Translocation of active mitochondria during pig oocyte maturation, fertilization and early embryo development in vitro

The distribution of active mitochondria during pig oocyte maturation, ferti lization and early embryo development in vitro was revealed by using MitoTr acker Green staining and confocal laser scanning microscopy. The regulation of mitochondrial translocation by microfilaments and microtubules was also studied. In oocytes collected from small follicles, strong staining of act ive mitochondria was observed in the cell cortex. Accumulation of active mi tochondria in the peripheral cytoplasm and around the germinal vesicles was characteristic of fully grown oocytes collected from large follicles. Mito chondria accumulated in the perinuclear area during meiotic progression fro m germinal vesicle breakdown (GVBD) to anaphase I. Larger mitochondrial foc i were formed and moved to the inner cytoplasm in mature oocytes. Compared with the oocytes matured in vivo, in which large mitochondrial foci were di stributed throughout the cytoplasm, mitochondria were not observed in the c entral cytoplasm in most of the oocytes matured in vitro. Strong staining o f mitochondria was observed in the first polar bodies in metaphase II oocyt es. In fertilized eggs, active mitochondria aggregated in the pronuclear re gion. Perinuclear clustering and a cortical ring were the most marked featu res of early cleavage. Active mitochondria were distributed in both inner c ell mass cells and trophectoderm cells of the blastocysts. Disassembly of m icrotubules with nocodazole inhibited both mitochondrial aggregations to th e germinal vesicle area and their inward movement to the inner cytoplasm du ring oocyte maturation, as well as the translocation of mitochondria to the peri-pronuclear region during fertilization, whereas disruption of microfi laments by cytochalasin B had no effects. These data indicate that: (i) ooc yte maturation, fertilization and early embryo development in pigs are asso ciated with changes in active mitochondrial distribution; (ii) mitochondria l translocation is mediated by microtubules, but not by microfilaments; and (iii) in vitro maturation conditions may cause incomplete movement of mito chondria to the inner cytoplasm and thus affect cytoplasmic maturation.