Four-dimensional visualization and quantitative analysis of meiotic spindle movements in live mouse oocytes.
This paper made a different attempt of real-time observation of the meiotic spindle movements in living mouse oocyte using a convenient method. This method was based on an experimental phenomenon discovered in our work. In living mouse oocytes, a high concentration of calcium ions (Ca(2+)) was observed throughout the region occupied by the initial meiotic spindle. After Ca(2+) labelling with Fura-2, a weakly fluorescent area (WFA) appeared on each side of the chromosomes. The activities of the WFAs changed during spindle development. By real-time tracking of WFAs, we were able to indirectly observe the meiotic spindle movements. Occasionally, it was observed that the first meiotic spindle rotated from an orientation parallel to the cortex to become perpendicular, instead of migrating from the oocyte centre to the cortex along its axis. Moreover, we analysed this uncommon rotation of the first meiotic spindle and found that the whole rotation process can be divided into two phases: the early slow-speed rotation and the subsequent rapid-speed rotation. We further characterized this rotation with respect to rotational speed and acceleration at all the stages of development. By using a two-photon laser-scanning microscope in combination with Fura-2 dye that is nondamaging to oocytes, we provide a convenient method for indirect visualization and quantitative analysis of spindle movements by real-time tracking of WFAs. This method is easy to operate and master, and economical with time and effort.
State Key Laboratory of Low-Dimensional Quantum Physics, Department of Physics, Tsinghua University, Beijing 100084, PR China.
N
N
Tian
L
L
Zhang
B
B
Liu
P
P
Wang
Y
Y
Li
W
W
Ma
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002875:Chromosomes / Q000378:metabolism; D005260:Female; D005453:Fluorescence; D016257:Fura-2; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008540:Meiosis; D008677:Metaphase; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism; D009865:Oocytes / Q000166:cytology* / Q000378:metabolism; D012399:Rotation; D008941:Spindle Apparatus / Q000378:metabolism*
D002118:Calcium; D016257:Fura-2