Schlichter L C
Department of Physiology, University of Toronto, Ontario, Canada.
Dev Biol. 1989 Jul;134(1):59-71. doi: 10.1016/0012-1606(89)90078-x.
Ionic currents in immature, ovulated Rana pipiens oocytes (metaphase I) were studied using the voltage-clamp technique. At this stage of maturity the oocyte can produce action potentials in response to depolarizing current or as an "off response" to hyperpolarizing current. Reducing external Na+ to 1/10 normal (choline substituted) eliminated the action potentials and both the negative-slope region and zero-crossing of the I-V relation. Reducing external Cl- to 1/10 or 1/100 normal (methanesulfonate substituted) lengthened the action potential. The outward current was reduced and a net inward current was revealed. By changing external Na+, Cl-, and K+ concentrations and using blocking agents (SITS, TEA), three voltage- and time-dependent currents were identified, INa, IK and ICl. The Na+ current activated at about 0 mV and reversed at very positive values which decreased during maturation. Inward Na+ current produced the upstroke of the action potential. During each voltage-clamp step the Na+ current activated slowly (seconds) and did not inactivate within many minutes. The Na+ current was not blocked by TTX at micromolar concentrations. The K+ current was present only in the youngest oocytes. Because IK was superimposed on a large leakage current, it appeared to reverse at the resting potential. When leakage currents were subtracted, the reversal potential for IK was more negative than -110 mV in Ringer's solution. IK was outwardly rectifying and strongly activated above -50 mV. The outward K+ current produced an after hyperpolarization at the end of each action potential. IK was blocked completely and reversibly by 20 mM external TEA. The Cl- current activated at about +10 mV and was outwardly rectifying. ICl was blocked completely and reversibly by 400 microM SITS added to the bathing medium. This current helped repolarize the membrane following an action potential in the youngest oocytes and was the only repolarizing current in more mature oocytes that had lost IK. The total leakage current had an apparently linear I-V relation and was separated into two components: a Na+ current (IN) and a smaller component carried by as yet unidentified ions.
采用电压钳技术研究了未成熟、已排卵的豹蛙卵母细胞（减数分裂中期I）中的离子电流。在这个成熟阶段，卵母细胞能够响应去极化电流产生动作电位，或者作为对超极化电流的“关闭反应”产生动作电位。将细胞外Na⁺浓度降至正常水平的1/10（用胆碱替代）可消除动作电位以及I-V关系曲线的负斜率区域和零交叉点。将细胞外Cl⁻浓度降至正常水平的1/10或1/100（用甲磺酸盐替代）可延长动作电位。外向电流减小，出现净内向电流。通过改变细胞外Na⁺、Cl⁻和K⁺浓度并使用阻断剂（SITS、TEA），鉴定出三种电压和时间依赖性电流，即INa、IK和ICl。Na⁺电流在约0 mV时激活，在非常正值时反转，在成熟过程中该值会降低。内向Na⁺电流产生动作电位的上升支。在每个电压钳制步骤中，Na⁺电流缓慢激活（数秒），在数分钟内不会失活。微摩尔浓度的TTX不会阻断Na⁺电流。K⁺电流仅存在于最年轻的卵母细胞中。由于IK叠加在较大的漏电流上，它似乎在静息电位处反转。减去漏电流后，在林格氏液中，IK的反转电位比 -110 mV更负。IK呈外向整流，在高于 -50 mV时强烈激活。外向K⁺电流在每个动作电位结束时产生一个超极化后电位。20 mM的细胞外TEA可完全且可逆地阻断IK。Cl⁻电流在约 +10 mV时激活，呈外向整流。向浴液中添加400 μM SITS可完全且可逆地阻断ICl。在最年轻的卵母细胞中，该电流有助于动作电位后膜的复极化，并且是在失去IK的更成熟卵母细胞中唯一的复极化电流。总漏电流具有明显的线性I-V关系，可分为两个成分：一个Na⁺电流（IN）和一个由尚未鉴定的离子携带的较小成分。
