Osmotic shock induces structural damage on equine spermatozoa plasmalemma and mitochondria

The present study aimed to elucidate the effects that osmotic shock exerts on equine spermatozoa. To achieve this goal, a retrospective study of the cellular volume of 40 equine ejaculates subjected to osmolarities ranging from 75 to 900 mOsm in Biggers-Whitten-Whittingham (BWW) media was performed using a Multisizer3 Coulter Counter®. The 300 mOsm BWW solution was used as control. The sperm volume ranged between 37.93 ± 0.6 (mean ± Standard Error of the Mean (SEM)) in 75 mOsm BWW to 21.61 ± 0.27 (mean ± SEM) for 900 mOsm BWW. Thus the spermatozoa behaved as linear osmometers when adjusted to the Boyle Van't Hoff equation (R2 = 0.9808). After the different osmotic challenges, spermatozoa were returned to 300 mOsm BWW and the cellular volume was measured again. The results showed that the spermatozoa were able to retrieve the isosmolar volume (20.81 ± 0.34; mean ± SEM). Also, an ultrastructural study of spermatozoa membrane and mitochondria was accomplished using Transmission Electron Microscopy (TEM) after the osmotic challenges in 2 ejaculates. As observed by TEM, sperm plasmalemma swelled and detached from the sperm head in hypotonic conditions (75 mOsm), with blebbing on return to isosmolarity. When subjected to 900 mOsm, the sperm plasmalemma shrank, with disarrangement and blebbing when returned to isosmolarity. Mitochondria were also found to change their volume; the main pathologic change was irreversible vacuolization and changes in their arrangement for all the osmotic challenges tested. The present work leads to a better understanding of how osmotic shock adversely affects equine spermatozoa structure.