Cytoskeletal organization of rat oocytes during metaphase II arrest and following abortive activation: a study by confocal laser scanning microscopy.

Abstract

In metaphase II arrested rat oocytes (M II), microtubules were found in the taper-shaped meiotic spindle and in the cytoplasm as asters and free microtubules. Whereas spindle microtubules were acetylated, those located in the cytoplasm were not. Cytoplasmic microtubules were also labile as assessed by mild cooling. In contrast to mouse oocytes, rat microtubule organizing centers (MTOCs) did not react with MPM-2 antibody by immunofluorescence despite the fact that this antibody reacts with several proteins as shown by immunoblot. However, cytoplasmic MTOCs in M II-arrested rat oocytes could be detected by their nucleating capacity in the presence of taxol, a drug that induced the formation of numerous cytoplasmic asters. In addition, taxol caused a change in the spindle shape and the formation of astral microtubules at the spindle poles. Meiotic spindles (as well as chromosomes devoid of microtubules after nocodazole-treatment) were overlaid by an actin-rich domain. Spontaneous abortive activation led to the extrusion of the second polar body followed by another metaphase arrest--metaphase III; however, normal spindles did not form and dispersed chromosomes surrounded by microtubules were observed. Electron microscopic studies confirmed these observations and revealed that the kinetochores, are located deep within the chromosomes in contrast to mouse kinetochores, and this might be responsible for the absence of a metaphase III spindle in the rat oocyte. Induced activation caused transition to interphase with the formation of a characteristic microtubule network. This study shows that there are several significant differences in the cytoskeletal organization of rat and mouse oocytes.