Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy.

Abstract

The role of mitochondrial metabolism in the development of preimplantation embryos is unclear. To clarify the importance of mitochondria in early development, the fluorescent probes rhodamine 123 (Rh123: stains active mitochondrial membrane) and nonyl acridine orange (NAO: stains active and inactive mitochondrial membrane) were used with confocal laser scanning microscopy to study the distribution of mitochondria in hamster unfertilized follicular and oviductal eggs (11.5 hr and 16 hr post-hCG, respectively) and preimplantation embryos (1-cell to blastocyst). Rh123 staining indicated that active mitochondria were homogeneously distributed in unfertilized follicular and oviductal eggs. At 3 hr post egg activation (PEA) by sperm, active mitochondria were still found throughout the cytoplasm of the activated egg although they were slightly clustered around the pronuclei and were intensely active in the second polar body. During the next 9 hr, the majority of active mitochondria encircled the apposing pronuclei. By this time the second polar body no longer stained. In 2-cell, 4-cell and 8-cell embryos, there was a striking constancy in the pattern of active mitochondria which were clustered around the nuclei and delineating the cytocortex subjacent to the plasma membrane. In the blastocyst, active mitochondria were most readily detected in the trophectoderm cells in a homogeneous distribution. Staining mitochondria with NAO showed the same distribution patterns as Rh123, indicating that perinuclear clustering of active mitochondria involves the physical movement of these organelles rather than simply changes in their activity. Distribution of actin microfilaments and microtubules showed similar patterns to mitochondria and may be involved in their movement. This migration of mitochondria, beginning during the early stages of fertilization in the hamster egg and persisting until blastocoel formation, must have some functional correlation with successful preimplantation development.