Culture of the Intact Postnatal Naked Mole-Rat Ovary: From Meiotic Prophase to Single-Cell RNASeq.

ABSTRACT

Recently, we reported that, in the naked mole-rat (Heterocephalus glaber) ovary, there is mitotic expansion of the primordial germ cells (PGCs), and the initiation of the meiotic program occurs postnatally. This is opposite to almost all other mammals, including humans and mice, whose reproductive cycle begins very early in development. In both mouse and human, the ovaries become populated with PGCs in utero; these PGCs will later generate the oogonia. After mitotic proliferation, these cells will trigger the meiotic program and initiate meiotic prophase I. Given that all these processes happen in utero, their analysis has been very challenging; so the ability to study them postnatally and to manipulate them with inhibitors or other substances, in the naked mole-rat, opens new possibilities in the field. In this chapter, we present a comprehensive collection of protocols that permit the culture of whole naked mole-rat ovaries, followed by analysis of germ cells, from PGCs to oocytes, in meiotic prophase I, as well the obtention of single-cell suspension or single-nuclei suspension for RNASeq.