BRAIN MACROPHAGES AND REPRODUCTIVE AGING

当衰老的雌性大鼠不再能产生促性腺激素的激增来回应周期中期的雌激素激增(正反馈)时，她的生殖生命就结束了。正常情况下，这种下丘脑老化是渐进的，表现为过氧化物酶在下丘脑的星形胶质细胞中积累。我们已经证明了雌激素诱导的突触收缩、神经胶质细胞突起的细化和正反馈之间的关系。为了解释这种衰老，我们提出，在周期的大鼠，驻留在弓状核的脑巨噬细胞被雌激素反复激活并产生自由基，进而使下丘脑功能丧失。在对这一假设的检验中：1)在周期雌性大鼠中，我们将表征弓状核巨噬细胞的生理性中期激活，并利用光镜和电子显微镜免疫标记神经元标记物微管相关蛋白2(MAP 2)、星形胶质细胞标记物胶质纤维酸性蛋白(GFAP)、巨噬细胞标记物OX42以及细胞黏附分子1(I-CAM-1)的出现来确定弓状核突触/神经元、星形胶质细胞和脑巨噬细胞的突触可塑性之间的形态关系。2)我们建议使用维生素E来延缓正常衰老大鼠的生殖衰老，维生素E是一种抗氧化剂，以前已被证明可以抑制中枢神经系统中自由基的产生。正常衰老动物的生殖周期将通过阴道涂片和血液促黄体生成素测量进行监测。在三个不同的衰老里程碑(3个月龄、11个月龄和15个月龄)，对照组和维生素E组的雌性将通过光镜和电子显微镜免疫细胞化学进行脑巨噬细胞标志物的研究，并结合定量突触学进行研究。繁殖失败通常在11个月后明显，在15个月后完成。因此，我们将从功能和形态上描述衰老的特征，并确定维生素E的保护作用。3)为了在体外确定雌激素激活脑巨噬细胞的细胞基础，我们将处理小胶质细胞、星形胶质细胞以及表达α、β或α/β雌激素受体的神经细胞系的原代培养，单独和联合使用，评估雌激素单独或与维生素E或超氧化物歧化酶一起对这些细胞的影响。我们将通过原位化学发光和组织化学来评估这些培养物中活性氧的产生。

The aging female rat's reproductive life ends when she can no longer produce a surge of gonadotrophin in response to the midcycle surge of estrogen (positve feedback). Normally, this hypothalamic aging is gradual, marked by peroxidase accumulation in astroglia in the hypothalamus. We have shown a relationship between estrogen-induced synaptic retraction, elaboration of glial processes and positive feedback. To explain this aging, we propose that in the cycling rat, resident brain macrophages in the arcuate nucleus are repeatedly activated by estrogen and produce free radicals that, in turn, disables hypothalamic function. In testing this hypothesis: 1) In cycling female rats, we will characterize the physiological mid cycle activation of arcuate nucleus macrophages and determine the morphological relationship between synaptic plasticity of arcuate nucleus synapses/neurons, astrocytes and brain macrophages using light and electron microscopic immunolabeling for the neuronal marker microtubule-associated protein 2 (MAP 2), the astroglia marker glial fibrillary acidic protein (GFAP), the macrophage marker OX42, and, the appearance of a cell adhesion molecule 1 (I-CAM-1) to mark activated macrophages. 2) We propose to delay the onset of reproductive senescence in normally aging rats with the administration of vitamin E, an anti-oxidant that previously has been shown to suppress free radical production in the central nervous system. The reproductive cycles of normally aging animals will be monitored by vaginal smears and blood LH measurements. At three distinct aging milestones (3 months old, 11 months old and 15 months), control and vitamin E-treated females will be studied by light- and electron microscopic immunocytochemistry for brain macrophage markers in combination with quantitative synaptology will be carried out. The failure of reproduction is normally evident by eleven months and completed by 15 months. Thus, we will characterize aging functionally and morphologically and determine the protective effect of vitamin E. 3) To determine in vitro the cellular basis of activation of brain macrophages by estrogen, we will treat primary cultures of microglia, astrocytes, and, neuronal cell lines that express alpha, beta or alpha/beta estrogen receptors, alone and in combination, assess the effects of estrogen, alone or with vitamin E or superoxide dismutase on these. We will assess production of reactive oxygen species in these cultures by in-situ chemiluminescence and histochemistry.