The mechanism of calcium channel facilitation in bovine chromaffin cells
The mechanism of calcium channel facilitation in bovine chromaffin cells
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DOI：
10.1113/jphysiol.1996.sp021524
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发表时间：
1996-08-01
影响因子：
5.5
通讯作者：
Garcia, AG
中科院分区：
文献类型：
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作者：
Albillos, A;Gandia, L;Garcia, AG
1.本研究旨在阐明去极化预脉冲对电压钳位牛嗜铬细胞钙通道易化的机制。通过研究可溶性囊泡裂解物（SVL）对全细胞Ba 2+电流（I-Ba）的影响，验证了自分泌调节此类通道的假设。SVL由牛肾上腺髓质匀浆制备。该浓缩SVL中的ATP含量为3.18 +/- 0.12 mM（n = 4）。SVL中存在的去甲肾上腺素和肾上腺素的浓度分别为11.2 +/- 0.97和15.2 +/- 2 mM（n = 5）。SVL在外部溶液中的1：1000稀释度使I-Ba的幅度减半，并使其活化动力学减慢7倍。SVL的阻断作用呈浓度依赖性，洗脱后可迅速逆转. SVL对I-Ba动力学的抑制和减慢可被强去极化预脉冲（+90 mV，45 ms）部分逆转。这种抑制的逆转，称为Ca 2+通道促进，在3 μ M硝苯地平的存在下持续存在。胞内透析GDP-β-S（0.5 mM）或用百日咳毒素预处理细胞（100 ng ml（-1）18-24 h）可防止SVL 1：100稀释引起的峰电流降低;在这些条件下未观察到前脉冲易化。5.受体阻断剂纳洛酮（10 μ M）或苏拉明（100 μ M）和PPADS（100 μ M）在很大程度上拮抗SVL的作用。用碱性磷酸酶处理SVL或用盐水缓冲液透析以除去低分子量物质（
1. This study was planned to clarify the mechanism of Ca2+ channel facilitation by depolarizing prepulses given to voltage-clamped bovine chromaffin cells. The hypothesis for an autocrine modulation of such channels was tested by studying the effects of a soluble vesicle lysate (SVL) on whole-cell Ba2+ currents (I-Ba).2. SVL was prepared from a bovine adrenal medullary homogenate. The ATP content in this concentrated SVL amounted to 3.18 +/- 0.12 mM (n = 4). The concentration of noradrenaline and adrenaline present in the SVL was 11.2 +/- 0.97 and 15.2 +/- 2 mM, respectively (n = 5). A 1:1000 dilution of SVL in the external solution halved the magnitude of I-Ba and produced a 7-fold slowing of its activation kinetics. The blocking effects of SVL were concentration dependent and quickly reversed upon washout.3. Inhibition and slowing of the kinetics of I-Ba by SVL could be partially reversed by strong depolarizing prepulses (+90 mV, 45 ms). This reversal of inhibition, called Ca2+ channel facilitation, persisted in the presence of 3 mu M nifedipine.4. Intracellular dialysis of GDP-beta-S (0.5 mM) or pretreatment of the cells with pertussis toxin (100 ng ml(-1) for 18-24 h) prevented the reduction in peak current caused by a 1:100 dilution of SVL; no prepulse facilitation could be observed under these conditions.5. The receptor blockers naloxone (10 mu M) or suramin (100 mu M) and PPADS (100 mu M) largely antagonized the effects of SVL. Treatment of SVL with alkaline phosphatase or dialysis against a saline buffer to remove low molecular mass materials (