Holmsen H, Daniel J L, Dangelmaier C A, Molish I, Rigmaiden M, Smith J B
Thromb Res. 1984 Dec 1;36(5):419-28. doi: 10.1016/0049-3848(84)90298-6.
Gel-filtered platelets prelabeled with [3H]-arachidonate and [14C]-adenine or [32P]-orthophosphate were stimulated with thrombin in the presence of various concentrations of trifluoperazine (TFP). Based on the presence of [14C]- or [32P]-labeled extracellular adenine nucleotides, TFP, above 50 microM, caused platelet lysis which reached 30-40% at 100 microM. In the non-lytic range (0-50 microM) TFP caused marked inhibition of [3H]-arachidonic acid liberation and [3H]-phosphatidylcholine breakdown which was complete at 25 microM. Breakdown of [3H]-phosphatidylinositol was partially (about 50%) inhibited at 25 microM TFP and little further inhibition occurred above this concentration. These results show that thrombin-induced liberation of [3H]-arachidonic acid occurs entirely by a TFP-sensitive mechanism, and suggest that the major portion of the arachidonate is liberated from phosphatidylcholine with a possible contribution from phosphatidylinositol. Dense granule secretion and acid hydrolase secretion were progressively inhibited by TFP, while the thiazine had only a small effect on phosphorylation of myosin. These results indicate that the inhibition of the secretory processes by TFP is not caused by action of TFP on myosin light chain kinase. It is suggested that the profound effect of TFP on arachidonic acid liberation but not myosin phosphorylation is due to different subcellular localization of these calmodulin-requiring enzymes: phospholipase A2 and myosin light chain kinase. The lipophilic TFP dissolves preferentially in the membranes where it has access to phospholipase A2 but not to myosin light chain kinase.
用[3H] - 花生四烯酸、[14C] - 腺嘌呤或[32P] - 正磷酸盐预标记的凝胶过滤血小板，在不同浓度的三氟拉嗪（TFP）存在下用凝血酶刺激。根据[14C] - 或[32P] - 标记的细胞外腺嘌呤核苷酸的存在情况，浓度高于50微摩尔的TFP会导致血小板裂解，在100微摩尔时裂解率达到30 - 40%。在非裂解范围内（0 - 50微摩尔），TFP显著抑制[3H] - 花生四烯酸的释放和[3H] - 磷脂酰胆碱的分解，在25微摩尔时完全抑制。[3H] - 磷脂酰肌醇的分解在25微摩尔TFP时被部分抑制（约50%），高于此浓度时进一步抑制作用很小。这些结果表明，凝血酶诱导的[3H] - 花生四烯酸释放完全通过TFP敏感机制发生，并表明花生四烯酸的主要部分从磷脂酰胆碱释放，磷脂酰肌醇可能有一定贡献。致密颗粒分泌和酸性水解酶分泌被TFP逐渐抑制，而噻嗪对肌球蛋白磷酸化只有很小的影响。这些结果表明，TFP对分泌过程的抑制不是由TFP对肌球蛋白轻链激酶的作用引起的。有人认为，TFP对花生四烯酸释放有深远影响而对肌球蛋白磷酸化没有影响，是由于这些需要钙调蛋白的酶在亚细胞定位不同：磷脂酶A2和肌球蛋白轻链激酶。亲脂性的TFP优先溶解在膜中，在那里它可以接触到磷脂酶A2，但接触不到肌球蛋白轻链激酶。
