udy has been characterized by maintaining a relatively constant sperm head morphometry throughout the cryopreservation (response pattern 1). This would not imply the absence of changes in sperm shape during cryopreservation, but that changes could occur rapidly at each stage. Males showing this response pattern have been characterized by smaller sperm head dimensions in fresh semen and have showed an intermediate sperm quality at thawing. As we pointed out above, many authors have associated this sperm shape with good freezability. In this study, sperm quality at thawing has been higher for those males that have undergone the change on head dimensions mainly during cooling (response pattern 3). It is possible that spermatozoa from these males have responded in a better way to the structural changes resulting from the addition of cryoprotectant. The membrane structure function would not have been impaired during cooling allowing the movement of water and cryoprotectant in an efficient manner. Thus, sperm from these males would be less susceptible to damage from freezing and thawing. Finally, we have identified a third pattern of response, in which sperm have changed their morphometry in a progressive way (response pattern 2). The males having this response pattern have been characterized by large sperm head dimensions on fresh semen and small dimensions and low quality on thawed semen. Two hypotheses could support the behavior of this sperm: that the sperm membrane structure has been significantly damaged during cooling so the exchange of water and cryoprotectant has occurred in a suboptimal way, or that given the characteristics of their cell membranes, more time is needed to complete the exchange. As a result, spermatozoa would not have responded appropriately to changes in their environment during freezing and thawing, leading to a decrease of quality on thawed semen. We think it would be of utmost importance to further study the cryopreservation process in males that have showed a progressive change on their sperm head dimensions. Confirmation that the membrane structure of spermatozoa would require more time to complete the water and cryoprotectant exchange during cooling would allow the design of specific freezing protocols for each type of male.
This study has allowed us to evaluate whether different ejaculates of a male had the same sperm head dimensions and if these ejaculates showed the same pattern of response throughout the cryopreservation process. Results have showed that, in fresh semen, all sperm head morphometric parameters were quite similar for the different ejaculates of a male, being the coefficients of variation of low magnitude. Also, males have always showed the same pattern of response to cryopreservation for the three ejaculates assessed. The low variability on sperm head dimensions between ejaculates of a male has already been stated for this breed. Thus, Maroto-Morales et al.  have reported low CV’s for all the sperm head dimensions. These authors have presented the important role of sperm head morphometry on the characterization of spermatozoa of a male given its low intra-male variability, and have supported the genetic determinism of the sperm morphometry pointed out by other authors , . To further explore this idea, we examined the degree of relatedness (obtained from the additive relationship matrix) between males showing the same (within-) and different (between-) response patterns. Results have showed that additive relationship coefficients for males belonging to the same group were twice larger than those among groups of response patterns. Thus, males who share the same pattern of response were found to be more closely related to each other than with those showing different responses. This result supports the idea of a genetic control of sperm cryoresistance.
In conclusion, the present study shows that sperm head morphometry is a key feature to be included in conservation and breeding programs. The assessment of response patterns suggested here allows us to characterize the sperm cryore