McCulloh D H, Chambers E L
Department of Physiology and Biophysics, University of Miami School of Medicine, Florida 33101.
J Gen Physiol. 1992 Feb;99(2):137-75. doi: 10.1085/jgp.99.2.137.
The early events of fertilization that precede and cause activation of an egg have not been fully elucidated. The earliest electrophysiological change in the sea urchin egg is a sperm-evoked increase of the egg's membrane conductance. The resulting depolarization facilitates entry of the fertilizing sperm and precludes the entry of supernumerary sperm. The sequence of the increase in the egg's membrane conductance, gamete membrane fusion, egg activation, and sperm entry, including causal relationships between these events, are not known. This study reports the use of whole egg voltage clamp and loose patch clamp to monitor simultaneously changes of membrane conductance and capacitance at the site of sperm-egg contact. Measurements were made during sperm-egg interactions where sperm entry readily proceeded or was precluded by maintaining the egg's membrane potential either at large, negative values or at positive values. Whenever the sperm evoked an increase of the egg's membrane conductance, that increase initiated abruptly, was localized to the site of sperm attachment, and was accompanied by a simultaneous abrupt increase of the membrane capacitance. This increase of capacitance indicated the establishment of electrical continuity between gametes (possibly fusion of the gametes' plasma membranes). If sperm entry was blocked by large negative membrane potentials, the capacitance cut off rapidly and simultaneously with a decrease of the membrane conductance, indicating that electrical continuity between gametes was disrupted. When sperm entry was precluded by positive membrane potentials, neither conductance nor capacitance increased, indicating that sperm entry was halted before the fusion of membranes. A second, smooth increase of capacitance was associated with the exocytosis of cortical granules near the sperm in eggs that were activated. Electrical continuity between the gametes always preceded activation of the egg, but transient electrical continuity between the gametes alone was not always sufficient to induce activation.
在卵子激活之前发生并导致卵子激活的早期受精事件尚未完全阐明。海胆卵子最早的电生理变化是精子诱发的卵子膜电导增加。由此产生的去极化促进了受精精子的进入,并阻止了多余精子的进入。卵子膜电导增加、配子膜融合、卵子激活和精子进入的顺序,包括这些事件之间的因果关系,尚不清楚。本研究报告了使用全卵电压钳和膜片钳同时监测精子与卵子接触部位的膜电导和电容变化。在精子与卵子相互作用期间进行测量,此时精子进入很容易进行,或者通过将卵子的膜电位维持在大的负值或正值来阻止精子进入。每当精子诱发卵子膜电导增加时,这种增加会突然开始,局限于精子附着部位,并伴随着膜电容的同时突然增加。这种电容增加表明配子之间建立了电连续性(可能是配子质膜的融合)。如果精子进入被大的负膜电位阻断,电容会迅速切断,同时膜电导降低,这表明配子之间的电连续性被破坏。当精子进入被正膜电位阻止时,电导和电容都不会增加,这表明精子进入在膜融合之前就停止了。在被激活的卵子中,靠近精子的皮质颗粒的胞吐作用与电容的第二次平滑增加有关。配子之间的电连续性总是先于卵子的激活,但仅配子之间的短暂电连续性并不总是足以诱导激活。