MICROSCOPIC ANALYSIS OF ENZYME-ACTIVITY, MITOCHONDRIAL DISTRIBUTION AND HYDROGEN-PEROXIDE IN 2-CELL RAT EMBRYOS

A developmental block is induced by phosphate in rat embryos at the la te two-cell stage. The present study was designed to examine the energ y metabolism of rat two-cell blocked and non-blocked embryos. Enzyme a ctivity was measured in individual embryos by histochemical techniques . The activities of malate dehydrogenase, isocitrate dehydrogenase, la ctate dehydrogenase, pyruvate dehydrogenase, glyceraldehyde-3-phosphat e dehydrogenase, glutamate dehydrogenase, glucose-6-phosphate dehydrog enase, glucose-6-phosphatase, and phosphorylase did not differ among n on-blocked and blocked embryos. However, the activity of succinate deh ydrogenase was significantly decreased in blocked embryos compared wit h non-blocked embryos. In blocked embryos, cytochrome oxidase activity was distributed homogeneously, but was located at the perinuclear reg ion in non-blocked embryos. Active mitochondrial organization was visu alized using the fluorescent probe rhodamine 123 and laser scanning co nfocal microscopy. In both non-blocked and blocked embryos, mitochondr ia were distributed homogeneously. The concentration of H2O2 measured fluorometrically in embryos cultured without phosphate did not change significantly during the culture period, but decreased in embryos cult ured with phosphate. The timing corresponded to the occurrence of the two-cell block. In summary, these results suggest that the development al block in rat two-cell embryos is induced by disturbance of mitochon drial energy metabolism.