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Zinc (Zn) is a vital trace element for the body and its bioavailability influences numerous reproductive events. However, the mechanisms that regulate Zn homeostasis in the cumulus-oocyte complex (COC) are yet to be elucidated. The aim of this study was to investigate the role of estradiol 17-beta (E 2), FSH and LH in Zn homeostasis regulation in bovine COC matured in vitro and Zn transporters gene expression. For this purpose, intracellular Zn levels in oocytes and cumulus cells (CC) were assessed using a Zn-specific fluorescent indicator. In addition, gene expression and sequencing of six Zn transporters (Slc39a6, Slc39a8, Slc39a14, Slc30a3, Slc30a7 and Slc30a9) were assessed. Our results demonstrated that the simultaneous presence of E 2 , FSH, and LH during oocyte maturation altered intracellular zinc levels and transporters expression in both oocytes and CC. Transporter's gene expression was different in oocytes and CC, possibly due to cell-specific changes in Zn levels during maturation. The interaction effects of Zn with hormonal treatments influenced the results. This study emphasizes that Slc39a6 is highly sensitive to hormone induction. Overall, the hormonal modulation of Zn homeostasis in the COC was evidenced. Also, a preponderant role of FSH as a modulator of Zn intracellular levels and transporter gene expression is suggested. • Zinc availability is beneficial for follicular growth, oocyte maturation, fertilization, and embryo development. • Reproductive hormones affected Zn homeostasis and gene expression of Zn transporters during IVM of bovine COC. • The gene expression of Zn transporters in oocytes and cumulus cells were different. • The role of FSH in the modulation of intracellular Zn levels and transporter gene expression is evident. • The Zn transporter Slc39a6 is highly sensitive to hormonal regulation. [ABSTRACT FROM AUTHOR]