Vitrification of germinal vesicle (GV) stage oocytes has been shown to be closely associated with decreased rates of meiosis maturation and increased rates of aneuploidy. However, little is known about the effects of melatonin on these events in mice vitrified GV oocytes. In this study, the effects of melatonin on meiosis maturation potential and the incidence rate of aneuploidy in mouse vitrified oocytes were analyzed by supplementing in vitro maturation (IVM) solution with melatonin at different concentrations. This study, for the first time, showed that the mitochondrial heat production was markedly increased in vitrified oocytes (P < 0.05), which compromised the first polar body extrusion (PBE) of vitrified oocytes (73.3% vs. 85.1%, P < 0.05). However, 10−11 mol/L melatonin could significantly decrease mitochondrial heat production and ROS level (9.1 vs. 12.0 pixels, P < 0.05), meanwhile increase ATP level (1.1 vs. 0.88 pmol, P < 0.05) and mtDNA copies (107438 vs. 67869, P < 0.05), which rescued the abnormal chromosome alignment (32% vs. 69%, P < 0.05) and reduced the incidence of aneuploidy (15.6% vs. 38.5%, P < 0.05) in vitrified oocytes. The meiosis maturation ability of vitrified oocytes with melatonin supplementation was similar to that of fresh ones (83.4% vs. 85.1%, P > 0.05). Collectively, our data revealed that melatonin has a protective action against vitrification-induced injuries of oocytes meiosis maturation. The mitochondrial in vitrified oocytes was prone to generate heat rather than ATP, which resulted in the higher rates of abnormal chromosome alignment and aneuploidy. The mitochondrial heat product in vitrified oocytes was associated with mitochondrial DNA copies and ROS level, and melatonin could significantly rescue the aneuploidy in vitrified oocytes by decreasing mitochondria product heat and increasing the ATP synthesis. Image 1 • Mitochondrial was prone to generate more heat rather than ATP in vitrified oocytes. • Melatonin rescue the aneuploidy in vitrified oocyte by decreasing mitochondria product heat and increasing the ATP synthesis. • The motichondrial heat product in vitrified oocytes was associated with mitochondrial DNA copies and ROS level. • Melatonin increase the mitochondrial DNA copies and decline the ROS level, which reduced the mitochondrial heat production. [ABSTRACT FROM AUTHOR]