Objective To explore the characteristics and molecular mechanisms of dynamic changes in ovarian function during natural aging in mice. Methods Eighteen female ICR mice of 2, 6, 10 and 14 months of age (referred to as 2M, 6M, 10M, and 14M) were included. The estrous cycle, ovarian index, pregnancy rate and embryo number were detected. The number of follicles was observed using HE staining. Serum anti-Müllerian hormone (AMH) levels were measured using ELISA. Protein and mRNA expression of P16, P21, proliferator-activated receptor-γ coactivator-1α (PGC-1α), and mitochondrial transcription factor A (TFAM) were detected through IHC and qPCR, respectively, and the protein expression of PGC-1α and TFAM was detected by Western blot. Relationships between indicators were evaluated based on Spearman’s rank correlation coefficients and logistic regression analyses. Results Compared with estimates in the 2M group, the percentage of estrous cycle disorders (P<0.05 in 10M group, P<0.01 in 14M group), ovarian index (P<0.05 in 10M group, P<0.01 in 14M group), serum AMH level (P<0.01 in 10M group, P<0.01 in 14M group), number of embryos (P<0.01 in the 10M group, P<0.01 in 14M group), and pregnancy rate (P<0.01 in 14M group) were significantly lower; the number of follicles at all levels and total number of follicles were lower (all P<0.01), and the number of atretic follicles (all P<0.01) was higher. Furthermore, P16 and P21 mRNA (P16: P<0.01 in 10M and 14M groups; P21: P<0.05 in 10M group, P<0.01 in 14M group) and protein levels (P16: P<0.01 in each group; P21: P<0.01 in 10M and 14 M groups) were elevated. Spearman’s correlation analyses showed that the age in months was negatively correlated with the ovarian index, serum AMH level, primordial follicle number, number of embryos, and expression of PGC-1α and TFAM and positively correlated with the expression of atretic follicle, P16 and P21 (all P<0.01). Logistic regression analysis revealed a significant negative correlation between the age in months and pregnancy rate (P<0.01). PGC-1α mRNA was significantly decreased (all P<0.01), and TFAM mRNA was significantly decreased in 10M and 14M groups (P<0.01). PGC-1α levels were significantly decreased in the 10M group (IHC: P<0.05; Western blot: P<0.01) and 14M group (all P<0.01); TFAM levels were significantly lower in 6M, 10M, and 14 M groups than in the 2M group (IHC: P<0.01; Western blot: P<0.01). Compared with the 2M group, expression levels of PGC-1α and TFAM were positively correlated with the ovarian index and serum AMH level and negatively correlated with P16 and P21 expression (all P<0.01). Conclusions Ovarian function in mice declines progressively with age in months, as reflected by reductions in follicles and fertility and the up-regulation of aging markers, which may be associated with the decreased expression of the mitochondrial biogenesis factors PGC-1α and TFAM.