Dynamics Of Excitatory Synaptic Transmission In The CNS

大脑以突触连接的模式将信息存储在大型神经元网络中。新的信息通过不完全理解的机制修改连接而被纳入神经网络。一个基本的问题是，各个连接是否独立地起作用，或者它们是否受到相邻突触活动的影响。突触连接是通过释放与受体神经元上的受体结合的可扩散神经递质分子来实现的;最近的证据表明，神经递质可能会逃离释放它的突触并扩散到相邻的突触中。突触连接之间的神经递质的这种“溢出”将对神经网络的信息容量以及它们在发育过程中构建的机制产生深远的影响。这个实验室的工作是确定兴奋性神经递质谷氨酸在海马体和视网膜中突触之间溢出的程度，海马体是大脑中学习和记忆储存的主要场所，而视网膜是视觉刺激被编码以便沿着视神经传输的地方。利用电生理技术在急性制备的大鼠视网膜和海马切片中，我们发现谷氨酸从释放它的突触逃逸并扩散到邻近的突触中。这种扩散受到谷氨酸转运蛋白的严格调节，谷氨酸转运蛋白主要位于神经胶质细胞膜上，结合谷氨酸并将其从脑脊液中清除。此外，似乎受体神经元的电状态影响受体是否对从远端突触释放的低水平谷氨酸有反应。目前正在继续研究这些机制的调制及其对神经元网络信息处理的影响。此外，我们已经记录了转运蛋白介导的海马星形胶质细胞的突触反应，以定量估计如何快速突触释放谷氨酸清除细胞外空间。在成年大鼠海马中，谷氨酸在释放后1毫秒内被吸收。这一速率如此之快，表明摄取实际上受到转运蛋白效率的限制，即，当它们结合谷氨酸而不是不结合谷氨酸时，它们将转运谷氨酸的可能性。转运蛋白将它们结合的谷氨酸释放回细胞外空间的大约50%，其速率接近我们测量的摄取速率。这表明转运蛋白缓冲了谷氨酸的扩散，并且谷氨酸实际上比从其测量的细胞外寿命预期的扩散得更少。工作继续确定什么样的突触外受体被激活，尽管这种快速吸收和可能的作用，神经元转运蛋白提供底物的合成抑制性神经递质，因此，限制癫痫。 我们在视网膜中的工作表明，某些类型的受体可能是专门定位的，以限制它们在某些条件下的激活。在神经节细胞上，NMDA型谷氨酸受体似乎位于突触周围，因此它们的激活被谷氨酸转运蛋白阻止，除非同时释放许多谷氨酸囊泡。实验室最近的工作表明，这些突触周围受体扩大了神经节细胞对光刺激的反应范围。此外，含有不同亚基组成的NMDA受体可能介导不同的功能输入到相同的神经节细胞。实验室中的大量工作现在都是针对理解这种独特的NMDA受体靶向的分子机制。 我们同时记录突触耦合视网膜神经元的其他实验表明，带状突触能够非常快速地释放递质，即使它们的生理释放是缓慢的。此外，我们的实验表明，带状突触协调多个囊泡在诱发反应的同时释放。此外，我们还发现，在这个突触的突触抑制几乎完全是由于神经递质囊泡的耗尽。虽然这被认为是高概率突触抑制的常见机制，但我们的实验为这一想法提供了相对定量的证据。这些结果可能为了解突触带的功能提供新的见解。 我们还研究了A17无长突细胞对视杆双极细胞的抑制性GABA能反馈。这种反馈似乎是由GABA-A和GABA-C受体介导的成分的复杂组合介导的。我们继续测试的想法，这两个组件的反馈可能会被激活在不同的释放条件。最近的研究表明，GABA从A17无长突细胞的释放是由钙进入钙渗透AMPA型谷氨酸受体引起的，而不是电压门控钙通道。我们已经发现，我们可以用有毒的5-羟色胺类似物特异性地消融A17无长突细胞，使我们能够比较来自A17的相互反馈和来自其他无长突细胞类型的反馈的特征。

The brain stores information in patterns of synaptic connections within large networks of neurons. New information is incorporated into a neural network through the modification of connections via mechanisms that are incompletely understood. One fundamental question is whether individual connections behave independently, or whether they are influenced by the activity of neighboring synapses. Synaptic connections are made through the release of diffusible neurotransmitter molecules that bind to receptors on the recipient neuron; recent evidence suggests that the neurotransmitter may escape the synapse in which it is released and diffuse into neighboring synapses. This "spillover" of neurotransmitter between synaptic connections would have a profound impact on the information capacity of neural networks and the mechanisms by which they are constructed during development. Work in this laboratory is directed towards determining the extent to which the excitatory neurotransmitter glutamate spills over between synapses in the hippocampus, a major site of learning and memory storage in the brain, and in the retina, where visual stimuli is encoded for transmission along the optic nerve. Using electrophysiological techniques in acutely prepared slices of rat retina and hippocampus, we have found that glutamate escapes the synapse from which it is released and diffuses into neighboring synapses. This diffusion is tightly regulated by glutamate transporters, pump proteins located primarily on glial membranes that bind glutamate and remove it from the cerebrospinal fluid. Moreover, it appears that the electrical state of the recipient neuron influence whether the receptors are responsive to low levels of glutamate released from a distant synapse. Work is continuing to investigate the modulation of these mechanisms and their impact on information processing in networks of neurons. In addition, we have recorded transporter-mediated synaptic responses in hippocampal astrocytes to estimate quantitatively how fast synaptically released glutamate is cleared from the extracellular space. In the adult rat hippocampus, glutamate is taken up within 1 millisecond following release. This rate is so fast that it suggests that uptake is actually limited by the efficiency of transporters, i.e., the probability that they will transport glutamate when they bind it rather than unbind it. Transporters release about 50% of the glutamate they bind back into the extracellular space, at a rate that approximates our measured rate of uptake. This suggests that transporters buffer the diffusion of glutamate and that glutamate actually diffuses less far than expected from the its measured extracellular lifetime. Work continues to determine what kind of extrasynaptic receptors are activated despite this rapid glutmaate uptake and a possible role for neuronal transporters in providing substrate for the synthesis of inhbitory neurotransmitter and, consequently, limiting epileptogenesis. Our work in the retina indicates that certain typed of receptors may be localized specifically to limit their activation under certain conditions. On ganglion cells, NMDA-type glutamate receptors appear to be located perisynaptically, such that their activation is prevented by glutamate transporters unless many vesicles of glutamate are released simultaneously. More recent work in the lab indicates that these perisynaptic receptors extend the range over which ganglion cells respond to light stimulation. In addition, NMDA receptors containing different subunit compositions may mediate defferent functional inputs to the same ganglion cell. A great deal of work in the lab is now directed toward understanding the molecular mechanisms underlying this unique targeting of NMDA receptors. Other experiments in which we record simultaneously from synaptically coupled retinal neurons indicate that ribbon synapses are capable of very fast transmitter release, even though their physiological release is slow. In addition, our experiments indicate that ribbon synapses coordinate the simultaneous release of multiple vesicles during evoked responses. In addition, we have discovered that synaptic depression at this synapse is due almost entirely to the depletion of neurotransmitter vesicles. While this is thought to be a common mechanism for depression at high-probability synapses, our experiments have provided relatively quantitative evidence for this idea. These results may provide new insights into the function of the synaptic ribbon. We also study inhbitory, GABAergic feedback from A17 amacrine cells onto rod bipolar cells. This feedback appears to be mediated by a complex combination of GABA-A and GABA-C receptor-mediated components. We continue to test the idea that these two components of feedback may be activated under different release conditions. Recent work indicates that GABA release from the A17 amacrine cell is elicited by calcium entering through calcium-permeagble AMPA-type glutamate receptors, not voltage-gated calcium channels. We have found that we can ablate A17 amacrine cells specifically with a toxic serotonin analog, allowing us to compare the characteristics of reciprocal feedback from A17s and feedback coming from other amacrine cell types.