Institute for Physiology II, University Muenster, Muenster, Germany.
Department of Pharmacy and Applied Sciences, La Trobe Institute for Molecular Sciences, La Trobe University, Bendigo, Victoria, Australia.
Acta Physiol (Oxf). 2019 Jul;226(3):e13261. doi: 10.1111/apha.13261. Epub 2019 Mar 4.
Dantrolene interacts with ryanodine receptors in skeletal and cardiac muscle affecting sarcoplasmic reticulum calcium release. Since dantrolene is lipophilic it could also affect intra-sarcoplasmic reticulum calcium handling proteins such as calsequestrin. This study investigated whether dantrolene (1-30 µmol/L) alters the polymerization state and the calcium binding capacity of recombinant cardiac calsequestrin and cardiac sarcoplasmic reticulum calcium handling.
Human recombinant cardiac calsequestrin was used to make simultaneous measurements of turbidity (to indicate calsequestrin polymerization) and calcium binding to calsequestrin in the presence and absence of dantrolene. Caffeine-induced Ca transients were used to investigate the effects of dantrolene on sarcoplasmic reticulum calcium loading and release in saponin-permeabilized cardiomyocytes laid down in monolayers in 96-well array plates.
Dantrolene (1-30 µmol/L) increased the polymerization state of calsequestrin and its calcium binding capacity. In the presence of dantrolene, calsequestrin-dependent turbidity increased 2.5-11.5-fold at 1.0 mmol/L calcium added to unbuffered Ca solutions and 3-10-fold when calcium was raised from 0.06 to 30 µmol/L. The dantrolene-dependent increase in turbidity at 30 µmol/L dantrolene was associated with a 3-fold increase in the number of calcium ions bound per calsequestrin molecule at 30 and 100 µmol/L calcium. The caffeine-induced releasable calcium loaded by the sarcoplasmic reticulum at 0.63 µmol/L free calcium in permeabilized cardiomyocytes was also increased in the presence of 30 µmol/L dantrolene.
Dantrolene alters the polymerization state and the calcium binding properties of cardiac calsequestrin and increases sarcoplasmic reticulum calcium loading in permeabilized cardiomyocytes.
丹曲林钠与骨骼肌和心肌中的兰尼碱受体相互作用，影响肌浆网钙离子释放。由于丹曲林钠具有亲脂性，它也可能影响肌浆网内的钙处理蛋白，如钙调蛋白。本研究旨在探讨丹曲林钠（1-30 μmol/L）是否改变重组人心肌钙调蛋白的聚合状态和钙结合能力，以及心脏肌浆网钙离子处理。
使用人重组人心肌钙调蛋白，在存在和不存在丹曲林钠的情况下，同时测量浊度（表示钙调蛋白聚合）和钙与钙调蛋白的结合。在单层铺板的 96 孔阵列板中用皂素通透的心肌细胞，用丹曲林钠处理，测量咖啡因诱导的 Ca 瞬变，研究丹曲林钠对肌浆网钙离子加载和释放的影响。
丹曲林钠（1-30 μmol/L）增加了钙调蛋白的聚合状态和钙结合能力。在丹曲林钠存在的情况下，当无缓冲 Ca 溶液中加入 1.0 mmol/L 钙离子时，钙调蛋白依赖的浊度增加 2.5-11.5 倍，当从 0.06 增加到 30 μmol/L 时，浊度增加 3-10 倍。在 30 μmol/L 丹曲林钠存在下，丹曲林钠依赖的浊度增加与 30 和 100 μmol/L 钙时钙调蛋白结合的钙离子数量增加 3 倍有关。在 0.63 μmol/L 游离钙的皂素通透的心肌细胞中，肌浆网可释放的钙离子加载量也增加了，在 30 μmol/L 丹曲林钠存在的情况下，肌浆网可释放的钙离子加载量也增加了。
丹曲林钠改变了心肌钙调蛋白的聚合状态和钙结合特性，并增加了皂素通透的心肌细胞中肌浆网的钙离子负载。
