Morbidity of aging non-incubated chicken blastoderms: further cytological evidence and interpretation.

Cytological analysis was carried out on the blastodermal cells of White Leghorn eggs subjected to several pre-incubation treatments. These treatments were storage, ranging from 0 days to 4 weeks, together with one or more of the following: (1) variation of ambient temperature (5 °C, 15-5 °C, 24 °C), (2) variation of carbon dioxide level (normal air, 1-5% CO2-enriched air), (3) increased humidity (wrapping in plastic bags). To facilitate analysis, the chromosomal configuration of metaphase cells was classified into four types, I-IV, according to their increasingly 'abnormal' appearance, which included condensation, dispersal and/or clumping of chromosomes. In interphase cells, the degree of 'abnormality' was rated on the staining capacity and shape of the nuclei. The study yielded the following results. (1) Temperature of storage was the most important single factor determining the state of 'normality' of the nuclei. The CO2 level in the storage chamber or the use of plastic bags (to provide the eggs in storage with a special' mini-environment', believed largely due to increased humidity) had little effect on the cytological picture of the affected blastoderms. (2) As the blastoderms of eggs stored at 15-5 °C aged, the proportion of Type IV chromosomal configurations steadily rose. At 24 °C the aging process frequently followed a different route: both metaphase and post-metaphase chromosomes often simply disintegrated; the progression from Type I to Type IV was not in evidence. Aging at 5 °C resulted in an early appearance of uniformly dark-stained, spherical or oval nuclei. These were similar to those observed in the terminal stages of retrogression seen in the interphase nuclei. (3) During extended storage at 15-5 °C, mitosis was shown to be blocked at metaphase. No active mitoses were observed in the blastoderms of eggs stored at 5 °C. At 24 °C however, limited mitotic activity was present, up to and including anaphase. (4) The presence or absence of a high level of mitotic activity during pre-incubation storage was not crucial to the survival of the blastoderm. However, an environment that permitted limited mitosis was important if the cells were to have the best possible chance for remaining alive during storage. The CO2 content of the air or the use of plastic bags played no role in this respect. (5) Two explanations, at the nuclear level, are suggested for the observed chain of events in the blastoderm of a stored chicken egg.