There is an evident relationship between the fertilizing capacity of sperm and the normal morphology, quality chromatin, and motility of sperm. It is well known that thyroid hormones are the important regulators of testicular function. A correlation was found between the hypothyroidism and sperm damages. The present study was conducted to investigate the effects of hypothyroidism on sperm morphology, chromatin quality, and motility. For this purpose, 20 male mice were divided into the control and the hypothyroid groups that received 0.05% 6-n-propyl-2-thiouracil (PTU) for 35 days. Sperm morphology with Papanicolaou staining and sperm chromatin quality with both Aniline Blue (AB) and Toluidine blue (TB) staining were assessed. Besides, immunohistochemistry and real-time PCR were performed to evaluate the changes of cation sperm channel (CatSper) genes. A significant increase in the sperm chromatin condensation was found in the hypothyroid mice compared to the control mice (p < 0.05). Furthermore, a significant decrease was observed in the morphology of normal sperm in hypothyroid mice compared to the controls (p < 0.05). The results showed that Hypothyroidism could downregulate the expression of CatSper genes. Immunohistochemical data confirmed the real time-PCR results. Furthermore, the results showed that hypothyroidism could adversely affect sperm morphology, sperm chromatin condensation, and CatSper gene expression in mice and these abnormalities may be related to the excessive production of reactive oxygen species (ROS) in a hypothyroid state.