Tesi C, Colomo F, Nencini S, Piroddi N, Poggesi C
Dipartimento di Scienze Fisiologiche, Università degli Studi di Firenze, Italy.
Biophys J. 2000 Jun;78(6):3081-92. doi: 10.1016/S0006-3495(00)76845-7.
In striated muscle, force generation and phosphate (P(i)) release are closely related. Alterations in the [P(i)] bathing skinned fibers have been used to probe key transitions of the mechanochemical coupling. Accuracy in this kind of studies is reduced, however, by diffusional barriers. A new perfusion technique is used to study the effect of [P(i)] in single or very thin bundles (1-3 microM in diameter; 5 degrees C) of rabbit psoas myofibrils. With this technique, it is possible to rapidly jump [P(i)] during contraction and observe the transient and steady-state effects on force of both an increase and a decrease in [P(i)]. Steady-state isometric force decreases linearly with an increase in log[P(i)] in the range 500 microM to 10 mM (slope -0.4/decade). Between 5 and 200 microM P(i), the slope of the relation is smaller ( approximately -0.07/decade). The rate constant of force development (k(TR)) increases with an increase in [P(i)] over the same concentration range. After rapid jumps in [P(i)], the kinetics of both the force decrease with an increase in [P(i)] (k(Pi(+))) and the force increase with a decrease in [P(i)] (k(Pi(-))) were measured. As observed in skinned fibers with caged P(i), k(Pi(+)) is about three to four times higher than k(TR), strongly dependent on final [P(i)], and scarcely modulated by the activation level. Unexpectedly, the kinetics of force increase after jumps from high to low [P(i)] is slower: k(Pi(-)) is indistinguishable from k(TR) measured at the same [P(i)] and has the same calcium sensitivity.
在横纹肌中，力的产生与磷酸盐（P(i)）的释放密切相关。已利用浴于皮肌纤维的[P(i)]变化来探究机械化学偶联的关键转变。然而，这种研究的准确性因扩散屏障而降低。一种新的灌注技术用于研究[P(i)]对兔腰大肌肌原纤维单根或非常细的束状纤维（直径1 - 3微米；5摄氏度）的影响。利用该技术，能够在收缩过程中快速改变[P(i)]，并观察[P(i)]增加和减少对力的瞬态和稳态影响。在500微摩尔至10毫摩尔范围内，稳态等长力随log[P(i)]的增加呈线性下降（斜率为-0.4/十倍）。在5至200微摩尔P(i)之间，该关系的斜率较小（约-0.07/十倍）。在相同浓度范围内，力发展的速率常数（k(TR)）随[P(i)]的增加而增加。在[P(i)]快速变化后，测量了力随[P(i)]增加而降低的动力学（k(Pi(+))）以及力随[P(i)]降低而增加的动力学（k(Pi(-))）。正如在使用笼蔽P(i)的皮肌纤维中所观察到的，k(Pi(+))比k(TR)高约三到四倍，强烈依赖于最终的[P(i)]，且几乎不受激活水平的调节。出乎意料的是，从高[P(i)]跃变到低[P(i)]后力增加的动力学较慢：k(Pi(-))与在相同[P(i)]下测量的k(TR)无法区分，并且具有相同的钙敏感性。
