SARCOPLASMIC-RETICULUM CALCIUM RELEASE IN FROG SKELETAL-MUSCLE FIBERS ESTIMATED FROM ARSENAZO-III CALCIUM TRANSIENTS
SARCOPLASMIC-RETICULUM CALCIUM RELEASE IN FROG SKELETAL-MUSCLE FIBERS ESTIMATED FROM ARSENAZO-III CALCIUM TRANSIENTS
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DOI：
10.1113/jphysiol.1983.sp014959
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发表时间：
1983-01-01
影响因子：
5.5
通讯作者：
MARSHALL, MW
中科院分区：
文献类型：
--
作者：
BAYLOR, SM;CHANDLER, WK;MARSHALL, MW
从青蛙肌肉中分离出的单根抽搐纤维被注入金属变色染料偶氮胂III。在650或660纳米处测量的与染料相关的吸光度变化被用于估计在动作电位刺激或电压钳制去极化（温度为15 - 17℃）后肌质游离[Ca²⁺]的时间进程。进行了计算机模拟以估计肌浆网（sr）和肌质之间Ca²⁺的通量（在染料含量不超过0.8毫摩尔的纤维中）。在动作电位刺激后，0.2 - 0.3毫摩尔的Ca²⁺从肌浆网进入肌质。与游离[Ca²⁺]的波形相比，肌浆网Ca²⁺释放的波形出现早且短暂。肌钙蛋白速率常数的选择以及小白蛋白的包含对释放波形的形状都没有太大影响；改变这些参数的主要影响是改变幅度。在肌浆网Ca²⁺释放的初始快速阶段之后，有一个更长的、持续的Ca²⁺摄取时期。该摄取的峰值幅度除以肌浆网Ca²⁺泵位点的浓度被用于估计这些位点的周转率。如此获得的数值，2.4 - 6.3秒⁻¹，与先前基于生化实验发表的值非常吻合。肌浆网Ca²⁺泵已知的生化特性可以解释计算出的肌浆网Ca²⁺通量的最后阶段，因此，在该模型的框架内，可以解释游离[Ca²⁺]瞬变的下降阶段。基于在间隔10毫秒的一系列动作电位期间获得的Ca²⁺瞬变进行的模拟表明，与第二个或后续动作电位相关的肌浆网Ca²⁺释放不超过与第一个动作电位相关释放的20 - 30%。同样，基于在电压钳制实验中测量的Ca²⁺瞬变进行的模拟表明，在接近阈值的阶跃去极化期间，肌浆网Ca²⁺释放达到一个早期峰值，随后下降到一个较低水平。这些关于Ca²⁺释放减少的观察结果似乎与肌浆网Ca²⁺通透性的降低更相符，而不是与离子驱动力的降低相符。模拟表明，细胞内Ca²⁺结合位点的存在使得对肌浆网Ca²⁺释放的电压依赖性的估计不如对Ca²⁺瞬变峰值所观察到的那么陡峭。例如，两个实验显示峰值游离[Ca²⁺]有3毫伏的e倍变化，而Ca²⁺释放峰值需要4毫伏的e倍变化。通过使用一个考虑到肌质中已知Ca²⁺结合位点的动力学和稳态特性的定量模型，有助于从肌质游离[Ca²⁺]的测量中推断肌浆网和肌质之间Ca²⁺的移动。
Single twitch fibers, dissected from frog muscle, were injected with the metallochromic dye Arsenazo III. Changes in dye-related absorbance measured at 650 or 660 nm were used to estimate the time course of myoplasmic free [Ca2+] following either action potential stimation or voltage-clamp depolarization (temperature, 15.degree.-17.degree. C). Computer simulations were carried out to estimate the flux of Ca2+ between the sr [sarcoplasmic reticulum] and myoplasm (in fibers containing no more than 0.8 mM-dye). Following action potential stimulation 0.2-0.3 mM-Ca2+ enters the myoplasm from the sr. The wave form of sr Ca2+ release is early and brief compared with the wave form of free [Ca2+]. Neither the selection of troponin rate constants nor the inclusion of parvalbumin has much effect on the shape of the release wave form; the main effect of varying these parameters is to change the magnitude. After the initial, rapid phase of Ca2+ release from the sr there is a longer, maintained period of Ca2+ uptake. The peak magnitude of this uptake divided by the concentration of sr Ca2+ pump sites is used to estimate the turnover rate of the sites. The numbers so obtained, 2.4-6.3 s-1, are in good agreement with previoulsy published values based on biochemical experiments. The known biochemical properties of the sr Ca2+ pump can explain the final phase of the computed sr Ca2+ flux and therefore, within the framework of the model, the falling phase of the free [Ca2+] transient. Simulations, based on Ca2+ transients obtained during a train of action potentials spaced 10 ms apart, indicate that sr Ca2+ release associated with a 2nd or subsequent action potential is no more than 20-30% of the release associated with the 1st action potential. Similarly, simulations based on Ca2+ transients measured in voltage-clamp experiments show that during a step depolarization near threshold the sr Ca2+ release reaches an early peak which is followed by a decline to a smaller level. These observations of reduced Ca2+ release appear to be more consistent with a decrease in sr Ca2+ permeability than with a decrease in driving force on the ion. The simulations show that the presence of intracellular binding sites for Ca2+ makes the estimate of the voltage dependence of sr CA2+ release less steep than that observed for the peak of the Ca2+ transient. For example, two experiments which showed a 3 mV e-fold change in peak free [Ca2+] required a 4 mV e-fold change for the peak of Ca2+ release. Inferences about Ca2+ movements between sr and myoplasm from measurements of myoplasmic free [Ca2+] are aided by the use of a quantitative model that takes into account the kinetic and steady-state properties of the known Ca2+-binding sites in myoplasm.