Date of Award
Leibo, Stanley P.
Reproductive science plays an important role in conservation biology. Quantitative studies of basic reproductive biology in wildlife are critical for the development of successful assisted reproductive technologies. Investigation of the volume regulatory mechanism of spermatozoa could produce options to improve the cryopreservation of spermatozoa and provide a non-hormonal contraceptive option for men, both of which could have significant impacts on global biodiversity preservation. Volume regulation of somatic cells involves the movement of osmolytes through various channels, including potassium channels. The potassium channels involved in volume regulation of human, monkey, and murine spermatozoa were investigated. Flow cytometry was used to gauge the sensitivity of the volume regulatory process of spermatozoa to various potassium channel inhibitors and a simultaneous hypotonic challenge. Channels potentially involved in regulatory volume decrease of spermatozoa varied with species but included voltage-gated (Kv) channels 1.4, 1.5, 1.7, 4.1, 4.2 and 4.3 as well as TWIK1, TWIK2, TASK1, TASK2, TASK3, TREK2 , and minK. The presence of some of these channels was confirmed by western blotting and immunocytochemistry. Changes in the motility patterns of human and monkey spermatozoa in the presence of potassium channel inhibitors during hypotonic stress were also observed, suggesting a relationship between volume regulation and motility. To evaluate potential organic osmolytes involved in, and compare effects of CPAs on, volume regulation, the isotonicity of murine epididymal spermatozoa was measured using a null point method. Spermatozoa were then exposed to high concentrations of various osmolytes and cryoprotective agents in isotonic medium to evaluate which compounds were able to penetrate the sperm plasma membrane. The osmotic responses of spermatozoa from strains of mice known to have spermatozoa of high (B6D2F1) and low (C57BL6) post-thaw fertility were compared during various osmotic challenges in various media. These experiments indicated that spermatozoa from B6D2F1 mice may have better volume regulation capabilities than spermatozoa from C57BL6 mice, suggesting that better post-thaw fertility of murine spermatozoa could be influenced by the volume regulatory process. The knowledge gained from these experiments could contribute to improved sperm handling and preservation techniques and be used to develop non-hormonal male contraceptives based on inhibiting volume regulation.
Barfield, Jennifer, "Regulation of Volume by Spermatozoa and Its Significance for Conservation Biology" (2007). University of New Orleans Theses and Dissertations. 581.