Quality evaluation of cat’s epididymal spermatozoa chilled to -1ºC and 4ºC

In cryopreservation of domestic cat’s semen, chilling at 4oC apparently avoids the thermal shock and irreversible loss of sperm cells. However, studies demonstrating the behavior of feline’s sperm cells at different chilling temperatures were not found in the available literature. The aim of this study was to evaluate the quality of fresh domestic cats spermatozoids obtained from epididymis tail, and compare after chilling at -1oC and 4oC for 24 to 48 hours. Twenty nine adult cats were used, mixed breeds, in good nutritional status, weighting 2 to 6 kg, in the period from november 2015 to february 2016. The study was conducted in two stages: in the first was used testes of 14 cats and the chilling temperature of -1oC; the second stage was realized with 15 cats and spermatozoids were chilled at 4o C. After elective orchiectomy, spermatozoids were recovered from epididymis tail and diluted with ACP-117® medium. Spermatozoids quality was evaluated in three stages: fresh, after 24 hours and 48 hours of chilling. Spermatozoids kinetics was calculated by computer analysis (CASA, Hamilton-Thorne IVOS II®, Beverly, USA), concentration by Neubauer’s chamber, morphology by Karras modified staining and membrane integrity by eosin negrosin staining. The results were analyzed using the R 3.2.5 software. The Friedman’s nonparametric test was used to compare the times (fresh, 24 and 48 hours), with 5% of significance levels for all tests. There was no difference in motility when compared the two chilling temperatures (-1oC and 4oC). The average motility of the fresh spermatozoids, 24 and 48 hours after chilling were different even at -1oC (67.5 ± 18.7%; 40 ± 20.9%; 29.2 ± 19.9%) as the 4oC (70.4 ± 11.9%, 28.6 ± 21.7%; 17 ± 19.7%). However, it was visually remarkable the reduction of progressive motility in the chilling at 4°C, which was different in three stages (40.4 ± 9.4%; 12.7 ± 11.7%; 10.5 ± 6.5 %), whereas at -1°C this drop was less pronounced (27.6 ± 14.2%, 17.6% ± 13.5 and 12.2 ± 13%). In morphology, although there was no difference in the percentage of normal cells of the three times at -1o C (52.5 ± 20.3%; 42.3 ± 22.9%; 35 ± 23.3%) and at 4 °C (55.9 ± 43.1%; 43.1 ± 15.2%; 43.4 ± 18.6%), there was a significant increase in larger defects between the chilling at 24 and 48 hours - 1°C (28.4 ± 16.7%; 36.2 ± 22.9%). Nevertheless, 4° C chilling had significant increase in the sperm membrane damage (4.9 ± 3.1%; 8.6% ± 5, 12 ± 7.3%), and was not observed that damage cells when chilled at -1°C (7.1 ± 3.6%; 9.1 ± 5.4%; 10.1 ± 4.6%). The recovered spermatozoids from epididymis tails chilled at -1oC showed acceptable results both in motility as in percentage of spermatozoids integrity. Therefore we conclude that the chilling temperature of -1 °C is effective to maintain the quality of sperm recovered from the epididymis, a new technique can be used in the cryopreservation of domestic cats.