡＭＰＡ,ＱＡ对ＫＡ反应存在一定的抑制作用,提示ＡＭＰＡ,ＱＡ可能与ＫＡ作用于同一受体。抑制性氨基酸ＧＡＢＡ的受体被证明大部分为ＧＡＢＡＡ亚型,但有小部分的ＧＡＢＡ反应不能为荷包牡丹碱（ｂｉｃｕｃｕｌｌｉｎｅ）所阻断。     

新型γ－氨基丁酸受体：GABAc受体

众多的证据表明,在神经系统,特别是视觉神经通路中,除通常的ＧＡＢＡＡ和ＧＡＢＡＢ受体之外,还存在着一种具有不同药理特性的ＧＡＢＡＣ受体。这种受体不为荷包牡丹碱（ｂｉｃｕｃｕｌｌｉｎｅ）所阻遏,亦不为氯苯氨丁酸（ｂａｃｌｏｆｅｎ）所激活,在激活后并不显示失敏现象,可能在视网膜中视杆通路的信,自传递和调控中起重要作用。     

糖皮质激素对谷氨酸和GABA受体的电生理反应的快速调制作用

在大鼠下丘脑薄片和豚鼠腹腔神经节上,分别用玻璃微电极细胞外和细胞内记录方法,观察了１０＾６ｍｏｌ／Ｌ糖皮质激素（ＧＣ）对谷氨酸和ＧＡＢＡ受体分层效应的快速调制作用。结果表明,ＧＣ灌流后５ｍｉｎ,对谷氨酸受体介导的效应起抑制作用,而对ＧＡＢＡ受体介导的效应起增强作用。撤除ＧＣ后,神经元对谷氨酸和ＧＡＢＡ的反应恢复到对照水平。低钙高镁灌流液不能取消ＧＣ的调制作用。结果提示,ＧＣ在不需要突触环路条件下,     

糖皮质激素对谷氨酸和DGABA受体的电生理反应的快速调制作用

在大鼠下丘脑薄片和豚鼠腹腔神经节上,分别用玻璃微电极细胞外和细胞内记录方法,观察了１０－６ｍｏｌ／Ｌ糖皮质激素（ＧＣ）对谷氨酸和ＧＡＢＡ受体介导效应的快速调制作用。结果表明,ＧＣ灌流后５ｍｉｎ,对谷氨酸受体介导的效应起抑制作用,而对ＧＡＢＡ受体介导的效应起增强作用。撤除ＧＣ后,神经元对谷氨酸和ＧＡＢＡ的反应恢复到对照水平。低钙高镁灌流液不能取消ＧＣ的调制作用。结果提示,ＧＣ在不需要突触环路条件下,可能通过非基因组途径影响谷氨酸和ＧＡＢＡ受体介导的效应。     

鲫鱼视网膜谷氨酸受体和GABA受体在爪蟾卵母细胞中的表达

我们以两栖类卵母细胞为功能表达系统,通过注射鲫鱼（Ｃａｒａｓｓｉｕｓｃａｒａｓｓｉｕｓ）视网膜ｍＲＮＡ,利用电压箝及药物灌流手段,系统地研究了鲫鱼视网膜内氨基酸受体的类型和特征,结果如下：（１）Ｇｌｕ受体：ＫＡ可以诱发明显的去极化电流,而且Ｄｉａｚｏｘｉｄｅ能增强ＫＡ诱导的反应,这提示鲫鱼视网膜内某些Ｃｌｕ受体是ＡＭＰＡ选择性亚型（ＡＭＰＡ－ｐｒｅｆｅｒｒｉｎｇｓｕｂｔｙｐｅ）。（２）ＣＡＢＡ受体：ＧＡＢＡ能诱发一个快速、光滑的内向电流,绝大部分对ＧＡＢＡ的反应可被ｂｉｃｕｃｕｌｌｉｎｅ所压抑,而ＧＡＢＡ＿Ｂ受体的激动剂ｂａｃｌｏｆｅｎ则无任何作用,这提示,鲫鱼视网膜内大部分是ＧＡＢＡ＿Ａ受体。     

GABAB受体与腺苷酸环化酶偶联环节的脱敏研究

将突触体膜与佛波脂（ＰＭＡ）,ＧＡＢＡＢ受体激动剂巴氯芬（Ｂａｃｌｏｆｅｎ,ＢＡＬ）预孵育一定时间后ＢＡＬ对腺苷酸环化酶（ＡＣ）基础活性及ｆｏｒｓｋｏｌｉｎ刺激的ＡＣ活性的抑制率显著降低,而ｆｏｒｓｋｏｌｉｎ预孵育时,ＢＡＬ对基础及ｆｏｒｓｋｏｌｉｎ刺激的ＡＣ活性的抑制率不变,表明ＧＡＢＡＢ受体与ＡＣ偶联环节的脱敏机制涉及蛋白激酶激活。而与蛋白激酶Ａ无关,脱敏时ＧＡＢＡＢ受体的Ｋｄ值增加,本 实验     

GABA and neuroligin signaling: linking synaptic activity and adhesion in inhibitory synapse development

GABA-mediated synaptic inhibition is crucial in neural circuit operations. In mammalian brains, the development of inhibitory synapses and innervation patterns is often a prolonged postnatal process, regulated by neural activity. Emerging evidence indicates that gamma-aminobutyric acid (GABA) acts beyond inhibitory transmission and regulates inhibitory synapse development. Indeed, GABA(A) receptors not only function as chloride channels that regulate membrane voltage and conductance but also play structural roles in synapse maturation and stabilization. The link from GABA(A) receptors to postsynaptic and presynaptic adhesion is probably mediated, partly by neuroligin-reurexin interactions, which are potent in promoting GABAergic synapse formation. Therefore, similar to glutamate signaling at excitatory synapse, GABA signaling may coordinate maturation of presynaptic and postsynaptic sites at inhibitory synapses. Defining the many steps from GABA signaling to receptor trafficking/stability and neuroligin function will provide further mechanistic insights into activity-dependent development and possibly plasticity of inhibitory synapses.     

BDNF-mediated modulation of GABA and glycine release in dorsal horn lamina II from postnatal rats

Recent studies show that excitatory glutamatergic transmission is potentiated by BDNF in superficial dorsal horn, both at the pre- and the postsynaptic site. The role of BDNF in modulating GABA and glycine-mediated inhibitory transmission has not been fully investigated. To determine whether the neurotrophin is effective in regulating the spontaneous release of the two neurotransmitters, we have recorded miniature inhibitory postsynaptic currents (mIPSCs) in lamina II of post-natal rats. We show that application of BDNF enhanced the spontaneous release of GABA and glycine, in presence of tetrodotoxin. The effect was blocked by the trk-receptor inhibitor k-252a. Amplitude and kinetics of mIPSCs were not altered. Evoked GABA and glycine IPSCs (eIPSCs) were depressed by BDNF and the coefficient of variation of eIPSC amplitude was significantly increased. By recording glycine eIPSCs with the paired-pulse protocol, an increase of paired-pulse ratio during BDNF application was observed. We performed parallel ultrastructural studies to unveil the circuitry involved in the effects of BDNF. These studies show that synaptic interactions between full length functional trkB receptors and GABA-containing profiles only occur at non peptidergic synaptic glomeruli of types I and II. Expression of trkB in presynaptic vesicle-containing dendrites originating from GABAergic islet cells, indicates these profiles as key structures in the modulation of inhibitory neurotransmission by the neurotrophin. Our results thus describe a yet uncharacterized effect of BDNF in lamina II, giving further strength to the notion that the neurotrophin plays an important role in pain neurotransmission.     

Activation and deactivation rates of recombinant GABA(A) receptor channels are dependent on alpha-subunit isoform.

The role of subunit composition in determining intrinsic maximum activation and deactivation kinetics of GABA(A) receptor channels is unknown. We used rapid ligand application (100-micros solution exchange) to examine the effects of alpha-subunit composition on GABA-evoked activation and deactivation rates. HEK 293 cells were transfected with human cDNAs encoding alpha1beta1gamma2- or alpha2beta1gamma2-subunits. Channel kinetics were similar across different transfections of the same subunits and reproducible across several GABA applications in the same patch. Current rise to peak was at least twice as fast for alpha2beta1gamma2 receptors than for alpha1beta1gamma2 receptors (reflected in 10-90% rise times of 0.5 versus 1.0 ms, respectively), and deactivation was six to seven times slower (long time constants of 208 ms versus 31 ms) after saturating GABA applications. Thus alpha-subunit composition determined activation and deactivation kinetics of GABA(A) receptor channels and is therefore likely to influence the kinetics and efficacy of inhibitory postsynaptic currents.     

Declusterization of GABAA receptors affects the kinetic properties of GABAergic currents in cultured hippocampal neurons

Speed and reliability of synaptic transmission are essential for information coding in neuronal networks and require the presence of clustered neurotransmitter receptors at the plasma membrane in precise apposition to presynaptic terminals. Receptor clusterization is the result of highly regulated processes involving functional and structural proteins. Among the structural elements, microtubules are known to play a crucial role in anchoring of gamma-aminobutyric acid, type A (GABA(A)) receptors. Here we show that microtubule depolymerization with nocodazole induces the declusterization of GABA(A) receptors and modifies the kinetic properties of GABAergic currents in cultured hippocampal neurons. In particular, this drug, applied either in the bath or via the patch pipette, induced the acceleration of the onset kinetics of miniature inhibitory postsynaptic currents (mIPSCs) without significantly affecting their frequency, thus suggesting a main postsynaptic site of action. After nocodazole treatment, current responses to ultrafast applications of GABA exhibited a faster rise time and an accelerated onset of desensitization. A quantitative analysis of GABA-evoked currents and model simulations suggest that declusterization affects the gating properties of GABA(A) receptors. In particular, a faster entry into the desensitized state of declustered GABA(A) receptors may account for the changes in the kinetic properties of mIPSCs after nocodazole treatment. Hence it appears that the clustered condition of GABA(A) receptors contributes in shaping GABAergic currents.     

GABAA receptor kinetics in the cerebellar nuclei: evidence for detection of transmitter from distant release sites

Neurons of the cerebellar nuclei receive GABAergic input from Purkinje cells. Purkinje boutons have several closely spaced presynaptic densities without GABA transporters, raising the possibility that neurotransmitter released by one presynaptic site diffuses to multiple postsynaptic sites. To test whether such local spillover may contribute to transmission, we studied gating of GABA(A) receptors at 31-33 degrees C in cerebellar nuclear neurons acutely dissociated from mice. Currents were evoked by rapid application of long steps, brief pulses, and high-frequency trains of GABA to outside-out patches. Receptors desensitized and deactivated rapidly, and dose-response measurements estimated an EC(50) of approximately 30 microM. From these data, a kinetic scheme was developed that replicated the recorded currents. Next, we simulated diffusion of GABA in the synaptic cleft, constrained by previous electron microscopic data, and drove the kinetic GABA(A) receptor model with modeled concentration transients. Simulations predicted receptor occupancies of approximately 100% directly opposite the release site and approximately 50% at distant postsynaptic densities, such that receptors up to 700 nm from a release site opened on the timescale of the inhibitory postsynaptic currents before desensitizing. Further simulations of probabilistic release from multiple-site boutons suggested that local spillover-mediated transmission slows the onset and limits the extent of depression during high-frequency signaling.     

GABA generates excitement

In the CNS, gamma-aminobutyric acid (GABA) acts as an inhibitory transmitter via ligand-gated GABA(A) receptor channels and G protein-coupled GABA(B) receptors. Both of these receptor types mediate inhibitory postsynaptic transmission throughout the nervous system. For GABA(A) receptors, this inhibitory action is associated with a hyperpolarization due to an increase in conductance to chloride ions. Previous studies show that GABA(A) receptor activation in neonatal neurons and spinal cord neurons can be excitatory. Two papers recently appeared that clearly demonstrate that GABA can have a depolarizing and excitatory action in mature cortical neurons. Here we discuss the evolving story on chloride ion homeostasis in CNS neurons and its role in the bipolar life of the GABA(A) receptor.     

Gating of Ca2+-activated K+ channels controls fast inhibitory synaptic transmission at auditory outer hair cells

Fast inhibitory synaptic transmission in the central nervous system is mediated by ionotropic GABA or glycine receptors. Auditory outer hair cells present a unique inhibitory synapse that uses a Ca2+-permeable excitatory acetylcholine receptor to activate a hyperpolarizing potassium current mediated by small conductance calcium-activated potassium (SK) channels. It is shown here that unitary inhibitory postsynaptic currents at this synapse are mediated by SK2 channels and occur rapidly, with rise and decay time constants of approximately 6 ms and approximately 30 ms, respectively. This time course is determined by the Ca2+ gating of SK channels rather than by the changes in intracellular Ca2+. The results demonstrate fast coupling between an excitatory ionotropic neurotransmitter receptor and an inhibitory ion channel and imply rapid, localized changes in subsynaptic calcium levels.     

The enigma of transmitter-selective receptor accumulation at developing inhibitory synapses

The control of synaptic inhibition is crucial for normal brain function. More than 20 years ago, glycine and gamma-aminobutyric acid (GABA) were shown to be the two major inhibitory neurotransmitters. They can be released independently from different terminals or co-released from the same terminal to activate postsynaptic glycine and GABA(A) receptors. The anchoring protein gephyrin is involved in the postsynaptic accumulation of both glycine and GABA(A) receptors. In lower brain regions, both receptors can be concentrated in synapses, whereas in higher brain regions, glycine receptors are mostly excluded from postsynaptic sites. The activation of glycine and/or GABA(A) receptors determines the strength and precise timing of inhibition. Therefore, tight regulation of postsynaptic glycine versus GABA(A) receptor localization is crucial for optimizing synaptic inhibition in neurons. This review focuses on recent findings and discusses questions concerning the specificity of postsynaptic inhibitory neurotransmitter receptor accumulation during inhibitory synapse formation and development.     

GABAA receptor trafficking-mediated plasticity of inhibitory synapses

Proper developmental, neural cell-type-specific, and activity-dependent regulation of GABAergic transmission is essential for virtually all aspects of CNS function. The number of GABA(A) receptors in the postsynaptic membrane directly controls the efficacy of GABAergic synaptic transmission. Thus, regulated trafficking of GABA(A) receptors is essential for understanding brain function in both health and disease. Here we summarize recent progress in the understanding of mechanisms that allow dynamic adaptation of cell surface expression and postsynaptic accumulation and function of GABA(A) receptors. This includes activity-dependent and cell-type-specific changes in subunit gene expression, assembly of subunits into receptors, as well as exocytosis, endocytic recycling, diffusion dynamics, and degradation of GABA(A) receptors. In particular, we focus on the roles of receptor-interacting proteins, scaffold proteins, synaptic adhesion proteins, and enzymes that regulate the trafficking and function of receptors and associated proteins. In addition, we review neuropeptide signaling pathways that affect neural excitability through changes in GABA(A)R trafficking.     

The GABAB1b isoform mediates long-lasting inhibition of dendritic Ca2+ spikes in layer 5 somatosensory pyramidal neurons

The apical tuft of layer 5 pyramidal neurons is innervated by a large number of inhibitory inputs with unknown functions. Here, we studied the functional consequences and underlying molecular mechanisms of apical inhibition on dendritic spike activity. Extracellular stimulation of layer 1, during blockade of glutamatergic transmission, inhibited the dendritic Ca2+ spike for up to 400 ms. Activation of metabotropic GABAB receptors was responsible for a gradual and long-lasting inhibitory effect, whereas GABAA receptors mediated a short-lasting (approximately 150 ms) inhibition. Our results suggest that the mechanism underlying the GABAB inhibition of Ca2+ spikes involves direct blockade of dendritic Ca2+ channels. By using knockout mice for the two predominant GABAB1 isoforms, GABAB1a and GABAB1b, we showed that postsynaptic inhibition of Ca2+ spikes is mediated by GABAB1b, whereas presynaptic inhibition of GABA release is mediated by GABAB1a. We conclude that the molecular subtypes of GABAB receptors play strategically different physiological roles in neocortical neurons.