Monitoring of Sperm-Independent Calcium Oscillations in Immature Oocytes of Mice.

Repetitive increases of intracellular calcium ions (Ca2+ oscillations) control cellular functions in various biological events, including meiotic resumption after fertilization. Sperm-derived substances enter the cytoplasm of mature oocytes by sperm fusion, causing Ca2+ oscillations. Sperm-independent Ca2+ oscillations are also induced in immature oocytes isolated from the ovaries of neonatal to adult mice. The presence of Ca2+ oscillations may contribute to subsequent oocyte quality; however, its physiological role and molecular mechanism are unclear. Here, we describe a method of collecting immature oocytes from the ovaries of juvenile (12, 15, and 21 days after birth) and adult mice and monitoring their Ca2+ oscillations. Since mouse oocytes are larger than other types of cells, they are a useful model for studying spatiotemporal patterns and the mechanism of Ca2+ oscillations in various types of cells. This method can be applied to other rodents due to similarities in oocyte size and developmental processes. Furthermore, the use of various fluorescent probes enables visualization of organelle rearrangement. The mechanism of interaction between oocytes and somatic cells differs between juvenile and adult mice. Therefore, two distinct methods are employed for oocyte collection. Key features • Isolation of immature oocytes from juvenile ovaries [use of ethylenediaminetetraacetic acid (EDTA)]. • Isolation of immature oocytes from adult ovaries (no treatment with protease and EDTA). • Monitoring of Ca2+ oscillations in immature oocytes.