Synaptic Regulation of Neurogenesis in the Dentate Gyrus

在成年期，齿状回的颗粒细胞（GC）层不断地添加新的神经元。 在2周内，超过50%的颗粒细胞死亡，大多数存活的GC并入齿状突 电路.尽管齿状核神经发生的功能意义尚不清楚， 神经发生损害了联想记忆，而联想学习任务增加了新GC的存活。 因此，GC存活的调节对于联想学习显得很重要。我们的初步研究显示 低频刺激穿通通路，主要的海马回投射到齿状回， 使8天龄的未成熟GC神经元完全存活。此外，随后的激活 相同的穿通路径纤维1周后诱导未成熟神经元中的立即早期基因（IEG）Zif 268。 我们的主要假设是传入活动促进GC存活，最有可能是通过GABA α受体激活。 这反过来又启动了未成熟GC向功能性成熟GC的快速发育， 受体和突触可塑性，反映在IEG Zif 268的活性依赖性表达。测试 这些假设，我们将使用成年大鼠与永久植入电极在穿通通路， 齿状回通过给神经元给药来评估穿通通路刺激对GC神经发生的影响。 溴脱氧尿苷（BrdU），一种仅在分裂细胞中掺入的胸苷类似物， 体视学和荧光免疫细胞化学来定量BrdU阳性细胞、IEG蛋白表达和 GC发育的标志物（祖细胞、未成熟和成熟）。在具体目标1的研究中，我们将 确定有丝分裂后的“关键期”，在此期间，穿孔通路刺激可以赋予新的- 生成GC神经元。我们还将探讨由传入活动提供的因素， 存活，并确定局部或全身施用GABA a、AMPA或NMDA受体 拮抗剂阻断由穿通通路刺激介导的GC存活。在具体目标2中，我们将确定 给予8天大GC的存活也加速了它们的功能发育。 确定NR 2B NMDAR亚基和Zif 268（一种涉及NR 2B NMDAR亚基的IEG蛋白）的最早时间点， 突触可塑性，可以表达后，传入诱导的GC生存时，与未受刺激 年龄相同的GC。我们还将探讨传入活动提供的可能因素，这些因素对 Zif 268表达，并确定GABA a、AMPA或NMDA受体拮抗剂是否阻断Zif 268在细胞中的表达。 幸存的GC在具体目标3中，我们将确定是否增加或减少8日龄的种群， 生存能力GC促进或损害空间或情节记忆任务的性能。这些研究 将有助于理解干细胞存活，这对治疗神经系统疾病至关重要， 促进PI的发展目标，获得主流资金，以保持一个可行的 研究/教学实验室环境在少数民族机构。

New neurons are continually added to the granule cell (GC) layer of the dentate gyrus during adulthood. Within 2 weeks, over 50% of these granule cells die, with most surviving GCs incorporating into the dentate circuit. Although the functional significance of dentate neurogenesis is unclear, treatments that reduce GC neurogenesis impair associative memory, and associative learning tasks increase the survival of new GCs. Thus the regulation of GC survival appears important for associative learning. Our preliminary studies show low-frequency stimulation of the perforant pathway, the principal glutamatergic projection to the dentate gyrus, confers complete survival of immature GC neurons 8 days of age. Furthermore, subsequent activation of the same perforant path fibers 1 week later induces the immediate-early gene (IEG) Zif268 in immature neurons. Our primary hypothesis is afferent activity promotes GC survival, most likely via GABAa receptor activation. This, in turn, initiates rapid development of immature GCs into functional, mature GCs that display NMDA receptors and synaptic plasticity as reflected in the activity-dependent expression of the IEG Zif268. To test these hypotheses, we will use adult rats with permanently implanted electrodes in the perforant pathway and dentate gyrus. The effects of perforant path stimulation on GC neurogenesis is assessed by administering bromodeoxyuridine (BrdU), a thymidine analog incorporated only in dividing cells, in conjunction with unbiased stereology and fluorescent immuno-cytochemistry to quantify BrdU-positive cells, IEG protein expression and markers of GC development (progenitor, immature and mature). In the studies of Specific Aim 1, we will determine the `critical period' after mitosis during which perforant path stimulation can confer survival of newly- generated GC neurons. We also will explore the factor(s) that are provided by afferent activity that confer GC survival, and determine whether local or systemic administration of GABAa, AMPA, or NMDA receptor antagonists block GC survival mediated by perforant path stimulation. In Specific Aim 2, we will determine if conferring survival of 8 day old GCs also accelerates their functional development This will be assessed by determining the earliest time point when NR2B NMDAR subunits and Zif268, an IEG protein implicated in synaptic plasticity, can be expressed following afferent-induced GC survival when compared with unstimulated GCs of the same age. We also will explore possible factors provided by afferent activity that are crucial for Zif268 expression, and determine if GABAa, AMPA, or NMDA receptor antagonists block Zif268 expression in surviving GCs. In Specific Aim 3, we will determine if increasing or depleting the population of 8-day old, survival-competent GCs facilitates or impairs performance in spatial or episodic memory tasks. These studies will contribute to understanding stem cell survival, crucial for treating neurological disorders, and will facilitate the PI's development goal of obtaining mainstream funding to maintain a viable research/teaching laboratory environment at a minority institution.