Live and Time-Lapse Imaging of Early Oogenesis and Meiotic Chromosomal Dynamics in Cultured Juvenile Zebrafish Ovaries.

Oocyte production is crucial for sexual reproduction. Recent findings in zebrafish and other established model organisms emphasize that the early steps of oogenesis involve the coordination of simultaneous and tightly sequential processes across cellular compartments and between sister cells. To fully understand the mechanistic framework of these coordinated processes, cellular and morphological analysis in high temporal resolution is required. Here, we provide a protocol for four-dimensional live time-lapse analysis of cultured juvenile zebrafish ovaries. We describe how multiple-stage oocytes can be simultaneously analyzed in single ovaries, and several ovaries can be processed in single experiments. In addition, we detail adequate conditions for quantitative image acquisition. Finally, we demonstrate that using this protocol, we successfully capture rapid meiotic chromosomal movements in early prophase for the first time in zebrafish oocytes, in four dimensions and in vivo. Our protocol expands the use of the zebrafish as a model system to understand germ cell and ovarian development in postembryonic stages.