Chronic scrotal heat stress induces dose-dependent collapse of male sexual behavior in mice

Phuc Dang-Ngoc; Tam Le-Minh; Tung Nguyen-Thanh

doi:10.31893/jabb.2026006

Keywords:

Abstract

Scrotal hyperthermia is known to impair male reproductive physiology. However its dose- and time-dependent effects on copulatory behaviors have not been fully characterized. This study aimed to systematically quantify the longitudinal effects of chronic scrotal heat stress at moderate (37°C) and severe (40°C) temperatures on the sexual behavior of male mice and correlate these behaviors with testicular mass. Male ICR mice were randomized into control, moderate-heat (H37), and severe-heat (H40) groups. Copulatory behaviors, including mounting, intromission, and ejaculation parameters, were assessed weekly for seven weeks. Testicular and body weights were measured at the conclusion of the study. Control mice maintained stable sexual behavior throughout the study. In the H37 group, progressive impairment was observed starting at week three; with increasing mount, intromission, and ejaculation latencies and decreasing, whereas the frequencies of these behaviors correspondingly decreased over the seven-week period. The H40 group exhibited more severe and rapid deterioration. Behavioral latency increased sharply through week five before all copulatory activities,—including mounting, intromission, and ejaculation,—ceased entirely from week six onward. These behavioral deficits were linked to a significant, dose-dependent reduction in testicular mass. Chronic scrotal heat stress impairs male sexual performance in a dose- and time-dependent manner, ranging from gradual functional decline to complete behavioral collapse. This study established a robust murine model with sensitive behavioral endpoints that is useful for investigating the pathophysiology of heat-induced reproductive dysfunction and for evaluating potential therapeutics.
References
1. Aldahhan, R. A., & Stanton, P. G. (2021). Heat stress response of somatic cells in the testis. Molecular and Cellular Endocrinology, 527, 111216.
2. Bui-Le, T. N., & Hoang-Tan, Q. (2023). Protective effect of Curculigo orchioides Gaertn. extract on heat stress-induced spermatogenesis complications in murine model. Zhongguo Zhong Yao Za Zhi, 45(4), 3255–3267.
3. Cai, H., Qin, D., & Peng, S. (2021). Responses and coping methods of different testicular cell types to heat stress: Overview and perspectives. Zhonghua Nan Ke Xue, 41(6), 564–569.
4. Durairajanayagam, D., Agarwal, A., & Ong, C. (2015). Causes, effects and molecular mechanisms of testicular heat stress. Reproductive BioMedicine Online, 30(1), 14–27.
5. Dutta, S., Majzoub, A., & Agarwal, A. (2019). Oxidative stress and sperm function: A systematic review on evaluation and management. Arab Journal of Urology, 17(2), 87–97.
6. Feige-Diller, J., Krakenberg, V., Bierbaum, L., Seifert, L., Palme, R., Kaiser, S., Sachser, N., & Richter, S. H. (2020). The effects of different feeding routines on welfare in laboratory mice. Frontiers in Veterinary Science, 6, 479. https://doi.org/10.3389/fvets.2019.00479
7. Hafez Hafez, M., El-Sayed El-Kazaz, S., El-Neweshy, M. S., Shukry, M., Ghamry, H. I., & Tohamy, H. G. (2025). Resveratrol mitigates heat stress-induced testicular injury in rats: enhancing male fertility via antioxidant, antiapoptotic, pro-proliferative, and anti-inflammatory mechanisms. Naunyn-Schmiedeberg’s Archives of Pharmacology, 398(7), 8359–8373. https://doi.org/10.1007/s00210-024-03759-4
8. Hamilton, T. R. S., Mendes, C. M., Castro, L. S., Assis, P. M., Perez Siqueira, A. F., Delgado, J. C., Goissis, M. D., Muiño-Blanco, T., Cebrián-Pérez, J. Á., Visintin, J. A., & Assumpção, M. E. O. D’A. (2016). Evaluation of lasting effects of heat stress on sperm profile and oxidative status of ram semen and epididymal sperm. Oxidative Medicine and Cellular Longevity, 2016, 1687657. https://doi.org/10.1155/2016/1687657
9. Hansen, P. J. (2009). Effects of heat stress on mammalian reproduction. Philosophical Transactions of the Royal Society B: Biological Sciences, 364(1534), 3341–3350.
10. Hoang-Thi, A. P., Dang-Thi, A. T., Phan-Van, S., Nguyen-Ba, T., Truong-Thi, P. L., Le-Minh, T., Nguyen-Vu, Q. H., & Nguyen-Thanh, T. (2022). The impact of high ambient temperature on human sperm parameters: a meta-analysis. Iranian Journal of Public Health, 51(4), 710–723. https://doi.org/10.18502/ijph.v51i4.9232
11. Houston, B. J., Nixon, B., King, B. V., & Aitken, R. J. (2018). Heat exposure induces oxidative stress and DNA damage in the male germ line. Biology of Reproduction, 98(4), 593–606.
12. Khan, I., & Mesalam, A. (2023). Heat stress as a barrier to successful reproduction and potential alleviation strategies in cattle. Animals, 13(14), 2145.
13. Kim, B., Park, K., & Rhee, K. (2013). Heat stress response of male germ cells. Cellular and Molecular Life Sciences, 70(15), 2623–2636.
14. Lin, P. H., Chen, Y. K., Zhao, J. H., & Wang, Z. (2021). Exertional heat stroke on fertility, erectile function, and testicular morphology in male rats. Scientific Reports, 11(1), 3539.
15. Liu, Z.-W., Wang, L., Wang, Z.-W., & Zhang, X.-X. (2020). Assessment of sexual behavior of male mice. Journal of Visualized Experiments, 157, e60154.
16. Luengo-Mateos, M., Borràs, M., & Esteve-Codina, A. (2024). Protocol for ovariectomy and estradiol replacement in mice. STAR Protocols, 5(1), 102910.
17. Nguyen-Thanh, T., Pham, N. T., Nguyen, T. H., & Bui, H. T. (2022). Chronic scrotal heat stress causes testicular interstitial inflammation and fibrosis: An experimental study in mice. International Journal of Reproductive Biomedicine, 20(7), 569–580.
18. Ogawa, S., Lubahn, D. B., Korach, K. S., & Pfaff, D. W. (2000). Abolition of male sexual behaviors in mice lacking estrogen receptors alpha and beta (αβ ERKO). Proceedings of the National Academy of Sciences of the United States of America, 97(26), 14737–14741.
19. Paul, C., Teng, S., & Saunders, P. T. (2009). A single, mild, transient scrotal heat stress causes hypoxia and oxidative stress in mouse testes, which induces germ cell death. Biology of Reproduction, 80(5), 913–919.
20. Phumsatitpong, C., Wagenmaker, E. R., & Moenter, S. M. (2021). Neuroendocrine interactions of the stress and reproductive axes. Frontiers in Neuroendocrinology, 63, 100928.
21. Plant, T. M. (2015). 60 years of neuroendocrinology: The hypothalamo-pituitary-gonadal axis. Journal of Endocrinology, 226(2), T41–T54.
22. Rhodes, G. J. (2017). Surgical preparation of rats and mice for intravital microscopic imaging of abdominal organs. Methods, 128, 129–138.
23. Robinson, B. R., Netherton, J. K., Ogle, R. A., & Baker, M. A. (2023). Testicular heat stress: A historical perspective and two postulates for why male germ cells are heat sensitive. Biological Reviews, 98(2), 603–622. https://doi.org/10.1111/brv.12921
24. Xiong, Y., Li, J., & He, S. (2022). Zinc protects against heat stress-induced apoptosis via the inhibition of endoplasmic reticulum stress in TM3 Leydig cells. Biological Trace Element Research, 200(2), 728–739.
25. Zhang, M. H., Zhang, X., Wang, M., Li, J., Xu, Y., & Wu, B. (2015). Scrotal heat stress causes sperm chromatin damage and cysteinyl aspartate-specific proteinases 3 changes in fertile men. Journal of Assisted Reproduction and Genetics, 32(5), 747–755.
26. Zhang, P., Zheng, Y., Lv, Y., Li, F., Su, L., Qin, Y., & Zeng, W. (2020). Melatonin protects the mouse testis against heat-induced damage. Molecular Human Reproduction, 26(2), 65–79. https://doi.org/10.1093/molehr/gaaa002
27. Zhao, Z., Cai, Y., Lin, X., Liu, N., Qin, Y., & Wu, Y. (2024). The role of heat-induced stress granules in the blood-testis barrier of mice. International Journal of Molecular Sciences, 25(7), 3637.

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

How to cite

Dang-Ngoc, P., Le-Minh, T., & Nguyen-Thanh, T. (2026). Chronic scrotal heat stress induces dose-dependent collapse of male sexual behavior in mice. Journal of Animal Behaviour and Biometeorology, 14(1), 2026006. https://doi.org/10.31893/jabb.2026006

[1] Aldahhan, R. A., & Stanton, P. G. (2021). Heat stress response of somatic cells in the testis. Molecular and Cellular Endocrinology, 527, 111216.

[2] Bui-Le, T. N., & Hoang-Tan, Q. (2023). Protective effect of Curculigo orchioides Gaertn. extract on heat stress-induced spermatogenesis complications in murine model. Zhongguo Zhong Yao Za Zhi, 45(4), 3255–3267.

[3] Cai, H., Qin, D., & Peng, S. (2021). Responses and coping methods of different testicular cell types to heat stress: Overview and perspectives. Zhonghua Nan Ke Xue, 41(6), 564–569.

[4] Durairajanayagam, D., Agarwal, A., & Ong, C. (2015). Causes, effects and molecular mechanisms of testicular heat stress. Reproductive BioMedicine Online, 30(1), 14–27.

[5] Dutta, S., Majzoub, A., & Agarwal, A. (2019). Oxidative stress and sperm function: A systematic review on evaluation and management. Arab Journal of Urology, 17(2), 87–97.

[6] Feige-Diller, J., Krakenberg, V., Bierbaum, L., Seifert, L., Palme, R., Kaiser, S., Sachser, N., & Richter, S. H. (2020). The effects of different feeding routines on welfare in laboratory mice. Frontiers in Veterinary Science, 6, 479. https://doi.org/10.3389/fvets.2019.00479

[7] Hafez Hafez, M., El-Sayed El-Kazaz, S., El-Neweshy, M. S., Shukry, M., Ghamry, H. I., & Tohamy, H. G. (2025). Resveratrol mitigates heat stress-induced testicular injury in rats: enhancing male fertility via antioxidant, antiapoptotic, pro-proliferative, and anti-inflammatory mechanisms. Naunyn-Schmiedeberg’s Archives of Pharmacology, 398(7), 8359–8373. https://doi.org/10.1007/s00210-024-03759-4

[8] Hamilton, T. R. S., Mendes, C. M., Castro, L. S., Assis, P. M., Perez Siqueira, A. F., Delgado, J. C., Goissis, M. D., Muiño-Blanco, T., Cebrián-Pérez, J. Á., Visintin, J. A., & Assumpção, M. E. O. D’A. (2016). Evaluation of lasting effects of heat stress on sperm profile and oxidative status of ram semen and epididymal sperm. Oxidative Medicine and Cellular Longevity, 2016, 1687657. https://doi.org/10.1155/2016/1687657

[9] Hansen, P. J. (2009). Effects of heat stress on mammalian reproduction. Philosophical Transactions of the Royal Society B: Biological Sciences, 364(1534), 3341–3350.

[10] Hoang-Thi, A. P., Dang-Thi, A. T., Phan-Van, S., Nguyen-Ba, T., Truong-Thi, P. L., Le-Minh, T., Nguyen-Vu, Q. H., & Nguyen-Thanh, T. (2022). The impact of high ambient temperature on human sperm parameters: a meta-analysis. Iranian Journal of Public Health, 51(4), 710–723. https://doi.org/10.18502/ijph.v51i4.9232

[11] Houston, B. J., Nixon, B., King, B. V., & Aitken, R. J. (2018). Heat exposure induces oxidative stress and DNA damage in the male germ line. Biology of Reproduction, 98(4), 593–606.

[12] Khan, I., & Mesalam, A. (2023). Heat stress as a barrier to successful reproduction and potential alleviation strategies in cattle. Animals, 13(14), 2145.

[13] Kim, B., Park, K., & Rhee, K. (2013). Heat stress response of male germ cells. Cellular and Molecular Life Sciences, 70(15), 2623–2636.

[14] Lin, P. H., Chen, Y. K., Zhao, J. H., & Wang, Z. (2021). Exertional heat stroke on fertility, erectile function, and testicular morphology in male rats. Scientific Reports, 11(1), 3539.

[15] Liu, Z.-W., Wang, L., Wang, Z.-W., & Zhang, X.-X. (2020). Assessment of sexual behavior of male mice. Journal of Visualized Experiments, 157, e60154.

[16] Luengo-Mateos, M., Borràs, M., & Esteve-Codina, A. (2024). Protocol for ovariectomy and estradiol replacement in mice. STAR Protocols, 5(1), 102910.

[17] Nguyen-Thanh, T., Pham, N. T., Nguyen, T. H., & Bui, H. T. (2022). Chronic scrotal heat stress causes testicular interstitial inflammation and fibrosis: An experimental study in mice. International Journal of Reproductive Biomedicine, 20(7), 569–580.

[18] Ogawa, S., Lubahn, D. B., Korach, K. S., & Pfaff, D. W. (2000). Abolition of male sexual behaviors in mice lacking estrogen receptors alpha and beta (αβ ERKO). Proceedings of the National Academy of Sciences of the United States of America, 97(26), 14737–14741.

[19] Paul, C., Teng, S., & Saunders, P. T. (2009). A single, mild, transient scrotal heat stress causes hypoxia and oxidative stress in mouse testes, which induces germ cell death. Biology of Reproduction, 80(5), 913–919.

[20] Phumsatitpong, C., Wagenmaker, E. R., & Moenter, S. M. (2021). Neuroendocrine interactions of the stress and reproductive axes. Frontiers in Neuroendocrinology, 63, 100928.

[21] Plant, T. M. (2015). 60 years of neuroendocrinology: The hypothalamo-pituitary-gonadal axis. Journal of Endocrinology, 226(2), T41–T54.

[22] Rhodes, G. J. (2017). Surgical preparation of rats and mice for intravital microscopic imaging of abdominal organs. Methods, 128, 129–138.

[23] Robinson, B. R., Netherton, J. K., Ogle, R. A., & Baker, M. A. (2023). Testicular heat stress: A historical perspective and two postulates for why male germ cells are heat sensitive. Biological Reviews, 98(2), 603–622. https://doi.org/10.1111/brv.12921

[24] Xiong, Y., Li, J., & He, S. (2022). Zinc protects against heat stress-induced apoptosis via the inhibition of endoplasmic reticulum stress in TM3 Leydig cells. Biological Trace Element Research, 200(2), 728–739.

[25] Zhang, M. H., Zhang, X., Wang, M., Li, J., Xu, Y., & Wu, B. (2015). Scrotal heat stress causes sperm chromatin damage and cysteinyl aspartate-specific proteinases 3 changes in fertile men. Journal of Assisted Reproduction and Genetics, 32(5), 747–755.

[26] Zhang, P., Zheng, Y., Lv, Y., Li, F., Su, L., Qin, Y., & Zeng, W. (2020). Melatonin protects the mouse testis against heat-induced damage. Molecular Human Reproduction, 26(2), 65–79. https://doi.org/10.1093/molehr/gaaa002

[27] Zhao, Z., Cai, Y., Lin, X., Liu, N., Qin, Y., & Wu, Y. (2024). The role of heat-induced stress granules in the blood-testis barrier of mice. International Journal of Molecular Sciences, 25(7), 3637.

Abstract

References

How to cite