NEURAL REGULATION OF THE FETAL PARAVENTRICULAR NUCLEUS

我们已经在绵羊身上证明了胎儿下丘脑室旁核 核团(PVN)是：1)ACTH的分泌是必需的。 下丘脑-垂体-肾上腺轴(HHAA)对低氧血症的反应和 2)导致分娩的HHAA活性持续增加。在……里面 成年动物从脑干和海马区传入的神经元具有强大的 修饰对HHAA活性的影响。此外，脑干和 海马体是大脑中高浓度的 糖皮质激素反馈受体(GR)。GR的实验操作 对HHAA的产量有深远的影响。我们建议继续我们的 胎羊胎儿下丘脑室旁核在低氧血症中作用的研究 在分娩的时候。 具体目标1将确定6-羟基多巴胺(6-OHDA)的作用 下丘脑室旁核激素反应中的敏感神经元传入 胎儿HHAA对低氧血症和分娩时机的影响。这将是 通过使用：a)立体定位和尼氏染色技术来完成 制作120天胎羊脑的立体定位图谱； B)荧光金和免疫细胞化学染色的逆行追踪 多巴胺β-羟基酶和c-fos即刻蛋白的表达 早期基因确定脑干儿茶酚胺神经元投射到 PVN和这些神经元在低氧血症时被激活 胎儿分娩；c)立体定向分娩技术 微量注射6-OHDA，一种广泛用于研究的神经毒素 儿茶酚胺能神经元，进入下丘脑室旁核本身即可清除易感 神经元输入。 特定目标2将确定来自 在大脑的荷尔蒙反应中通过穹隆移动的海马体 HHAA对低氧血症和分娩时机有影响。这将是 使用我们的立体定位手术技术完成横断 穹隆。 具体目标3将确定糖皮质激素反馈在 脑干和海马体对胎儿HIS激素反应的影响 立体定向植入皮质醇晶体引起的低氧血症 脑干和海马体的预选区域。特定站点 选择将基于ICC和mRNA测量的GR浓度 分析。在所有三个特定目标中，外周血浆 放射免疫法测定CRH和PVN中ACTH、AVP、皮质醇和PVN含量 AVP信使核糖核酸将作为终点进行测量。 低氧血症和早产是发病率和死亡率的重要原因。 在人类怀孕期间。此外，低氧血症在本提案中被用作 可重复的、精确确定的急性应激刺激使我们能够 调整我们的方法，并与长期的研究进行比较 分娩开始的原因更复杂，更不为人所知。 这些研究合乎逻辑地源于前一年的项目I和II 在资金阶段，我们展示了，除其他结果外， PVN的损伤消除了胎儿对低氧血症的ACTH反应，并 低血压和妊娠延长。详细对照实验 胎儿对低氧血症的反应和体内启动机制的数据 只有在胚胎动物身上才能获得分娩的机会。这个项目 与项目I和项目III在跨物种和 机械化的水平。我们的实验与 胎儿损伤和浪费的问题。此外，这些研究也将得到改进 我们对神经和内分泌相互作用在控制高血压中的理解 HHAA功能。

We have shown in sheep that the fetal hypothalamic paraventricular nucleus (PVN) is necessary for: 1) secretion of ACTH by the hypothalamo-hypophyseal-adrenal axis (HHAA) in response to hypoxemia and 2) the prolonged increase in HHAA activity that leads to parturition. In adult animals neuronal inputs from brainstem and hippocampus have strong modifying effects on HHAA activity. In addition, the brainstem and hippocampus are areas of the brain with high concentrations of glucocorticoid feedback receptors (GR). Experimental manipulation of GR has profound influence on HHAA output. We propose to continue our investigation in fetal sheep of the role of the fetal PVN in hypoxemia and in parturition. Specific Aim 1 will determine the role of 6-hydroxydopamine (6-OHDA) susceptible neuronal inputs to the PVN in the hormonal response of the fetal HHAA to hypoxemia and in timing of parturition. This will be accomplished by using: a) stereotaxic and nissl staining techniques to produce a stereotaxic atlas of the 120 days gestation fetal sheep brain; b) retrograde tracing of fluorogold and immunocytochemical (ICC) staining for dopamine beta-hydroxylase and protein expression of c-fos immediate early gene to determine which brainstem catecholamine neurons project to the PVN and that these neurons are activated during hypoxemia and parturition in the fetus; c) stereotaxic technique to deliver micro-injections of 6-OHDA, a neurotoxin used extensively to study catecholaminergic neurons, into the PVN itself to remove susceptible neuronal input. Specific Aim 2 will determine the role of neuronal inputs from the hippocampus that travel via the fornix in the hormonal response of the HHAA to hypoxemia and in timing of parturition. This will be accomplished using our stereotaxic surgical technique to transect the fornix. Specific Aim 3 will determine the role of glucocorticoid feedback in brainstem and hippocampus on hormonal response of the fetal HIM to hypoxemia by stereotaxically implanting cortisol crystals into preselected areas of the brainstem and hippocampus. Specific site selection will be based on GR concentration as measured by ICC and mRNA analysis. In all three Specific Aims, peripheral plasma radioimmunoassayable ACTH, AVP, and cortisol and PVN content of CRH and AVP mRNA's will be measured as end points. Hypoxemia and prematurity are important causes of morbidity and mortality in human pregnancy. In addition, hypoxemia is used in this proposal as a repeatable, precisely determined acute stress stimulus allowing us to fine tune our methods and make comparison to the long term studies of the more complex and less understood causes of initiation of parturition. These studies arise logically out of Project I and II of the previous funding period in which we demonstrated, among other results, that lesions of the PVN abolished the fetal ACTH response to hypoxemia and hypotension and prolonged gestation. Detailed controlled experimental data on fetal responses to hypoxemia and in vivo mechanisms of initiation of parturition can only be obtained in fetal animals. This project closely interacts with Projects I and III at both cross-species and mechanistic levels. Our experiments have important relevance to the problem of fetal damage and wastage. Also, these studies will improve our understanding of nervous and endocrine interactions in control of HHAA function.