BRAIN MACROPHAGES AND REPRODUCTIVE AGING

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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.