Cryopreservation of macropodid spermatozoa: new insights from the cryomicroscope

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Title: Cryopreservation of macropodid spermatozoa: new insights from the cryomicroscope
Author(s): Holt, W. V.; Penfold, L. M.; Johnston, S. D.; Temple-Smith, P.; McCallum, C.; Shaw, J.; Lindemans, W.; Blyde, D.
Abstract: This study examined the effects of cooling and cryopreservation upon macropod spermatozoa (eastern grey kangaroo, Macropus giganteus and red-necked wallaby, Macropus rufogriseus). Sperm survival during and after freezing to -30°C or-70°C in minimum essential medium (MEM) + 5, 10, 20 or 30% (v/v) glycerol, MEM + 10 or 20% (v/v) ethylene glycol and MEM containing a mixture of 7.5% (v/v) glycerol + 10% (v/v) dimethylsulphoxide was examined by cryomicroscopy. The MEM/glycerol mixtures permitted better post-thaw sperm recovery than the other cryoprotectants. After freezing to -30°C at 10°C min-1 in 20% glycerol, then rewarming at 20°C min-1, flagellar activity resumed in more than 50% of spermatozoa when the temperature increased into the range 5-10°C. However, as the temperature increased, into the range 20-25°C, motility declined rapidly so that less than 5% motile cells were seen at 35°C. Spermatozoa in MEM without cryoprotectant were also examined by cryomicroscopy to evaluate changes in flagellar configuration, swimming behaviour and viability during cooling from 35°C to approximately -7°C, and rewarming to 35°C. Cooling from 35 to 28°C induced kangaroo spermatozoa to exhibit rigid principal-piece bending and non-linear motility, which was reversed by further cooling and the spermatozoa resumed their normal linear movement. Rewarming induced principal-piece bending in the range of 20-30°C, but this effect was reversed by further warming. Although red-necked wallaby spermatozoa showed these effects, they also exhibited a tendency to form rosette-like clusters during rewarming, especially when the temperature reached approximately 14°C. The clusters were induced when the flagellar end-pieces became anteriorly reflected, producing hook-like flagellar conformations, which then became interlinked.
Publication type: Journal article
Source: Reproduction, Fertility and Development, Vol. 11, no. 6 (1999), pp. 345-353
Publication year: 1999
Keyword(s): Genetic resource banks; Kangaroos; Plasma membrane; Semen; Wallabies
Publisher: CSIRO Publishing
ISSN: 1031-3613
Publisher URL: http://dx.doi.org/10.1071/RD99076
Peer reviewed