STEROID HORMONES AND NEURONAL REGENERATION

这项提议代表着我们正在进行的 神经营养剂治疗作用的研究， 丙酸睾丸酮(TP)，促进神经再生。模型 我们一直使用的系统是仓鼠面部运动神经元(HFMN)，一种 含有雄激素受体的外周神经元群。 上一笔赠款目标的实现确立了TP 显著加快面神经再生的速度 为了缩短发芽前的延迟，茶多酚可以触发神经细胞 身体对损伤的反应通过加速发病和增加 核仁反应的大小，存在内在的性别差异 面神经再生及雄激素对面神经的影响 再生是有性别差异的。这种神经营养剂是如何与众不同的 调节再生运动神经元的分子程序将是 此延续应用程序的焦点。具体目标是测试 以下是5个假设。首先，茶多酚加快了面神经的传导速度 细胞骨架基因的差异调控再生 轴突切断后的表达。茶多酚对大鼠血清白蛋白水平的影响 编码轴突再生所必需蛋白质的细胞骨架mRNAs将是 原位杂交法和Northern法检测轴突切断后的HFMN 用合适的细胞骨架基因探针进行印迹分析。第二，TP 通过加速面神经再生的速度 轴突运输的慢成分b的速率。立体定位法 将标记氨基酸注射到面部核团中将是 Done和TP对标记的转运和数量的影响 再生轴突节段内的细胞骨架蛋白在 确定了面神经损伤后的时间点。第三，茶多酚诱导 面神经元体对损伤的反应加速 通过核糖体RNA的快速转录激活而发生 吉恩。与2个特异性核糖体DNA探针的串联原位杂交 最初的转录本(前体)或稳定的(产物rRNA将在 损伤后选定的时间点以确定rRNA基因是否在 切断轴突的HFMN受TP的转录调控。第四，性别-- 茶多酚对金黄地鼠面神经再生的特异性影响 肾小球肾炎雄激素受体mRNA水平和/或调控的差异。在……里面 与雄激素受体mRNA探针的原位杂交将用于 确定雄激素受体mRNA水平的内在差异 男性和女性之间是否存在HFMN，以及是否存在性行为- 受体mRNA水平的特异性调节发生在轴突切断后， 带/不带TP。第五，TP可以以积极的方式改变 含雄激素受体中枢的再生反应模式 运动神经元。RDNA与β-微管蛋白基因的原位杂交 将使用探针来确定睾丸素是否可以触发 金黄地鼠红核脊髓神经元损伤后的再生反应模式 与在HFMN中观察到的类似。这些研究应该提供信息 与类固醇在神经元损伤中的作用机制相关，以及新的 有关人的内在分子程序差异的信息 受损的三叉神经节神经元与中枢神经系统神经元。

This proposal represents a direct continuation of our ongoing investigation of the therapeutic role of the neurotrophic agent, testosterone propionate (TP), in enhancing nerve regeneration. The model system we have been utilizing is the hamster facial motoneuron (HFMN), a population of peripheral neurons which contain androgen receptors. Accomplishment of the aims of the previous grant has established that TP significantly accelerates the rate of facial nerve regeneration without shortening the delay before sprouting occurs, TP triggers the nerve cell body response to injury by accelerating the onset and increasing the magnitude of the nucleolar reaction, intrinsic sex differences exist in facial nerve regeneration, and the effects of androgens on facial nerve regeneration are sex-specific. How this neurotrophic agent differentially regulates the molecular program of regenerating motoneurons will be the focus of this continuation application. The specific aims are to test the following 5 hypotheses. First, TP accelerates the rate of facial nerve regeneration through differential regulation of cytoskeletal gene expression following axotomy. The effects of TP on the levels of cytoskeletal mRNAs encoding proteins essential for axonal regrowth will be determined in axotomized HFMN using in situ hybridization and northern blot analyses with the appropriate cytoskeletal cDNA probes. Second, TP accelerates the rate of facial nerve regeneration through an acceleration of the rate of slow component b of axonal transport. Stereotaxic injections of labeled amino acids into the facial nuclear groups will be done and the effects of TP on the transport and quantity of labeled cytoskeletal proteins within segments of regenerating axons assessed at defined timepoints after facial nerve injury. Third, TP-induced acceleration of the onset of the facial nerve cell body response to injury occurs through the rapid transcriptional activation of the ribosomal RNA gene. Tandem in situ hybridization with 2 ribosomal DNA probes specific to the initial transcript (precursor) or stable (product rRNA will be done at selected timepoints after injury to determine if rRNA gene expression in axotomized HFMN is transcriptionally regulated by TP. Fourth, the gender- specific effects of TP on hamster facial nerve regeneration involve sex differences in androgen receptor mRNA levels and/or regulation in HFMN. In situ hybridization with androgen receptor mRNA probes will be done to determine if intrinsic differences in androgen receptor mRNA levels between male and female HFMN exist, as well as to establish whether sex- specific regulation of receptor mRNA levels occurs following axotomy, with/without TP. Fifth, TP can alter, in a positive manner, the regenerative response pattern in androgen receptor-containing central motoneurons. In situ hybridization with rDNA and betaII-tubulin cDNA probes will be used to determine if testosterone can trigger a regenerative response pattern in injured hamster rubrospinal neurons similar to that observed in HFMN. These studies should provide information relevant to the mechanism of steroid action in neuronal injury, and new information concerning differences in the intrinsic molecular programs of injured PNS vs. CNS neurons.