Bakker A J, Lamb G D, Stephenson D G
School of Physiology and Pharmacology, University of New South Wales, Sydney, Australia.
J Muscle Res Cell Motil. 1996 Feb;17(1):55-67. doi: 10.1007/BF00140324.
In this study, we investigated the effect of the Ca2+ pump inhibitor, 2,5-di-(tert-butyl)-1,4-hydroquinone on the function of the contractile apparatus, Ca2+ uptake, the permeability of the sarcoplasmic reticulum to Ca2+ and excitation-contraction coupling, in mechanically skinned muscle fibres of the rat and toad. 2,5-di-(tert-butyl)-1,4-hydroquinone had no significant effect on the maximum force and Ca2+ sensitivity of the contractile apparatus in rat and toad fibres at concentrations of 20 and 5 microM respectively. In rat fibres, 2,5-di-(tert-butyl)-1,4-hydroquinone was found to inhibit sarcoplasmic reticulum Ca2+ loading in a dose dependent manner, with a half maximal effect at 2 microM. In toad fibres, 5 microM 2,5-di-(tert-butyl)-1,4-hydroquinone completely blocked sarcoplasmic reticulum Ca2+ loading. Exposure to 5 mM BAPTA revealed a small resting sarcoplasmic reticulum Ca2+ leak in unstimulated rat fibres. This Ca2+ leak was not significantly affected by the presence of 20 microM 2,5-di-(tert-butyl)-1,4-hydroquinone, suggesting that 2,5-di-(tert-butyl)-1,4-hydroquinone does not substantially block or activate the sarcoplasmic reticulum Ca2+ release channels. Depolarisation-induced force responses elicited in rat and toad skinned fibres were not significantly affected by 0.5 microM 2,5-di-(tert-butyl)-1, 4-hydroquinone. In the rat fibres, 5 and 20 microM 2,5-di-(tert-butyl)-1,4-hydroquinone greatly increased the peak and duration of initial depolarisation-induced force responses, while subsequent responses were reduced. 2,5-di-(tert-butyl)-1,4-hydroquinone did not affect excitation contraction coupling, as depolarisation-induced force responses similar to initial controls could be elicited after 2,5-di-(tert-butyl)-1,4-hydroquinone exposure, provided that the initial Ca2+ release in 2,5-di-(tert-butyl)-1,4-hydroquinone was chelated with 0.5 mM EGTA (to prevent Ca(2+)-dependent damage) and the sarcoplasmic reticulum was reloaded with Ca2+. In the toad fibres, 5 microM 2,5-di-(tert-butyl)-1, 4-hydroquinone had a similar effect on depolarisation-induced force responses to that observed at 20 microM 2,5-di-(tert-butyl)-1, 4-hydroquinone in rat fibres. This study shows that 2,5-di-(tert-butyl)-1,4-hydroquinone specifically and reversibly inhibits the sarcoplasmic reticulum Ca2+ pump in skeletal muscle and therefore, 2,5-di-(tert-butyl)-1,4-hydroquinone could be a valuable tool for investigating the role of the sarcoplasmic reticulum in Ca2+ homeostasis in skeletal muscle.
在本研究中，我们调查了Ca2+泵抑制剂2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚对大鼠和蟾蜍机械去表皮肌纤维收缩装置功能、Ca2+摄取、肌浆网对Ca2+的通透性以及兴奋 - 收缩偶联的影响。在大鼠和蟾蜍纤维中，2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚浓度分别为20微摩尔和5微摩尔时，对收缩装置的最大力量和Ca2+敏感性没有显著影响。在大鼠纤维中，发现2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚以剂量依赖性方式抑制肌浆网Ca2+负载，在2微摩尔时达到半数最大效应。在蟾蜍纤维中，5微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚完全阻断肌浆网Ca2+负载。暴露于5毫摩尔的BAPTA显示未刺激的大鼠纤维中存在少量静息肌浆网Ca2+泄漏。20微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚的存在对这种Ca2+泄漏没有显著影响，表明2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚不会实质性地阻断或激活肌浆网Ca2+释放通道。0.5微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚对大鼠和蟾蜍去表皮纤维中去极化诱导的力量反应没有显著影响。在大鼠纤维中，5微摩尔和20微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚极大地增加了初始去极化诱导的力量反应的峰值和持续时间，而随后的反应则降低。2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚不影响兴奋 - 收缩偶联，因为在2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚暴露后，如果用0.5毫摩尔的EGTA螯合2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚中的初始Ca2+释放（以防止Ca(2 +)依赖性损伤）并使肌浆网重新负载Ca2+，则可以引发与初始对照相似的去极化诱导的力量反应。在蟾蜍纤维中，5微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚对去极化诱导的力量反应的影响与在大鼠纤维中20微摩尔的2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚观察到的相似。本研究表明，2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚特异性且可逆地抑制骨骼肌中的肌浆网Ca2+泵，因此，2,5 - 二 -（叔丁基）- 1,4 - 对苯二酚可能是研究肌浆网在骨骼肌Ca2+稳态中作用的有价值工具。
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