Effect of cryoprotectants and male on motility parameters and fertilization rate in paddlefish (Polyodon spathula) frozen-thawed spermatozoa

Otomar Linhart, Steve D. Mims, Boris Gomelsky, Ludmila I. Cvetkova, Jacky Cosson, Marek Rodina, A. Horváth, B. Urbányi

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

A sperm cryopreservation method using different cryoprotectants and sperm from different males was developed. Different percentages of pure cryoprotectants (methanol and DMSO) were added to extender 1 or 2 (20 mM tris pH 8 + 30 mM sucrose + 0.5 mM KCl or 20 mM tris pH 8 + 50 mM sucrose + 0.5 mM KCl, dilution 1:1) or non-extended sperm and every 1 ml of mixture was transferred to a 2-ml cryotube. The cryotubes were directly transferred to a pre-programmed PLANER Kryo 10 series III freezer at 0 °C and cooled from 0 °C to -5 °C at a rate of 3 °C.min-1, from -5 °C to -15 °C at a rate of 5 °Cmin-1, from-15 °C to -25 °C at a rate of 10 °C .min-1, from-25 °C to -80 °C at a rate of 20 °C .min-1. Thereafter the samples were held for 5 min at -80 °C and finally transferred into LN2 until next morning. The sperm was then thawed in a water bath at 40 °C for 105 s. Fertilization rate of control sperm was 81.5% (kept unfrozen; samples tested after 24-h storage at 3 °C), indicating that the gametes were of good quality. Percentage and the velocity of motile sperm from were evaluated in fresh and post-thawed sperm using video frames and subsequent image analysis. The results on hatching rates were significantly correlated with post-thawed sperm motility (r=0.49, P=0.035) and velocity (r=0.55, P=0.014) and not correlated with velocity of post-thawed spermatozoa (r=0.32, P=0. 177). The best fertilization rates were obtained with 64-75 % in post-thawed sperm (3.6.105 spermatozoa per egg) when sperm were treated either without any extender or with both extenders with methanol concentrations of 8 or 10 %. These results were not significantly different compared with those obtained using fresh sperm control samples. Hatching rate was very low, only 8-15 %, when sperm was frozen with 8 or 10 % DMSO. ANOVA showed a significant effect of males on sperm motility, velocity and fertilization rate in post-thawed sperm.

Original languageEnglish
Pages (from-to)389-394
Number of pages6
JournalJournal of Applied Ichthyology
Volume22
Issue numberSUPPL. 1
DOIs
Publication statusPublished - Dec 2006

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Polyodon spathula
Polyodontidae
motility
cryoprotectants
sperm
spermatozoa
fertilization (reproduction)
parameter
effect
rate
sperm motility
sucrose
hatching
methanol
cryopreservation
freezers
gamete

ASJC Scopus subject areas

  • Aquatic Science

Cite this

Effect of cryoprotectants and male on motility parameters and fertilization rate in paddlefish (Polyodon spathula) frozen-thawed spermatozoa. / Linhart, Otomar; Mims, Steve D.; Gomelsky, Boris; Cvetkova, Ludmila I.; Cosson, Jacky; Rodina, Marek; Horváth, A.; Urbányi, B.

In: Journal of Applied Ichthyology, Vol. 22, No. SUPPL. 1, 12.2006, p. 389-394.

Research output: Contribution to journalArticle

Linhart, Otomar ; Mims, Steve D. ; Gomelsky, Boris ; Cvetkova, Ludmila I. ; Cosson, Jacky ; Rodina, Marek ; Horváth, A. ; Urbányi, B. / Effect of cryoprotectants and male on motility parameters and fertilization rate in paddlefish (Polyodon spathula) frozen-thawed spermatozoa. In: Journal of Applied Ichthyology. 2006 ; Vol. 22, No. SUPPL. 1. pp. 389-394.
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