| Meiotic maturation and fertilization of mammalian oocytes is an important and complex process, the studies of oocytes parthenogenetic activation and the dynamic changes of cytoskeleton during oocytes activation will make us get a deeper understanding of the mechanism of this process. Recent years, there are some reports about the cytoskeletal distribution and roles of parthenotes, but the dynamic changes of cytoskeleton in the parthenotes activated by various stimuli are still a problem lacking systematic studies. To date, there are only a few internal reports about this field. This experiment aimed to systematically investigate and analyze the ethanol activated rates of mouse oocytes, affected factors of oocytes activation, types and development of parthenotes, and the organization and roles of the cytoskeleton during pronuclear formation and migration and early embryo cleavage by ethanol activation of oocytes, embryo culture in vitro and immunofluorescence cyto-chemistry. The results: (1) The ethanol activation of mouse oocytes showed that the activated and developmental rates of oocytes increased significantly with the increase of ethanol concentration and extension of exposure time, but over concentration and exposure time would result in increased fragment rates significantly. 7% ethanol treated oocytes for 7min was the optimum activated condition. With the increases of oocyte ages and temperature, the oocytes activation and development rates increased, while fragment rates increased significantly. Ovarian cumulus had no obvious effect on the activation of oocytes in vitro. Oocytes of sexual maturity had better activated effect than under sexual maturity mouse. (2) Degraded proportion of oocytes first polar body (PB1) and numbers of oocytes without PB1 both increased with the increases of oocyte ages after hCG injection. With or without PB1 had no significant effects on the activation, development and fragment rates. We also observed that oocytes with or without PB1 had the same distribution patterns of cytoskeleton. (3) Our fluorescence observation results indicated that microtubules were restricted to the region of the metaphase II (MII) meiotic spindle. With the extrusion of the second polar body (PB2), MII spindle began to disintegrate and disappear slowly, and the compact microtubule-containing structure known as the midbody was formed. The midbody located in the contact surfaces of cells that divided, and finally disappeared after cell division completed. Pronuclei migrated to the center of cytoplasm after their formation at the edge. (4) Microfilaments concentrated to the oocyte cortex and polar body at the different developmental stages of eggs. Microfilaments were mainly distributed in the cortex area which chromosomes located at mataphase of second meiosis (MII)and concentrated around pronuclei after pronuclear formation. After extrusion of polar body and the completion of the first cell cycle, microfilaments concentrated to the cleavage furrows. (5) The MII spindle showed abnormal assembly, and multiple microtubule asters formed in the cytoplasm, chromosomes were not being normal aligned and couldn't separated each other after treating the oocytes with taxol. While washed taxol completely, the activated oocytes could resume meiosis II, the MII spindle reorganized, the oocytes extruded PB2 and formed pronuclei. Our results showed that the parthenogenetic activation of mouse oocytes was affected by many factors, so the selection of optimum activated conditions was very important for the effect of activation. The organization and structure of cytoskeleton changed obviously during the resumption of meiotic and parthenogenetic activation of mammalian oocytes. Microtubules and microfilaments played important regulated roles in many events such as the resumption of meiotic, chromosomal movement, the formation, location and rotation of meiotic spindle, polar body extrusion, pronuclear formation and migration and also early embryo cleavage.
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