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United States Department of Agriculture

Agricultural Research Service

Title: Effects of Different Cryopreservation Methods on the Glyocalyx of Chicken Spermatozoa

Authors
item Pelaez, Jesus
item Long, Julie

Submitted to: Joint Meeting of the ADSA, AMSA, ASAS and PSA
Publication Type: Abstract Only
Publication Acceptance Date: March 2, 2007
Publication Date: July 18, 2007
Citation: Pelaez, J., Long, J.A. 2007. Effects of Different Cryopreservation Methods on the Glyocalyx of Chicken Spermatozoa. Joint Meeting of the ADSA, AMSA, ASAS and PSA. Journal of Poultry Science. 86:(Suppl 1)651.

Technical Abstract: The carbohydrate-rich zone on the sperm surface is essential for inmunoprotection in the female tract and early gamete interactions. We recently have shown the glycocalyx of chicken sperm to be extensively sialylated and contain residues of mannose, glucose, galactose, fucose, N-acetyl-galactosamine, N-acetyl-glucosamine and N-acetyl-lactosamine. Our objective here was to evaluate the effects of cryopreservation on the sperm glycocalyx. Semen was pooled from 6 roosters, diluted 1:1 (Lake’s pre-freeze diluent), cooled to 5°C and aliquoted for cryopreservation using 6% DMA, 11% DMSO or 11% glycerol. For the DMA method, semen was equilibrated for 1 min with DMA and rapidly frozen by dropping 25 µl aliquots into liquid nitrogen. For the DMSO and glycerol methods, semen was equilibrated for either 1 min (DMSO) or 20 min (glycerol), loaded into 0.25 ml straws and frozen (5 to -35°C, 7°C/min; -35 to -140°C, 20°C/min; nitrogen plunge). Thawed (rapid, DMA; moderate, DMSO, glycerol) semen was stained with 1 of 12 FITC-conjugated lectins (100 µg/mL; 30 min; 25°C; 100x106 cells/mL). Samples counterstained with PI were assessed by flow cytometry. On the day of cryopreservation, aliquots of fresh semen were stained with the panel of lectins and PI. For each lectin, the Mean Fluorescence Intensity (MnFI) of live sperm was compared among fresh and frozen/thawed (fr/th) treatments (n=5 replicates). For the majority of lectins (10/12), the MnFI was higher (p<0.05) for fr/th than fresh sperm. Exceptions included lectins specific for sialic acid and '-fucose, where DMSO and glycerol treatments, respectively, had MnFI similar (p>0.05) to fresh sperm. Among the fr/th treatments, the MnFI of sperm cryopreserved with DMSO was higher (p<0.05) for 4/10 lectins, including those specific for N-acetyl-lactosamine and N-acetyl-glucosamine. These data indicate that surface carbohydrates are altered during cryopreservation, and that cryoprotectant type and fr/th rates affect the degree of modification. While the specific functions of these glycoconjugates are not known, it is likely that the observed differences in fr/th sperm contribute to the reduced fertility of cryopreserved chicken semen.

Last Modified: 8/27/2014
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