Formation of mixed glycine and GABAergic synapses in cultured spinal cord neurons
Formation of mixed glycine and GABAergic synapses in cultured spinal cord neurons
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DOI：
10.1046/j.1460-9568.2000.00271.x
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发表时间：
2000-11-01
影响因子：
3.4
通讯作者：
Triller, A
中科院分区：
文献类型：
--
作者：
Dumoulin, A;Lévi, S;Triller, A
在脊髓中，GABA 和甘氨酸介导对含有甘氨酸和/或 GABA(A) 受体（分别为 GlyR 和 GABA(A)R）的单独或混合突触的抑制。我们在此分析了胚胎脊髓神经元体外 (DIV) 1 至 11 天突触发生过程中导致抑制性突触形成的事件顺序。我们使用免疫细胞化学方法同时检测抑制性末端特异性抗原、囊泡抑制性氨基酸转运蛋白 (VIAAT) 和以下突触后元件之一：GlyR、GABA(A)R 或 gephyrin（GlyR 的锚定蛋白，也与 GABA(A)R 相关）。定量分析显示，直到 5 DIV，大多数 gephyrin 簇都不与 VIAAT 阳性谱相邻，但在后期阶段与它们相关。相比之下，GlyR 和 GABA(A)R 在所有阶段主要聚集在包含 VIAAT 的终端前面。然而，大约 10% 的受体聚集体在非突触位点被检测到。 11 DIV 后，这两种受体共定位于 66.2 +/- 2.5% 的抑制性突触后结构域，而其中 30.3 +/- 2.6% 和 3.4 +/- 0.8% 分别仅包含 GlyR 和 GABA(A)R。有趣的是，在 3 DIV GABA(A)R 聚集在 gephyrin 和 GlyR 之前的突触后位置；因此，GABA(A)R 可能是混合甘氨酸和 GABA 能突触构建的起始元件。 gephyrin 与 GABA(A)R 的晚期共定位，以及其他研究小组证明，在没有 gephyrin 的情况下，未检测到突触后 GABA(A)R，表明 gephyrin 参与 GABA(A)R 的稳定，而不是参与其在突触位点的初始积累。
In the spinal cord, GABA and glycine mediate inhibition at separate or mixed synapses containing glycine and/or GABA(A) receptors (GlyR and GABA(A)R, respectively). We have analysed here the sequence of events leading to inhibitory synapse formation during synaptogenesis of embryonic spinal cord neurons between 1 and 11 days in vitro (DIV). We used immunocytochemical methods to detect simultaneously an antigen specific to inhibitory terminals, the vesicular inhibitory amino acid transporter (VIAAT), and one of the following postsynaptic elements: GlyR, GABA(A)R or gephyrin, the anchoring protein of GlyR, which is also associated with GABA(A)R. Quantitative analysis revealed that until 5 DIV most gephyrin clusters were not adjacent to VIAAT-positive profiles, but became associated with them at later stages. In contrast, GlyR and GABA(A)R clustered predominantly in front of VIAAT-containing terminals at all stages. However, about 10% of receptor aggregates were detected at nonsynaptic loci. The two receptors colocalized in 66.2 +/- 2.5% of the inhibitory postsynaptic domains after 11 DIV, while 30.3 +/- 2.6% and 3.4 +/- 0.8% of them contained only GlyR and GABA(A)R, respectively. Interestingly, at 3 DIV GABA(A)R clustered at a postsynaptic location prior to gephyrin and GlyR; GABA(A)R could thus be the initiating element in the construction of mixed glycine and GABAergic synapses. The late colocalization of gephyrin with GABA(A)R, and the demonstration by other groups that, in the absence of gephyrin, postsynaptic GABA(A)R is not detected, suggest that gephyrin is involved in the stabilization of GABA(A)R rather than in its initial accumulation at synaptic sites.