SPERM-ACTIVATED CHANNELS IN ASCIDIAN OOCYTE MEMBRANES

本研究的目的是研究海鞘中的离子电流 卵母细胞膜被精子激活。 的 研究应将精子激活通道的出现与 施肥潜力，表征离子通道 特异性和动力学，并测试通道是 从卵母细胞膜的细胞质侧被Ca激活。 这些实验使用海鞘，因为去角质卵母细胞是 由于海鞘卵母细胞缺乏皮质颗粒： 单通道记录提供了一个分子描述的 精子激活电流，以及皮质颗粒的缺失， 将通道引入卵母细胞的潜在机制 膜的 此外，膜片钳技术将特定区域 卵母细胞膜，我们将利用这一特点，以限制网站的 通过将精子放入记录吸管来观察配子相互作用。 这 实验，与添加精子的补丁实验相结合 到浴中，并结合全细胞记录， 受精流，将有助于确定渠道在哪里 有多少通道被精子激活， 以及通道从卵母细胞膜的哪一侧打开。 总的目的是了解精子激活通道如何与 受精过程中发生的其他事件，并确定主要的 卵母细胞激活的触发器。 这些实验将有助于整体 了解精子如何激活卵母细胞，他们将进一步 科学和体外受精的实验控制。

The purpose of this proposal is to study the ionic currents in the Ascidian oocyte membrane that are activated by the fertilizing spermatozoon. The research shall correlate the apearance of sperm-activated channels with the fertilization potential, characterize the channels with regard to ion specificity and kinetics, and test the hypothesis that the channels are activated from the cytoplasmic side of the oocyte membrane by Ca. The experiments make use of Ascidians because the dechorinated oocytes are easily patch-clamped and because Ascidian oocytes lack cortical granules: Single-channel recording provides a molecular description of the sperm-activated currents, and the absence of cortical granules elininates a potential mechanism for the introduction of channels into the oocyte membrane. In addition, the patch-clamp technique isolates a defined area of oocyte membrane, and we shall use this feature to restrict the site of gamete interaction by putting sperm in the recording pipette. This experiment, in combination with patch experiments in which sperm are added to the bath, and in combination with the whole-cell recording of fertilization currents, will help to determine where the channels are located, how many channels are activated by the fertilizing spermatozoon, and from which side of the oocyte membrane the channels are opened. The overall aim is to learn how sperm-activated channels interrelate with other events that occur during fertilization, and to determine the primary trigger of oocyte activation. These experiments will aid in the overall understanding of how sperm activate oocytes, and they will further the science and the experimental control of in vitro fertilization.