McCulloh D H, Chambers E L
Department of Physiology and Biophysics, University of Miami School of Medicine, Florida 33101.
J Gen Physiol. 1991 Mar;97(3):579-604. doi: 10.1085/jgp.97.3.579.
Although activation of a sea urchin egg by sperm leads to three phases of membrane conductance increase in the egg, the mechanism by which the sperm causes these conductance changes is not known. We used the loose patch clamp technique to localize the conductance changes in voltage clamped eggs. A patch of the egg's membrane was isolated from the bath by pressing the loose patch clamp pipette against the egg surface. Sperm added to the bath attached to the surface of the egg in a region other than at the isolated membrane patch. During phase 1 of the activation current, no changes of the membrane conductance were detected. At the time of, and subsequent to the onset of phase 2, large currents recorded between the interior of the patch pipette and the bath were attributed to changes of the seal resistance between the surface of the egg and the pipette. A local change of membrane conductance was observed during phase 2 despite the changes of seal resistance. During phase 2, the large amplitude and short duration of the local membrane conductance increase relative to the membrane, conductance increase for the whole egg during phase 2 indicated that the conductance increase occurred over the entire surface of the egg, but not simultaneously. The time when the peak conductance for the membrane patch occurred, relative to the time of onset for phase 2 in the whole egg, depended on the distance, measured in a straight line, between the site of sperm attachment and the tip of the pipette. These data indicate that the localized conductance increase progressed over the surface of the egg from the site of sperm attachment to the opposite pole of the egg. It is proposed that the local conductance increase, the cortical reaction, and the change of seal resistance are all evoked by a common cytoplasmic message that progresses throughout the cytoplasm of the egg from the site of sperm attachment to the opposite pole of the egg.
尽管精子激活海胆卵会导致卵膜电导增加出现三个阶段,但精子引起这些电导变化的机制尚不清楚。我们使用松散膜片钳技术来定位电压钳制卵中的电导变化。通过将松散膜片钳移液管压在卵表面,从浴液中分离出一片卵膜。添加到浴液中的精子附着在卵表面除分离膜片区域之外的其他区域。在激活电流的第1阶段,未检测到膜电导的变化。在第2阶段开始时及之后,在膜片移液管内部与浴液之间记录到的大电流归因于卵表面与移液管之间封接电阻的变化。尽管封接电阻发生了变化,但在第2阶段仍观察到了膜电导的局部变化。在第2阶段,局部膜电导增加的幅度大且持续时间短,相对于整个卵在第2阶段的膜电导增加而言,这表明电导增加发生在卵的整个表面,但并非同时发生。膜片的峰值电导出现的时间,相对于整个卵中第2阶段开始的时间,取决于精子附着位点与移液管尖端之间沿直线测量的距离。这些数据表明,局部电导增加是从精子附着位点开始,在卵表面朝着卵的相对极推进的。有人提出,局部电导增加、皮层反应和封接电阻的变化都是由一种共同的细胞质信号引发的,该信号从精子附着位点开始,在卵的细胞质中朝着卵的相对极传播。