Fernández-Belda F, Gómez-Fernández J C
Departamento de Bioquímica, Facultad de Veterinaria, Universidad de Murcia, Espinardo, Spain.
Biochim Biophys Acta. 1987 Oct 16;903(3):473-9. doi: 10.1016/0005-2736(87)90054-x.
Vesicular fragments of sarcoplasmic reticulum isolated from rabbit skeletal muscle were actively loaded with Ca2+ in the presence of ATP and an ATP-regenerating system using Arsenazo III as metallochromic indicator to monitor Ca2+ movements across the membrane. Once the Ca2+ release is triggered by the presence of tetraphenylboron in the reaction medium, the addition of verapamil or diltiazem gives rise to a net Ca2+ entry inside the vesicles. Preincubation in the presence of verapamil does not abolish the tetraphenylboron-induced Ca2+ release, the verapamil-induced Ca2+ accumulation being still observed. There appears to be a high-affinity site for verapamil titrated in the micromolar concentration range, whereas diltiazem demonstrates more complex behavior when its concentration is raised. This study suggests the existence of a Ca2+ pathway (putative channels) which is blocked by the drugs tested allowing Ca2+ accumulation inside the vesicles owing to the Ca2+-dependent ATPase activity.
从兔骨骼肌分离出的肌浆网囊泡片段，在ATP和ATP再生系统存在的情况下，利用偶氮胂III作为金属显色指示剂来监测Ca2+跨膜运动，从而被主动加载Ca2+。一旦反应介质中存在四苯硼引发Ca2+释放，加入维拉帕米或地尔硫卓会导致囊泡内出现净Ca2+内流。在维拉帕米存在下预孵育并不会消除四苯硼诱导的Ca2+释放，仍可观察到维拉帕米诱导的Ca2+积累。似乎存在一个在微摩尔浓度范围内可滴定的维拉帕米高亲和力位点，而当地尔硫卓浓度升高时，其表现出更复杂的行为。这项研究表明存在一条Ca2+途径（推测为通道），所测试的药物可阻断该途径，由于Ca2+依赖性ATP酶活性，使得Ca2+在囊泡内积累。
