STEROID-INITIATED EXOCYTOSIS IN SPERM

顶体反应(AR)是哺乳动物受精的中心事件 涉及精子头膜的融合和开窗。这个 启动这种胞吐过程的分子事件并不完全 明白了。这项研究的目的是增加我们对这些 人类精子中的事件。这些研究将在体外进行 人类精子孵化系统。我的实验室发现黄体酮，一种 类固醇激素存在于卵泡液中，由颗粒细胞分泌 和卵丘细胞，通过以下方式启动人类精子AR 增加精子[Ca2+]。这一点特别有趣，因为：1)精子 在活体人体内必须穿过卵丘细胞外基质 受精；2)[Ca~(2+)]升高如此之快(1分钟)，以至于精子 类固醇表面受体可能参与其中。 这些研究将包括：1)类固醇是否会增加心肌细胞内钙离子浓度 人精子头；2)测定必需的类固醇结构 [Ca~(2+)]和AR的增加；3)有助于确定 是否存在质膜类固醇受体；4)研究 类固醇抑制Na~+，K~+-ATPase和[Ca~(2+)]~(2+)的可能作用 -ATPase使精子头部[Ca~(2+)]升高，从而增加AR。 使用的方法包括：人类精子的体外孵化， 计算机荧光图像分析、电子传输 显微镜，AR的免疫荧光分析， 高效液相色谱、(45)钙外流研究和(32)磷底物的ATPase测定。 希望这些研究将有助于更好地理解 人类受精的分子基础以及未来的铅 新的避孕方法和不孕不育的治疗方法。在……里面 此外，类固醇通过细胞增加细胞内的[钙]离子 表面介导的机制可能与心肌细胞的钙需求事件有关 体细胞。因此，这些研究可能也具有重要意义 肥田。

The acrosome reaction (AR) is a central event of mammalian fertilization involving the fusion and fenestration of sperm head membranes. The molecular events that initiate this exocytotic process are not completely understood. The goal of this research is to increase our knowledge of those events in human sperm. These studies will be carried out with an in vitro human sperm incubation system. My laboratory has found that progesterone, a steroid hormone present in follicular fluid and secreted by granulosa cells and cells of the cumulus oophorus, can initiate the human sperm AR by increasing sperm [Ca2+]. This is of particular interest because: 1) sperm have to pass through the cumulus extracellular matrix during in vivo human fertilization; 2) the [Ca2+] increase is so fast (< 1 min) that a sperm steroid surface receptor may be involved. These studies will include: 1) whether steroids increase [Ca2+] in the human sperm head; 2) determination of the steroid structure necessary for the increase in [Ca2+] and the AR; 3) experiments to help determine whether or not there is a plasma membrane steroid receptor; 4) investigation of the possible role of steroid inhibition of Na+, K+ -ATPase and [Ca2+ + Mg2+] -ATPase in increasing sperm head [Ca2+] and thereby the AR. Methods to be used include: in vitro incubation of human sperm, computerized fluorescence image analysis, transmission electron microscopy,immunofluorescent assay of the AR, HPLC, (45)Ca-efflux studies and ATPase assays with (32)P-substrate. It is hoped that these studies will result in a greater understanding of the molecular basis of human fertilization and thereby lead in the future to new methods of contraception and of treatment for infertility. In addition, the increasing of intracellular [Ca2+] by a steroid via a cell surface-mediated mechanism might have relevance to Ca2+-requiring events in somatic cells. Thus, these studies may also be of importance outside the field of fertilization.