STUDIES OF SPERM FUNCTION IN MAMMALIAN FERTILIZATION

哺乳动物受精过程中精子功能的研究

• 批准号: 3312954
• 负责人: PATRICIA J OLDS-CLARKE
• 金额: $ 12.17万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-09-01 至 1995-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3312954
• 关键词: acidity /alkalinity; acrosome; calcium; cell membrane; chromosomes; developmental genetics; enzyme inhibitors; fertility promoting drug; fluorescent dye /probe; galactosyltransferases; gene mutation; genes; laboratory mouse; lactalbumin; sperm; sperm capacitation; sperm motility; acidity /alkalinity; acrosome; calcium; cell membrane; chromosomes; developmental genetics; enzyme inhibitors; fertility promoting drug; fluorescent dye /probe; galactosyltransferases; gene mutation; genes; laboratory mouse; lactalbumin; sperm; sperm capacitation; sperm motility

Very little is known about the molecular mechanisms controlling hyperactivation (HA) and how abnormal movements affect sperm transport through the female genital tract and penetration of the egg investments. The PI has shown that sperm from tw32/+ mice (tw32/+ sperm) have abnormal motility: HA occurs prematurely, and most of the nonhyperactivated motile sperm are abnormally slow. The PI will determine whether precocious HA or slow speed is also characteristic of other t haplotypes (Aim IA). Collaborative studies have shown that the axonemes of tw32/+ sperm are unusually sensitive to calcium (Ca). The PI will determine whether the very slow movements of sperm from mice carrying two t haplotypes are also sensitive, by (1) treating intact sperm with drugs that lower the intracellular free [Ca], and (2) measuring the [Ca] necessary to change flagellar curvature in demembranated sperm (Aim IB). Changes in invertebrate flagellar movement are controlled by cAMP, Ca-binding proteins, or both. Preliminary data suggest that tw32/+ sperm may contain elevated cAMP levels. The PI will determine whether there is a relationship between cAMP and premature HA by (1) measuring cAMP in tw32/+ sperm and (2) perturbing cAMP levels to affect the timing of HA (Aim IIA). Also the Ca-binding proteins of tw32/+ sperm flagella will be characterized by diagonal electrophoresis (Aim IIB). Since the PI has evidence that tw32/+ sperm have normal levels of capacitation, acrosome reaction and zona-binding, their only abnormality relevant to penetration of the investments is in motility. Therefore, the PI will determine whether tw32/+ sperm are abnormal in penetration of the investments by counting the numbers of sperm able to penetrate each investment (Aim II). Experiments from the PI's lab suggest that tw32/+ sperm that do not carry the tw32 haplotype (+ t sperm) are dysfunctional in fertilization in vivo. Although the mechanism of dysfunction is unknown, it must occur between the uterus and the site of gamete interaction and could involve abnormal sperm motility. The Pi will test the hypothesis that the dysfunction of + t sperm occurs in passage through the uterotubal junction by determining whether + t sperm can be recovered from oviductal populations, using the polymerase chain reaction to amplify a sequence which is different for + and t DNA (Aim IV). Determine the site of + t dysfunction will help to determine its nature. These experiments will increase our understanding of how Ca controls hyperactivation and how specific types of movements affect sperm fertilizing ability.

关于控制的分子机制知之甚少 过度激活（HA）以及异常运动如何影响精子运输 通过女性生殖道和鸡蛋投资的渗透。 PI表明TW32/+小鼠的精子（TW32/+精子）异常 运动性：HA过早发生，大多数非液化运动 精子异常缓慢。 PI将确定早熟的HA还是 慢速也是其他T单倍型的特征（AIM IA）。 协作研究表明，TW32/+精子的轴突是 对钙（CA）异常敏感。 PI将确定是否 携带两种T单倍型的小鼠的精子运动非常缓慢 敏感，通过（1）用降低的药物治疗完整的精子 细胞内无[CA]和（2）测量更改所需的[CA] 鞭毛精子中的鞭毛曲率（AIM IB）。 变更 无脊椎动物鞭毛运动由cAMP，CA绑定控制 蛋白质，或两者兼而有之。 初步数据表明TW32/+精子可能包含 高架营地。 PI将确定是否有 CAMP与HA之间的关系（1）在TW32/+中测量营地 精子和（2）扰动营地水平影响HA的时间（AIM IIA）。 TW32/+精子鞭毛的CA结合蛋白也将被描述 通过对角线电泳（AIM IIB）。 因为PI有证据表明 TW32/+精子具有正常的电容水平，Adrosom反应和 zona结合，它们与渗透有关的唯一异常 投资在运动。 因此，PI将确定是否 TW32/+精子通过计算投资的渗透率异常 能够穿透每项投资的精子数量（AIM II）。 实验 从Pi的实验室中，提示不带有TW32的TW32/+精子 单倍型（+ T精子）在体内受精中功能失调。 虽然 功能障碍的机制未知，必须在子宫之间发生 配子互动的位置，可能涉及异常精子 运动。 PI将检验 + T功能障碍的假设 精子通过确定通过子宫内交界处发生 是否可以从卵形种群中回收 + t的精子 聚合酶链反应以扩增 + +不同的序列 和T DNA（AIM IV）。 确定 + T功能障碍的位点将有助于 确定其本质。 这些实验将增加我们对 CA如何控制过度激活以及特定类型的运动如何影响 精子受精能力。