Analysis of Gene Expression Changes Associated With Long-Lasting Synaptic Enhancement in Hippocampal Slice Cultures After Repetitive Exposures to Glutamate
Analysis of Gene Expression Changes Associated With Long-Lasting Synaptic Enhancement in Hippocampal Slice Cultures After Repetitive Exposures to Glutamate
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DOI：
10.1002/jnr.22457
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发表时间：
2010-10-01
影响因子：
4.2
通讯作者：
Tashiro, Tomoko
中科院分区：
文献类型：
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作者：
Kawaai, Katsuhiro;Tominaga-Yoshino, Keiko;Tashiro, Tomoko
我们以前已经表明，反复暴露于谷氨酸（100 μ M，3分钟，三次，24小时间隔）诱导一个持久的突触增强伴随着突触在大鼠海马切片文化，一种现象称为上升（重复LTP诱导的突触增强）。为了研究RISE的分子机制，我们首先使用原始的寡核苷酸微阵列：“synaptoarray”分析了重复刺激后4小时至12天之间基因表达变化的时间过程。“结果表明，突触相关基因表达的变化在两个时间阶段被诱导，第三次刺激后24-96小时的早期阶段和6-12天的晚期阶段。在第三次刺激后48小时使用市售的高密度微阵列进行全面筛选，提供了负责RISE的候选基因。通过对这些基因和相关基因的实时PCR分析，鉴定了两类基因，1）先前报道的由生理和癫痫活性诱导的基因（bdnf、grm 5、rgs 2、syt 4、ania 4/carp/dc/k）和2）涉及基于cofilin的肌动蛋白丝动力学调节的基因（ywhaz、ssh 1 l、pak 4、limkl、cfl）。在第一类中，synaptotagmin 4也在蛋白质水平上显示出第三种刺激特异性上调。第二类中的五个基因通过第二次刺激协同上调，导致cofilin磷酸化的减少和肌动蛋白丝动力学的增强。相反，在第三次刺激后，它们被差异调节以增加cofilin磷酸化和增强肌动蛋白聚合，这可能是导致RISE建立的关键步骤。(C)2010 Wiley-Liss，Inc.
We have previously shown that repetitive exposures to glutamate (100 mu M, 3 min, three times at 24-hr intervals) induced a long-lasting synaptic enhancement accompanied by synaptogenesis in rat hippocampal slice cultures, a phenomenon termed RISE (for repetitive LTP-induced synaptic enhancement). To investigate the molecular mechanisms underlying RISE, we first analyzed the time course of gene expression changes between 4 hr and 12 days after repetitive stimulation using an original oligonucleotide microarray: "synaptoarray." The results demonstrated that changes in the expression of synapse-related genes were induced in two time phases, an early phase of 24-96 hr and a late phase of 6-12 days after the third stimulation. Comprehensive screening at 48 hr after the third stimulation using commercially available high-density microarrays provided candidate genes responsible for RISE. From real-time PCR analysis of these and related genes, two categories of genes were identified, 1) genes previously reported to be induced by physiological as well as epileptic activity (bdnf, grm5, rgs2, syt4, ania4/carp/dc/k) and 2) genes involved in cofilin-based regulation of actin filament dynamics (ywhaz, ssh1l, pak4, limkl, cfl). In the first category, synaptotagmin 4 showed a third stimulation-specific up-regulation also at the protein level. Five genes in the second category were coordinately up-regulated by the second stimulation, resulting in a decrease in cofilin phosphorylation and an enhancement of actin filament dynamics. In contrast, after the third stimulation, they were differentially regulated to increase cofilin phosphorylation and enhance actin polymerization, which may be a key step leading to the establishment of RISE. (C) 2010 Wiley-Liss, Inc.