Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography

In this study, we have used time-lapse video cinematography to studyfertilization in 50 human oocytes that had undergone intracytoplasmic sperminjection (ICSI). Time-lapse recording commenced shortly after ICSI andproceeded for 17-20 h. Oocytes were cultured in an environmental chamberwhich was maintained under standard culture conditions. Overall, 38 oocytes(76%) were fertilized normally, and the fertilization rate and embryoquality were not significantly different from 487 sibling oocytes culturedin a conventional incubator. Normal fertilization followed a defined courseof events, although the timing of these events varied markedly betweenoocytes. In 35 of the 38 fertilized oocytes (92%), there were circularwaves of granulation within the ooplasm which had a periodicity of 20-53min. The sperm head decondensed during this granulation phase. The secondpolar body was then extruded, and this was followed by the centralformation of the male pronucleus. The female pronucleus formed in thecytoplasm adjacent to the second polar body at the same time as, orslightly after, the male pronucleus, and was subsequently drawn towards themale pronucleus until the two abutted. Both pronuclei then increased insize, the nucleoli moved around within the pronuclei and some nucleolicoalesced. During pronuclear growth, the organelles contracted from thecortex towards the centre of the oocyte, leaving a clear cortical zone. Theoocyte decreased in diameter from 112 to 106 microm (P < 0.0001) duringthe course of the observation period. The female pronucleus wassignificantly smaller in diameter than the male pronucleus (24.1 and 22.4microm respectively, P = 0.008) and contained fewer nucleoli (4.2 and 7.0respectively, P < 0.0001). After time-lapse recording, oocytes werecultured for 48 h prior to embryo transfer or cryopreservation. Embryoquality was related to fertilization events and periodicity of thecytoplasmic wave, and it was found that good quality embryos arose fromoocytes that had more uniform timing from injection to pronuclear abuttaland tended to have a longer cytoplasmic wave. In conclusion, we have shownthat time-lapse video cinematography is an excellent tool for studyingfertilization and early embryo development, and have demonstrated thathuman fertilization comprises numerous complex dynamic events.