During the reproductive lifespan of a female, only a limited quantity of oocytes are naturally ovulated; therefore, the mammalian ovary possesses a substantial population of preantral follicles available to be handled and explored in vitro. Hence, the manipulation of preantral follicles enclosed in ovarian tissue aims to recover a considerable population of oocytes of high-value animals for potential application in profitable assisted reproductive technologies (ARTs). For this purpose, the technique of preantral follicle in vitro culture (IVC) has been the most common research tool, achieving extraordinary results with offspring production in the mouse model. Although promising outcomes have been generated in livestock animals after IVC of preantral follicles, the quantity and quality of embryo production with those oocytes are still poor. In recent years, the mare has become an additional model for IVC studies due to remarkable similarities with women and livestock animals regarding in vivo and in vitro ovarian folliculogenesis. For a successful IVC system, several factors should be carefully considered to provide an optimum culture environment able to support the viability and growth of preantral follicles enclosed in ovarian tissue. The cryopreservation of the ovarian tissue is another important in vitro manipulation technique that has been used to preserve the reproductive potential in humans and, in the future, may be used in highly valuable domestic animals or endangered species. Several improvements in cryopreservation protocols are necessary to support the utilization of ovarian tissue of different species in follow-up ARTs (e.g., ovarian fragment transplantation). This review aims to provide an update on the most current advances regarding supportive in vitro techniques used in equids to evaluate and manipulate preantral follicles and ovarian tissue, as well as methodological approaches used during IVC and cryopreservation techniques.