Abstract Cryptorchidism is associated with an increased risk of male infertility and testicular cancer. Persistent exposure to high temperature in cryptorchidism can lead to the apoptosis of spermatogenic cells. Transient receptor potential vanilloid 1 (TRPV1), a thermosensitive cation channel, has been found to have differential effects on various apoptosis processes. However, whether TRPV1 is involved in spermatogenic cell apoptosis induced by cryptorchidism remains unclear. Herein, we first observed the expression pattern of TRPV1 in the testes of mice with experimental cryptorchidism, and then investigated the role and mechanism of TRPV1 in spermatogenic cell apoptosis by using Trpv1 −/− mice. The results showed that TRPV1 was highly expressed on the membrane of spermatocytes in mouse testis, and the expression increased significantly in the testis of mice with experimental cryptorchidism. After the operation, Trpv1 −/− mice exhibited less reproductive damage and fewer spermatogenic cell apoptosis compared to the wild-type (WT) mice. Transcriptome sequencing revealed that the expression of apoptosis-related genes (Capn1, Capn2, Bax, Aifm1, Caspase 3, Map3k5, Itpr1 and Fas) was down-regulated in spermatocytes of cryptorchid Trpv1 −/− mice. Our results suggest that TRPV1 promotes the apoptosis of spermatocytes in cryptorchid mice by regulating the expression of apoptosis-related genes.