Dynamic instability and motile events of native microtubules from squid axoplasm.

Native microtubules from extruded axoplasm of squid giant axons were used as a paradigm to characterize the motion of organelles along free microtubules and to study the dynamics of microtubule length changes. The motion of large round organelles was visualized by AVEC-DIC microscopy and analyzed at a temporal resolution of 10 frames per second. The movements were smooth and showed no major changes in velocity or direction. During translocation, the organelles paused very rarely. Superimposed on the rather constant mean velocity was a velocity fluctuation, which indicated that the organelles are subject to considerable thermal motion during translocation. Evidence for a regular low-frequency oscillation was not found. The thermal motion was anisotropic such that axial motion was less restricted than lateral motion. We conclude that the crossbridge connecting the moving organelle to the microtubule has a flexible region that behaves like a hinge, which permits preferential movement in the direction parallel to the microtubule. The dynamic changes in length of native microtubules were studied at a temporal resolution of 1 Hz. About 98% of the native microtubules maintained their length ("stable" microtubules), while 2% showed phases of growing and/or shrinking typical for dynamic instability ("dynamic" microtubules). Gliding and organelle motion were not influenced by dynamic length changes. Transitions between growing and shrinking phases were low-frequency events (1-10 minutes per cycle). However, a new type of microtubule length fluctuation, which occurred at a high frequency (a few seconds per cycle), was detected. The length changes were in the 1-3 micron range. The latter events were very prominent at the (+) ends. It appears that the native axonal microtubules are much more stable than the purified microtubules and the microtubules of cultured cells that have been studied thus far. Potential mechanisms accounting for the three states of microtubule stability are discussed. These studies show that the native microtubules from squid giant axons are a very useful paradigm for studying microtubule-related motility events and microtubule dynamics.

鱿鱼巨大轴突挤出的轴浆中的天然微管被用作范例，以表征细胞器沿游离微管的运动，并研究微管长度变化的动力学。通过AVEC - DIC显微镜观察大的圆形细胞器的运动，并以每秒10帧的时间分辨率进行分析。运动是平滑的，在速度或方向上没有重大变化。在移位过程中，细胞器很少停顿。在相当恒定的平均速度上叠加有速度波动，这表明细胞器在移位过程中受到相当大的热运动影响。没有发现有规律的低频振荡的证据。热运动是各向异性的，轴向运动比侧向运动受到的限制更小。我们得出结论，将移动的细胞器连接到微管的横桥有一个像铰链一样的柔性区域，这允许在平行于微管的方向上优先运动。以1Hz的时间分辨率研究天然微管长度的动态变化。大约98%的天然微管保持其长度（“稳定”微管），而2%显示出动态不稳定性所特有的生长和/或收缩阶段（“动态”微管）。滑动和细胞器运动不受动态长度变化的影响。生长和收缩阶段之间的转变是低频事件（每个周期1 - 10分钟）。然而，检测到一种新型的微管长度波动，其发生频率较高（每个周期几秒）。长度变化在1 - 3微米范围内。后一种情况在（+）端非常显著。似乎天然的轴突微管比迄今为止所研究的纯化微管和培养细胞的微管要稳定得多。讨论了解释微管稳定性三种状态的潜在机制。这些研究表明，鱿鱼巨大轴突的天然微管是研究微管相关的运动事件和微管动力学的非常有用的范例。