CELLULAR CORRELATES OF AXONAL SPROUTING

哺乳动物下丘脑大细胞神经分泌神经元 系统（MNS）表现出异常强劲再生能力。它有 多年前就知道它们可以再生被切断的轴突。 垂体柄切片，这个实验室最近证明 未受伤的MNS神经元的轴突终末也将经历强烈的 成年大鼠的补偿性发芽。在摧毁了一半之后， 下丘脑-神经垂体束通过单侧下丘脑 损伤后，完整的对侧MNS神经元的轴突长出新芽， 使神经叶（NL）的轴突数量恢复到接近正常水平 在90天内。本项目的长期目标是确定 MNS的哪些特殊特性为这种情况提供了依据 可塑性。 这一建议建立在我们最近的发现基础上，即（a）加压素 (VP)和催产素（OT）神经元似乎是过度激活， 萌发反应和增加α 1和β II微管蛋白表达 同种型;和（B）在NL中发生瞬时神经胶质活化 在开始发芽之前， 血管生成伴随发芽。本提案的具体目标 （1）测试增加神经元活动的假设， 促进侧枝发芽，而活性降低则抑制侧枝发芽。 这将通过将神经元活动与由以下估计的神经元活动相关联来实现： 测量VP和OT基因表达、神经分泌和细胞凋亡。 代谢与幼年和成年大鼠轴突发芽程度的关系 在已知改变神经分泌活性的不同渗透条件下; (2)为了验证MNS神经元的侧支发芽 需要α 1、β II和β III微管蛋白mRNA的上调。这 将使用原位杂交（ISHH）完成， mRNA库随侧枝发芽程度的相对变化; 和（3）检验碱性成纤维细胞生长因子（bFGF）， 由MNS中的不同细胞类型产生并作用于MNS中的不同细胞类型， 刺激和协调轴突发芽和相关血管生成， 的NL。这将使用ISHH和ICC技术来检测 bFGF及其高亲和性受体FGF-R的表达 轴突发芽和相关血管生成过程中的MNS。履行 这些目标将为基本的细胞机制提供新的见解， 用中心肽能测定侧枝发芽的程度 神经元由于几乎所有的神经系统疾病都涉及某种程度的 组织变性，增加对细胞生物学的了解， 对中枢神经系统损伤的代偿性反应对于最终的 设计有效的治疗方法。

The neurons of the mammalian hypothalamic magnocellular neurosecretory system (MNS) exhibit unusually vigorous regenerative capabilities. It has been known for many years that they can regenerate axons severed by pituitary stalk section, and this laboratory has recently demonstrated that the axon terminals of uninjured MNS neurons will also undergo robust compensatory sprouting in the adult rat. Following destruction of one half of the hypothalamo-neurohypophysial tract via a unilateral hypothalamic lesion, axons of intact contralateral MNS neurons grow sprouts which return the axon population of the neural lobe (NL) to near normal levels within 90 days. The long-term objective of this project is to determine what special characteristics of the MNS provide the basis for this plasticity. This proposal builds upon our recent findings that (a) both vasopressin (VP) and oxytocin (OT) neurons appear to be hyperactivated during the sprouting response and increase expression of alpha1 and betaII tubulin isoforms; and (b) that transient glial activation occurs in the NL immediately prior to the initiation of sprouting, while continuous angiogenesis accompanies sprouting. The Specific Aims of this proposal are: (1) to test the hypothesis that increased neuronal activity facilitates collateral sprouting, whereas decreased activity inhibits it. This will be accomplished by correlating neuronal activity estimated by measures of VP and OT gene expression, neurosecretion, and cellular metabolism with the extent of axonal sprouting in juvenile and adult rats under different osmotic conditions known to alter neurosecretory activity; (2) to test the hypothesis that collateral sprouting by MNS neurons requires upregulation of alpha1, betaII, and betaIII tubulin mRNAs. This will be accomplished using in situ hybridization (ISHH) to correlate relative changes in mRNA pools with the extent of collateral sprouting; and (3) to test the hypothesis that basic Fibroblast Growth Factor (bFGF), produced by and acting upon different cell types in the MNS, serves to stimulate and coordinate axonal sprouting and associated angiogenesis in the NL. This will be accomplished using ISHH and ICC techniques to detect expression of bFGF and its high affinity receptor FGF-R by cells of the MNS during axonal sprouting and correlated angiogenesis. Fulfillment of these aims will provide new insights into basic cellular mechanisms which determine the extent of collateral sprouting by central peptidergic neurons. Since virtually all neural disorders involve some degree of tissue degeneration, increased knowledge of the cell biology underlying compensatory responses to injury of the CNS is crucial for the ultimate design of effective therapies.