Evaluation of fertilizing potential of frozen-thawed dog spermatozoa diluted in ACP-106 using an in vitro sperm--oocyte interaction assay.
The aim of present study was to evaluate frozen canine semen with ACP-106 (Powder Coconut Water) using an in vitro sperm--oocyte interaction assay (SOIA). Ten ejaculates from five stud dogs were diluted in ACP-106 containing 20% egg yolk, submitted to cooling in a thermal box for 40 min and in a refrigerator for 30 min. After this period, a second dilution was performed using ACP-106 containing 20% egg yolk and 12% glycerol. Samples were thawed at 38 degrees C for 1 min. Post-thaw motility was evaluated by light microscopy and by using a computer aided semen analysis (CASA). Plasma membrane integrity and sperm morphology/acrosomal status were evaluated by fluorescent probes (C-FDA/PI) and Bengal Rose respectively. Moreover, frozen-thawed semen was analysed by a SOIA. Subjective post-thaw motility was 52.0 +/- 14.8% and it was significant higher than the total motility estimated by CASA (23.0 +/- 14.8%) because this system considered the egg yolk debris as immotile spermatozoa. Although normal sperm rate and acrosomal integrity evaluated by Bengal Rose stain was 89.6 +/- 3.1% and 94.3 +/- 3.1%, respectively, post-thaw percentage of intact plasma membrane was only 35.1 +/- 14.3%. Regarding SOIA, the percentage of interacted oocytes (bound, penetrated and bound and/or penetrated) was 75.3%. Using regression analysis, it was found significant relations between some CASA patterns and data for SOIA. In conclusion, the freezing-thawing procedure using ACP-106 was efficient for maintain the in vitro fertility potential of dog spermatozoa.
Cardoso
R C S
RC
Laboratory of Carnivore Reproduction - FAVET, UECE, Itaperi, Fortaleza, Ceará, Brazil. cardosorcs@yahoo.com.br
Silva
A R
AR
Silva
L D M
LD
Chirinéa
V H
VH
Souza
F F
FF
Lopes
M D
MD
D000177:Acrosome / Q000502:physiology; D000818:Animals; D015925:Cryopreservation / Q000662:veterinary*; D003451:Cryoprotective Agents / Q000494:pharmacology*; D003470:Culture Media / Q000737:chemistry; D004285:Dogs / Q000502:physiology; D005434:Flow Cytometry; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility; D013084:Sperm-Ovum Interactions / Q000502:physiology*; D013094:Spermatozoa / Q000502:physiology*
D003451:Cryoprotective Agents; D003470:Culture Media