Woodward R M, Miledi R
Department of Psychobiology, University of California, Irvine 92717.
Proc Biol Sci. 1991 Apr 22;244(1309):11-9. doi: 10.1098/rspb.1991.0044.
Electrical recordings were used to study the sensitivity of native Xenopus oocytes to the octapeptide angiotensin II (AII). AII elicited oscillatory currents associated with an increase in membrane conductance to Cl-. Responsiveness to AII varied greatly between oocytes taken from different frogs, and to a lesser extent between oocytes from the same ovary. Oocytes from frogs showing high sensitivity had response thresholds between 0.5-1.0 nM AII, and at a holding potential of -60 mV, responded to 1 microM AII with currents greater than 3 microA. In contrast, oocytes from some frogs gave no response, even to 10 microM AII. A total of 618 oocytes from 79 frogs were tested for sensitivity to AII, and oocytes from 85% of frogs gave detectable electrical responses. Oscillatory Cl- currents elicited by AII were largely independent of extracellular Ca2+, were abolished by chelation of intracellular Ca2+ using EGTA and were mimicked by intraoocyte injection of inositol 1,4,5-trisphosphate (IP3). In addition to oscillatory Cl- currents, AII also evoked an influx of extracellular Ca2+, giving rise to a transient inward Cl- current on membrane hyperpolarizing steps. These experiments all suggested that AII responses were elicited through activation of an intracellular messenger pathway triggered by hydrolysis of inositolphospholipids, mobilization of intracellular Ca2+ by inositol polyphosphates, and activation of Ca(2+)-gated Cl- channels. The effect of manual or enzymic defolliculation on AII responses was studied in nine separate experiments recording from 70 defolliculated oocytes. Efficacy of defolliculation procedures was assayed using scanning electron microscopy, which confirmed removal of 90 to greater than 98% of follicular cells.(ABSTRACT TRUNCATED AT 250 WORDS)
采用电记录法研究非洲爪蟾卵母细胞对八肽血管紧张素II(AII)的敏感性。AII引发振荡电流,同时伴随着膜对Cl-的电导增加。取自不同青蛙的卵母细胞对AII的反应性差异很大,而取自同一卵巢的卵母细胞之间的差异较小。对AII敏感性高的青蛙的卵母细胞,其反应阈值在0.5 - 1.0 nM AII之间,在 - 60 mV的钳制电位下,对1 μM AII的反应电流大于3 μA。相比之下,一些青蛙的卵母细胞即使对10 μM AII也无反应。共检测了来自79只青蛙的618个卵母细胞对AII的敏感性,85%青蛙的卵母细胞给出了可检测到的电反应。AII引发的振荡Cl-电流在很大程度上不依赖细胞外Ca2+,用EGTA螯合细胞内Ca2+可消除该电流,且卵母细胞内注射肌醇1,4,5 - 三磷酸(IP3)可模拟该电流。除了振荡Cl-电流外,AII还引起细胞外Ca2+内流,在膜超极化时产生短暂内向Cl-电流。这些实验均表明,AII反应是通过激活由肌醇磷脂水解触发的细胞内信使途径、肌醇多磷酸动员细胞内Ca2+以及激活Ca(2 +)门控Cl-通道而引发的。在9个独立实验中,对70个去滤泡卵母细胞进行记录,研究了手动或酶促去滤泡对AII反应的影响。采用扫描电子显微镜检测去滤泡程序的效果,结果证实已去除90%至超过98%的滤泡细胞。(摘要截短于250字)