ng, with males showing the response pattern 2 having the lowest quality. The lack of differences of quality in fresh semen would not allow us to characterize the behavior of sperm of a male during cryopreservation based on these parameters , –. In contrast, changes in morphometry of sperm heads observed throughout the cryopreservation have proved useful to this purpose.
A number of studies have attempted to address the role of sperm head morphometry on the prediction of freezability , , –, . Although these studies have addressed this goal in different ways, most of them have used related measures of sperm head dimensions on fresh semen with measures of sperm quality at thawing. In general, these studies have reported that males with smaller sperm heads in fresh semen showed better cryoresistance than those with larger heads. Furthermore, some studies have observed that the sperm heads that had a lower rate of change on their dimension, from fresh to thawed semen, had better cryoresistance . The present study provides no information on which sperm parameters are most valuable as predictors of cryoresistance, since we perform a freezing-thawing process to gather information on sperm cryosurvival. Instead, we use this information to characterize the sperm cryoresistance of a male. Our work was based on the hypothesis that the response of spermatozoa to the freezing-thawing process could be an inherent feature of males and therefore remain constant. This idea is based on the fact that sperm head morphometry has been showed to be a quite constant feature of males . Here, we propose to characterize the sperm cryoability of a male rather than to predict this ability. Furthermore, we believe that this should be based on the study of patterns of response, rather than on the evaluation of a particular feature at a particular time. Had we focused only in the sperm head dimensions in fresh semen, this study would not have allowed us to identify the role of this feature in predicting cryoresistance. Males that suffered changes on their sperm head dimensions during cryopreservation (response patterns 2 and 3) showed significant differences in sperm quality at thawing but were characterized by a similar sperm head morphometry in fresh semen. And those males with response patterns 1 and 3 showed significant differences in fresh semen but resulted in quite similar quality after thawing. Therefore, it is important to know that the sperm dimensions have changed, but also how these changes occur and at which time of the cryopreservation process.
Cryopreservation involves major changes in sperm cell environment, and their success to survive this process will depend largely on their ability to respond to these changes within the finite time period allowed by the protocol . The responsiveness of spermatozoa to osmotic challenge and their ability to regulate cell volume is a characteristic closely related to cryopreservability . Five distinct processing steps can be recognized in relation with changes in cellular volume : extension and cooling, cryoprotectant addition and packaging, freezing, storage and thawing, each having its special relationship to membrane structure function and cell metabolism. Cooling is known to alter the physical properties of all cell membranes –. The second step demands a large volume change and a rapid recovery –. The addition of cryoprotectant leads to osmotically driven egress of intracellular water, followed by a slower return to the originally volume as the cryoprotectant enters. Freezing alters membrane structure and volume over a very short time period . Extracellular ice crystallization results in very large increases of all other solutes in the remaining liquid and the outward movement of water in response to high concentrations. Finally, thawing requires membrane recovery and cellular expansion.
The changes that occur in sperm volume during cryopreservation and their mechanisms allow us to hypothesize about sperm behavior in term of patterns of response. One of the patterns of response identified in this st