Novel morphological approaches for the study of oocyte maturation

Recent advances in light microscopy are discussed with respect to their application for the study of cell surface, cytoskeletal and organellar changes that occur during meiotic maturation in mammalian oocytes. Three techniques are considered: 1) multiple fluorochrome labeling using immunocytochemical or pharmacological probes to analyze the spatial and temporal disposition of components in oocytes fixed at various stages of meiosis, 2) the use of vital fluorescence stains to study the actual movement of organelles in living oocytes, and 3) video image intensification microscopy of living cells to record dynamic cellular changes with enhanced optical capabilities for fluorescence, polarization and differential interference contrast microscopy. A method is described for simultaneously localizing chromosomes, microtubules and f-actin in fixed rodent oocytes using, respectively, Hoechst 33258, antitubulin antibodies and NBD-phallicidin. Acridine orange and the laser dye rhodamine 123 are employed as vital stains to visualize lysosomes and mitochondria, respectively, in rat oocytes undergoing meiotic maturation. Finally, the application of time-lapse video image intensification microscopy for the study of fluorescently labeled cellular components is discussed with special reference to extended monitoring of cellular organelles for the analysis of dynamic movements of oocyte constituents during maturation.