MOLECULAR CORRELATES OF ADULT SYNAPTOGENESIS

成人突触发生的分子相关性

• 批准号: 6392615
• 负责人: WILLIAM B LEVY
• 金额: $ 22.7万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-01 至 2003-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6392615
• 关键词: developmental neurobiology; estradiol; estrus; female; gender difference; gene expression; genetic mapping; hippocampus; hormone regulation /control mechanism; in situ hybridization; laboratory rat; neurogenetics; ovariectomy; polymerase chain reaction; protein localization; synaptogenesis; voltage /patch clamp

DESCRIPTION (Verbatim from the Applicant's Abstract): The hippocampus is an important cognitive brain region by virtue of its critical role in learning and memory. In the adult hippocampus of the female rat, the CA3-CA1 synapses undergo phasic cycles of syanptogenesis and synapse shedding with each 4 or 5 day estrous cycle. Although this cycle requires the ovarian steroid estrogen, the excitatory pyramidal neurons in CA1 and CA3 lack the conventional, genomic estrogen receptor (ER alpha). This remarkable observation leads to many, unanswered questions about the control of ovarian steroid-dependent cycle of adult synaptogenesis in the hippocampus. It also begs us to ask whether this same cycle also occurs in cerebral cortex. Using the molecular biological techniques of PCR, Northern and Southern blots, and differential display, we will uncover a set of molecules that are selectively synthesized in an estradiol-dependent manner in hippocampal CA1 and CA3. The time course of selective synthesis will enable us to define a temporal cascade of expressed sequences from early time points, those presumably controlling transcription factors, through later ones, possibly including sequences involved in synapse assembly. By pharmacologically blocking estradiol-dependent synapse formation with an NMDA receptor antagonist, and independently, by hormonally undoing synapses already formed, we will identify a particularly interesting subset of these expressed transcripts. This selective subset will include estradiol-dependent transcripts specific to the synaptogenetic cascade. Interpreting the roles played by the estradiol-dependent transcripts will be facilitated by electrophysiological correlations, anatomical localization, and by structural similarities to known molecules. Using naturally cycling females and ovariectomized animals receiving estradiol replacement, we will electrophysiologically assess NMDA receptor function in CA1 and correlate this with altered gene expression. In situ hybridization, supplemented as necessary with microdissection, will permit the cellular localization of these novel gene products to pyramidal and/or nonpyramidal neurons and/or glia. In situ hybridization will subsequently be used 1) to search for similar expression patterns in cerebral cortex and hypothalamus, and 2) to compare this form of adult synaptogenesis with that in the developmentally immature, male rat. Finally, we begin a series of experiments, including cloning and mapping the newly discovered genes to mouse and human chromosomes. These studies will eventually culminate in testing the function of these novel gene products in the genetically altered mouse.

描述（逐字摘自申请人的摘要）：海马体是 重要的认知大脑区域，因为它在学习和学习中发挥着关键作用 记忆。在雌性大鼠的成年海马中，CA3-CA1 突触 经历突触发生和突触脱落的阶段性循环，每 4 或 5 个 日发情周期。虽然这个周期需要卵巢类固醇雌激素， CA1 和 CA3 中的兴奋性锥体神经元缺乏传统的、基因组的 雌激素受体（ERα）。这一非凡的观察导致许多， 关于控制卵巢类固醇依赖性周期的未解答的问题 海马体中的成人突触发生。这也让我们不禁要问，这是否 同样的循环也发生在大脑皮层。 使用 PCR、Northern 和 Southern 印迹等分子生物学技术， 和差异显示，我们将发现一组分子 在海马 CA1 中以雌二醇依赖性方式选择性合成 CA3。选择性合成的时间过程将使我们能够定义一个时间 来自早期时间点的表达序列的级联，这些大概 通过后来的转录因子控制转录因子，可能包括 参与突触组装的序列。通过药理阻断 与 NMDA 受体拮抗剂形成雌二醇依赖性突触，以及 独立地，通过激素破坏已经形成的突触，我们将识别 这些表达的转录本的一个特别有趣的子集。这 选择性子集将包括特定于雌二醇依赖性转录本 突触发生级联。 解释雌二醇依赖性转录本所发挥的作用将是 通过电生理相关性、解剖定位和 通过与已知分子的结构相似性。使用自然循环的女性 和接受雌二醇替代的卵巢切除动物，我们将 电生理学评估 CA1 中的 NMDA 受体功能并将其关联起来 基因表达改变。原位杂交，必要时补充 通过显微切割，将允许这些新基因的细胞定位 产品作用于锥体和/或非锥体神经元和/或神经胶质细胞。原位 随后将使用杂交1)来搜索相似的表达 大脑皮层和下丘脑的模式，2）比较这种形式 成年突触发生与发育未成熟的雄性大鼠的突触发生。 最后，我们开始一系列实验，包括克隆和绘制 新发现的小鼠和人类染色体基因。这些研究将 最终测试这些新基因产物的功能 转基因小鼠。