Gephyrin regulates the cell surface dynamics of synaptic GABAA receptors
Gephyrin regulates the cell surface dynamics of synaptic GABAA receptors
复制标题
DOI：
10.1523/jneurosci.2267-05.2005
复制
发表时间：
2005-11-09
影响因子：
5.3
通讯作者：
Moss, SJ
中科院分区：
文献类型：
--
作者：
Jacob, TC;Bogdanov, YD;Moss, SJ
快速突触抑制的功效主要取决于抑制性突触处GABA（A）受体的积累，这一过程仍然知之甚少。在这里，我们研究了细胞表面GABA（A）受体的动态使用N-末端细胞外黄道pHluorin报告修饰的受体亚基。在海马神经元中，发现GABA（A）受体结合pHluorin标记的亚单位聚集在突触部位，也表达为弥漫性突触外染色。通过结合FRAP（荧光恢复后光漂白）测量与FM 4 -64标记的活性突触前末梢的实时成像，很明显，集群突触受体表现出显着较低的迁移率在细胞表面相比，他们的突触外的同行。为了研究这种限制的基础，我们使用RNAi来抑制桥蛋白的表达，桥蛋白是一种调节GABA（A）受体在突触部位积累的蛋白质。然而，桥蛋白是否控制受体簇的实际形成、其稳定性，或者仅仅是受体细胞表面数量的全局调节剂仍然未知。抑制桥蛋白的表达不会改变神经元细胞表面表达的GABA（A）受体的总数，但会显著减少受体簇的数量。实时成像显示，与对照神经元相比，在没有桥蛋白的情况下形成的簇明显更移动的。总之，我们的研究结果表明，突触GABA（A）受体具有较低水平的横向流动性相比，它们的突触外的同行，并建议桥蛋白在减少GABA（A）受体的扩散，促进其积累在抑制性突触的特定作用。
The efficacy of fast synaptic inhibition is critically dependent on the accumulation of GABA(A) receptors at inhibitory synapses, a process that remains poorly understood. Here, we examined the dynamics of cell surface GABA(A) receptors using receptor subunits modified with N-terminal extracellular ecliptic pHluorin reporters. In hippocampal neurons, GABA(A) receptors incorporating pHluorin-tagged subunits were found to be clustered at synaptic sites and also expressed as diffuse extrasynaptic staining. By combining FRAP ( fluorescence recovery after photobleaching) measurements with live imaging of FM4-64-labeled active presynaptic terminals, it was evident that clustered synaptic receptors exhibit significantly lower rates of mobility at the cell surface compared with their extrasynaptic counterparts. To examine the basis of this confinement, we used RNAi to inhibit the expression of gephyrin, a protein shown to regulate the accumulation of GABA(A) receptors at synaptic sites. However, whether gephyrin acts to control the actual formation of receptor clusters, their stability, or is simply a global regulator of receptor cell surface number remains unknown. Inhibiting gephyrin expression did not modify the total number of GABA(A) receptors expressed on the neuronal cell surface but significantly decreased the number of receptor clusters. Live imaging revealed that clusters that formed in the absence of gephyrin were significantly more mobile compared with those in control neurons. Together, our results demonstrate that synaptic GABA(A) receptors have lower levels of lateral mobility compared with their extrasynaptic counterparts, and suggest a specific role for gephyrin in reducing the diffusion of GABA(A) receptors, facilitating their accumulation at inhibitory synapses.