The analysis of the potassium-independent chromatin cleavage patterns in oocytes proved intriguing. Although potassium apparently did not completely prevent DNA degradation, it did alter its pattern of electrophoretic movement as assessed by the comet assay. On the basis of the results of the ISEL analysis, we conclude that potassium prevents activation and/or activity of the endonuclease or endonucleases responsible for final degradation of oocyte chromatin to low-molecular weight fragments (as revealed by the absence of a plume of DNA outside of the oocyte when assessed by the comet assay) but that higher order DNA cleavage still occurs (as assessed by ISEL). The latter would only permit accumulation of the cleaved high-molecular weight chromatin against the oocyte plasma membrane.
This proposal is supported by our findings from analysis of DNA cleavage in granulosa cells of incubated rat ovarian follicles. In these experiments, internucleosomal, but not high-molecular weight, DNA cleavage was inhibited by KCl, NaCl, and LiCl. Furthermore, the occurrence of high-molecular DNA breaks in cells of KCl-treated follicles was sufficient for nuclear pyknosis to proceed. These findings are similar to those obtained in previous studies of rat ovarian follicles cultured with SAM, in which morphological criteria of apoptosis (e.g., nuclear pyknosis) were preserved in granulosa cells because of high-molecular weight DNA degradation, despite the complete absence of internucleosomal DNA cleavage.