Liévano A, Vega-SaenzdeMiera E C, Darszon A
Departamento de Bioquimica, Centro de Investigación y de Estudios Avanzados del IPN, México, DF.
J Gen Physiol. 1990 Feb;95(2):273-96. doi: 10.1085/jgp.95.2.273.
Ca2+ influx across the sea urchin sperm plasma membrane is a necessary step during the egg jelly-induced acrosome reaction. There is pharmacological evidence for the involvement of Ca2+ channels in this influx, but their presence has not been directly demonstrated because of the small size of this cell. Sea urchin sperm Ca2+ channels are being studied by fusing isolated plasma membranes into planar lipid bilayers. With this strategy, a Ca2+ channel has been detected with the following characteristics: (a) the channel exhibits a high mainstate conductance (gamma MS) of 172 pS in 50 mM CaCl2 solutions with voltage-dependent decaying to smaller conductance states at negative Em; (b) the channel is blocked by millimolar concentrations of Cd2+, Co2+, and La3+, which also inhibit the egg jelly-induced acrosome reaction; (c) the gamma MS conductance sequence for the tested divalent cations is the following: Ba2+ greater than Sr2+ greater than Ca2+; and (d) the channel discriminates poorly for divalent over monovalent cations (PCa/PNa = 5.9). The sperm Ca2+ channel gamma MS rectifies in symmetrical 10 mM CaCl2, having a maximal slope conductance value of 94 pS at +100 mV applied to the cis side of the bilayer. Under these conditions, a different single-channel activity of lesser conductance became apparent above the gamma MS current at positive membrane potentials. Also in 10 mM Ca2+ solutions, Mg2+ permeates through the main channel when added to the cis side with a PCa/PMg = 2.9, while it blocks when added to the trans side. In 50 mM Ca2+ solutions, the gamma MS open probability has values of 1.0 at voltages more positive than -40 mV and decreases at more negatives potentials, following a Boltzmann function with an E0.5 = -72 mV and an apparent gating charge value of 3.9. These results describe a novel Ca2(+)-selective channel, and suggest that the main channel works as a single multipore assembly.
钙离子穿过海胆精子质膜的内流是卵黄膜诱导顶体反应过程中的一个必要步骤。有药理学证据表明钙离子通道参与了这种内流,但由于这种细胞体积小,其存在尚未得到直接证实。通过将分离的质膜融合到平面脂质双分子层中来研究海胆精子钙离子通道。采用这种策略,检测到一种具有以下特征的钙离子通道:(a) 在50 mM氯化钙溶液中,该通道表现出172 pS的高主态电导(γMS),在负膜电位下电压依赖性衰减至较小的电导状态;(b) 该通道被毫摩尔浓度的镉离子、钴离子和镧离子阻断,这些离子也抑制卵黄膜诱导的顶体反应;(c) 测试的二价阳离子的γMS电导顺序如下:钡离子>锶离子>钙离子;(d) 该通道对二价阳离子和一价阳离子的区分能力较差(PCa/PNa = 5.9)。精子钙离子通道γMS在对称的10 mM氯化钙中整流,在施加到双分子层顺侧的+100 mV时最大斜率电导值为94 pS。在这些条件下,在正膜电位下,高于γMS电流时出现了一种电导较小的不同单通道活性。同样在10 mM钙离子溶液中,当镁离子添加到顺侧时,它以PCa/PMg = 2.9的比例透过主通道,而当添加到反侧时则阻断通道。在50 mM钙离子溶液中,γMS开放概率在电压比-40 mV更正时的值为1.0,在更负的电位下降低,遵循具有E0.5 = -72 mV和表观门控电荷值为3.9的玻尔兹曼函数。这些结果描述了一种新型的钙离子选择性通道,并表明主通道作为单个多孔隙组装体起作用。