Käser-Glanzmann R, Jakäbovä M, George J N, Lüscher E F
Biochim Biophys Acta. 1977 May 2;466(3):429-40. doi: 10.1016/0005-2736(77)90336-4.
The events involved in platelet shape change, aggregation, the release reaction and contraction are thought to be mediated by the availability of Ca2+. Increased cytoplasmic calcium, released from intracellular stores, triggers platelet activity, and increased concentration of adenosine 3',5'-cyclic monophosphate (cyclic AMP) inhibits platelet alterations. We have studied the hypothesis that cyclic AMP may regulate the level of platelet cytoplasmic calcium by stimulating calcium removal by a membrane system. Such a hypothesis would be consistent with the reversibility of most manifestations of platelet activation. Human platelets were sonicated and unlysed platelets, mitochondria and granules were removed by centrifugation at 19 000 X g. Electron microscopy shows that the sediment, after centrifugation of the supernatant at 40 000 X g consists to a large extent of membrane vesicles. Such preparations actively concentrate calcium, as measured by the uptake of 45Ca, and also have the maximal calcium-stimulated ATPase activity. Optimal calcium uptake requires ATP and oxalate, and release of calcium from loaded vesicles was stimulated by the calcium ionophore A23187 and inhibited by LaCl3. These data indicate that calcium was being actively concentrated within membrane vesicles. After washing of such preparations in the absence of ATP, their capacity to take up Ca2+ is reduced to an initial value of 2.8 nmol/mg protein per min. In the presence of 2 - 10(6) M cyclic AMP to which was added a protein kinase preparation from human platelets, up to a 3-fold increase of this rate of uptake was observed. These results suggest that in platelets, as in muscle, cyclic AMP is a regulatory factor in the control of cytoplasmic calcium. Although the cyclic nucleotide may have still other functions, it appears likely that the well-known inhibition of many platelet activities by high intracellular cyclic AMP concentrations is directly linked to the stimulation of the removal of Ca2+ from the cytoplasm.
血小板形状改变、聚集、释放反应及收缩所涉及的过程被认为是由钙离子的可利用性介导的。从细胞内储存库释放的细胞质钙离子增加会触发血小板活性，而腺苷3',5'-环磷酸（环磷酸腺苷，cAMP）浓度升高则会抑制血小板的改变。我们研究了这样一个假说，即环磷酸腺苷可能通过刺激膜系统移除钙离子来调节血小板细胞质钙离子水平。这样的假说与血小板激活的大多数表现的可逆性是一致的。将人血小板进行超声处理，然后通过19000×g离心去除未裂解的血小板、线粒体和颗粒。电子显微镜显示，上清液在40000×g离心后的沉淀物在很大程度上由膜囊泡组成。通过45Ca摄取量测定，这种制剂能主动浓缩钙离子，并且还具有最大的钙刺激ATP酶活性。最佳的钙摄取需要ATP和草酸盐，钙离子载体A23187能刺激加载囊泡中钙离子的释放，而LaCl3则能抑制这种释放。这些数据表明钙离子正在被主动浓缩在膜囊泡内。在无ATP的情况下洗涤这种制剂后，其摄取Ca2+的能力降低至每分钟每毫克蛋白质2.8 nmol的初始值。在加入来自人血小板的蛋白激酶制剂的2 - 10(6) M环磷酸腺苷存在的情况下，观察到这种摄取速率增加了高达3倍。这些结果表明，在血小板中，如同在肌肉中一样，环磷酸腺苷是控制细胞质钙离子的一个调节因子。尽管环核苷酸可能还有其他功能，但细胞内高浓度环磷酸腺苷对许多血小板活性的众所周知的抑制作用似乎很可能与刺激细胞质中钙离子的移除直接相关。
