ROLE OF NEURAL PLASTICITY IN PUBERTY

该项目的长期目标是了解在我们物种中引发青春期开始的机制。在人类和其他高等灵长类动物中，下丘脑GnRH脉冲生成系统在婴儿期起作用，在成年时，该系统为垂体-性腺轴提供主要驱动。然而，在青春期前，支配灵长类性腺功能的神经内分泌轴的活动被下丘脑GnRH脉冲发生器的非性腺约束所阻止并长期处于静止状态。因此，灵长类动物的青春期是由GnRH脉冲发生器活动的重新唤醒触发的。这种青春期控制系统似乎是人类、猿和猴子独有的。虽然青春期代表着人类发展的一个重要里程碑，与行为、认知功能、世界观的戏剧性变化有关，在某些情况下，还与情感障碍的发生有关，但人们对这一发育阶段开始背后的基本神经生物学知之甚少。这一问题将在本提案中使用雄性恒河猴作为实验范例来解决，恒河猴是具有代表性的高等灵长类动物。将涉及以下具体目标：具体目标1--检验假设，即下丘脑内侧基底核(MBH)NPY张力的增加是青春期前发育期间对脉动性GnRH释放施加抑制的关键组成部分；具体目标2--确定在青少年向青春期状态转变期间丢失的GnRH包膜突触输入的表型；具体目标3--确定除细胞体和树突外，在正中隆起水平上GnRH网络结构的重构是否与青春期脉动性GnRH释放的重新增强有关；具体目标4-确定在发育的青春期前阶段结束时抑制GnRH神经元网络内的结构重塑是否可以防止青春期的开始。核糖核酸酶保护分析和原位杂交将被用来跟踪NPY基因表达的发育变化。蛋白质水平将通过Western blotting确定。脑室注射NPY受体拮抗剂或抗体可阻断NPY在MBH中的作用。NPY的合成将被反义寡核苷酸阻断。将使用光学和电子显微镜来研究下丘脑的结构重塑。包埋前和包埋后的标准免疫细胞化学程序将用于定量GnRH包涵体、树突和轴突终末的突触输入和神经胶质包膜的变化。将试图通过在青春期下丘脑的这个区域注射或过度表达内源性神经氨酸酶来阻止青春期下丘脑GnRH网络的可塑性。

The long term goal of this project is to understand the mechanisms that are responsible for triggering the onset of puberty in our own species. In man and other higher primates, the hypothalamic GnRH pulse generating system, which, in the adult, provides the principal drive to the pituitary-gonadal axis, is functional during infancy. During prepubertal development, however, activity of the neuroendocrine axis governing primate gonadal function is arrested and held in a protracted state of quiescence by non-gonadal restraint of the hypothalamic GnRH pulse generator. Puberty in primates is thus triggered by a reawakening of GnRH pulse generator activity. Such a pubertal control system appears unique to man, apes and monkeys. Although puberty represents a major landmark in human development that is associated with dramatic changes in behavior, cognitive function, outlook, and, in some cases, with the onset of affective disorders, little is known about the fundamental neurobiology underlying the onset of this developmental stage. This problem will be addressed in the present proposal using the agonadal male rhesus monkey, a representative higher primate, as an experimental paradigm. The following Specific Aims Will be addressed: Specific Aim 1 - To test the hypothesis that an increase in NPY tone in the mediobasal hypothalamus (MBH) is a critical component of the restraint that is imposed upon pulsatile GnRH release during prepubertal development; Specific Aim 2 - To determine the phenotype of the synaptic input to GnRH perikarya that is lost during the transition from the juvenile to the pubertal state; Specific Aim 3 - To determine whether structural remodeling of the GnRH network at the level of the median eminence, in addition to that at the cell body and dendrites, occurs in association with the onset of the pubertal reaugmentation of pulsatile GnRH release; Specific Aim 4 - To determine whether inhibition of structural remodeling within the GnRH neuronal network at the end of the prepubertal phase of development prevents the onset of puberty. RNAse protection assays and in situ hybridization will be used to track developmental changes in NPY gene expression. Protein levels will be determined by Western blotting. The action of NPY in the MBH will be blocked using NPY receptor antagonists or antibodies administered intracerebroventricularly. Synthesis of NPY will be blocked with antisense oligodeoxynucleotides. Structural remodeling in the hypothalamus will be studied using both the light and electron microscope. Standard pre-embedding and post-embedding immunocytochemical procedures will be applied to quantitate changes in synaptic input and glial ensheathment of GnRH perikarya, dendrites and axonal terminals. Attempts will be made to block plasticity in the GnRH network of the pubertal hypothalamus by injecting or overexpressing endoneuraminidase in this region of the brain.