Lang R D, Bronk J R
J Cell Biol. 1978 Apr;77(1):134-47. doi: 10.1083/jcb.77.1.134.
The spray-freeze-etching technique has been used to study energy-linked mitochondrial structural changes in rat liver mitochondria incubated in vitro. The technique involves spraying the suspension of mitochondria into liquid propane at -190 degrees C, and does not require the use of cryoprotectants or chemical fixatives. The results confirmed that freshly isolated mitochondria have a condensed matrix and that this expands at the expense of the outer compartment to give the orthodox configuration when the mitochondria are incubated in a K+ medium in the presence of substrate and phosphate. Addition of adenosine diphosphate (ADP) caused a rapid shrinkage of the matrix compartment, and the time-course and extent of this shrinkage has been measured quantitatively by coupling a rapid sampling device to the spray-freezing apparatus. These data show that for orthodox mitochondria the onset of phosphorylation is accompanied by a reduction of 30% in the matrix volume in 20's, and there is no evidence that the decrease in matrical volume affects the phosphorylation efficiency. These results suggest that natural ionophores in the mitochondrial inner membrane make it permeable enough to permit a rapid readjustment of matrix volume after the addition of ADP, and that the associated ion movement does not cause uncoupling of oxidative phosphorylation.
喷雾冷冻蚀刻技术已被用于研究体外培养的大鼠肝脏线粒体中与能量相关的线粒体结构变化。该技术包括将线粒体悬浮液喷入-190℃的液态丙烷中，且不需要使用冷冻保护剂或化学固定剂。结果证实，新鲜分离的线粒体具有浓缩的基质，当线粒体在底物和磷酸盐存在的K+培养基中孵育时，基质会以牺牲外腔为代价而膨胀，从而呈现出正统的形态。添加二磷酸腺苷（ADP）会导致基质腔迅速收缩，通过将快速采样装置与喷雾冷冻设备相连，已对这种收缩的时间进程和程度进行了定量测量。这些数据表明，对于正统线粒体，磷酸化开始时，基质体积在20秒内减少30%，且没有证据表明基质体积的减少会影响磷酸化效率。这些结果表明，线粒体内膜中的天然离子载体使其具有足够的通透性，以便在添加ADP后能迅速重新调整基质体积，并且相关的离子移动不会导致氧化磷酸化的解偶联。
