Requirements for different Ca2+ pools in the activation of rabbit platelets. I. release reaction and protein phosphorylation.
Requirements for different Ca2+ pools in the activation of rabbit platelets. I. release reaction and protein phosphorylation.
复制标题
兔血小板活化对不同Ca2池的要求。
DOI：
10.1016/0304-4165(82)90159-3
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发表时间：
1982
期刊：
影响因子：
--
通讯作者：
Lyons,RM
中科院分区：
文献类型：
--
作者：
Shaw,JO;Lyons,RM
我们研究了钙离子的作用，细胞外和细胞内的起源，在释放反应和蛋白磷酸化在兔血小板刺激血小板活化因子（乙酰甘油醚磷酸胆碱），凝血酶，或离子载体A23187。在细胞外Ca ~（2+）存在的情况下，8-（N，N-二乙基氨基）辛基-3，4，5-三甲氧基苯甲酸酯（TMB-8），一种细胞内Ca ~（2+）转运的假定拮抗剂，在40 μM的半最大抑制剂浓度下阻断血小板活化因子引发的5-羟色胺释放，而凝血酶诱导的释放为350 μ M，A23187诱导的释放大于500 μM。TMB-8可抑制血小板活化因子诱导的两种血小板蛋白Mr = 41000（P7 P）和20000（P9 P）的磷酸化，这种作用与清除细胞外Ca ~（2+）所引起的作用是相加的。TMB-8对凝血酶或A23187刺激的血小板中的磷酸化仅表现出轻微至不存在的抑制作用。与P9 P磷酸化相反，由血小板活化因子引起的P7 P磷酸化更依赖于TMB-8敏感步骤而不是细胞外Ca 2+的可用性。用含有固定浓度游离Ca 2+的缓冲液进行的实验表明，当受血小板活化因子和凝血酶刺激时，这两个过程（释放和磷酸化）对细胞外游离Ca 2+具有相同的阈值要求（1-3 μM）。这些研究提供的证据表明，血小板活化因子对兔血小板的刺激比凝血酶引起的刺激更依赖于TMB-8敏感的细胞内Ca 2+源。此外，我们的数据表明，激活不同的细胞内过程参与血小板分泌（如P7 P和P9 P磷酸化）可能需要从不同的钙池。
We examined the role of Ca2+, both extracellular and intracellular in origin, in the release reaction and protein phosphorylation in rabbit platelets stimulated with platelet activating factor (acetylglyceryl ether phosphorylcholine), thrombin, or ionophore A23187. In the presence of extracellular Ca2+, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8), a putative antagonist of intracellular Ca2+transport, blocked platelet activating factor-initiated serotonin release at a half-maximal inhibitor concentration of 40 μM, compared to 350 μM for thrombin-induced release and greater than 500 μM, for A23187-induced release. Platelet activating factor-induced phosphorylation of two platelet proteins ofMr=41 000 (P7P) and 20 000 (P9P) was inhibited by TMB-8, an effect which was additive to that caused by removing extracellular Ca2+. TMB-8 demonstrated only minor to non-existant inhibitory effects on phosphorylation in thrombin- or A23187-stimulated platelets. In contrast to P9P phosphorylation, phosphorylation of P7P caused by platelet activating factor was more dependent on a TMB-8 sensitive step than on the availability of extracellular Ca2+. Experiments with buffers containing fixed concentrations of free Ca2+revealed that both processes (release and phosphorylation), when stimulated by platelet activating factor and thrombin, had the same threshold requirement (1–3 μM) for extracellular free Ca2+. These studies provide evidence that stimulation of rabbit platelets by platelet activating factor is more dependent on a TMB-8-sensitive intracellular Ca2+source than is stimulation caused by thrombin. Furthermore, our data indicate that activation of different intracellular processes involved in platelet secretion (such as P7P and P9P phosphorylation) may require Ca2+from different pools.