Academic literature on the topic 'Marsupial Spermatozoa Morphology'

SummaryThe morphology of the acrosome of spermatozoa an Australian daysyurid marsupial during in vivo fertilisation, as seen with the tranamission electron microscope, is described. In some spermatozoa close to the outer zona surface with largely intact acrosomes, eletron-demse struts between the inner and outer acrosomal membranes could be seen; in addition fusion between the outer acrosomal and overlying plasma membranes was sometimes evident. The zona matrix close to the acrosomal region of spermatozoa with heads party embedded within the zona pellucida was less electron-dense and more filamentous than elsewhere; perhaps indicative of local lytic activety. Nevertheless some of these spermatozoa at least had partly intact acrosomes with local regions of electron-dense bridges between the two acrosomal membranes. Spermatozoa with condensed chomatin were also observed within the egg cytoplsam some of which had membranes, vacuoles or close to the dorsal nuclear surface which could, in part, be acrosomal in orgin. These observations lend some support to the recent view, based on findings from an American species, that acrosomal changes of spermatozoa of some marsupials at the of fertillsation may be somewhat similar to those of eutherian mamals in spite of the considerable differences in gamete structure and design. How generally applicable these findings are to all marsupials remains, however, to be determined.

Australian marsupials exhibit a wide range of variation in sperm head morphology, and in thickness of the zona pellucida around the oocyte, suggesting interspecfic differences in the processes of sperm-egg interaction. The observations described here are largely based on the dasyurid Sminthopsis crassicaudata. They show that in oestrous females, after mating, a coagulum forms in the lateral vaginae and, within an hour of insemination, numerous spermatozoa congregate in the isthmus of the oviduct in which the vanguard population undergoes transformation with the head rotating on its axis with the tail to form a T-shape. Once oocytes are released, a few spermatozoa migrate to the higher reaches of the oviduct where sperm-zona binding occurs by way of the plasmalemma over the acrosomal region. The acrosome reaction takes place here and, as the egg rotates, the tail of the spermatozoon becomes parallel to the head. A small region of acrosome sometimes appears to remain intact at this time because spermatozoa with partly intact acrosomes have been found within the zona matrix. In some of these, electron-dense bridges between part of the inner and outer acrosomal membranes which may act as stabilizing structures, were also seen. The zona matrix is tightly packed around the penetrating spermatozoon, but that close to the acrosomal region becomes less electron-dense and more filamentous. Once incorporated into the egg, the spermatozoon lacks a cell membrane around the tail but vesicles close to the sperm head may, at least in part, be remnants of an inner acrosomal membrane. How generally applicable these observations are to other Australian marsupials remains to be determined.

The effects of long-term cooling and freezing on sperm motility are described for six marsupial species: the fat-tailed dunnart, koala, brushtail possum, long-footed potoroo, northern brown bandicoot and ring-tailed possum. The effects of up to eight days of cooling at 4 degrees C on the motility of dunnart spermatozoa and the effect of cryopreservation on spermatozoa of the other species were determined. The cryoprotectant used was a Tris-citrate-fructose-egg yolk-glycerol diluent. The percentage and rating of sperm motility, and sperm structure, as determined by light microscopy, were investigated. Sperm motility in the fat-tailed dunnart was retained for up to six days when cooled to 4 degrees C, suggesting that sperm from this species have some degree of tolerance to cold shock. After this time, however, the percentage of motile spermatozoa and their motility rating declined. In all species except the fat-tailed dunnart, reinitiation of motility following cryopreservation occurred across a range of glycerol concentrations (4-17%). Cryoprotectant containing 6% and/or 8% glycerol resulted in little change of motility rating or of the percentage of live sperm after thawing, although there was some decline in the percentage of motile sperm. The unusual structural and motility characteristics of dunnart spermatozoa may account for the lack of success of sperm cryopreservation in this species.

A male Tasmanian devil, aged 6.5 years, was euthanased with intravenous pentobarbitone overdose due to the diagnosis of squamous cell carcinoma. The testes and cauda epididymes were removed immediately after euthanasia and placed into TALP-HEPES medium containing 3% BSA and dissected. Smears of the isolated sperm were made, air-dried and stained using Diff-Quik. Sperm were classified using a modification of a system originally applied to the brush-tailed possum1 as follows: (a) Immature I – head is perpendicular or greater than 45° to the tail.(b) Immature II – head forms an acute angle of 45° or less with the tail.(c) Immature III – head and tail lie in a straight line but the sperm head still has a partly expanded acrosome.(d) Mature – the head is aligned with and has a diameter similar to the tail. No motility was seen in the testicular sperm, but >60% of sperm from the cauda epididymes were motile. The stained preparations showed a significant difference in the distribution of sperm between the testis and epididymis (χ2 = 150.45, df = 3, P < 0.001), with sperm within the testis being skewed towards the earlier stages of development whereas the epididymis was skewed towards the later stages of development (Table 1). These results confirm the similarity with several other marsupials in that sperm are formed in the testis with the head at right angles to the tail but rotation of the head to become aligned with the tail is completed during epididymal transport. Work is now required to identify optimum conditions for the culture, cryopreservation and insemination of sperm from this species. (1)Cummins JM. (1976). Epididymal maturation of spermatozoa in the marsupial Trichsurus vulpecula: changes in motility and gross morphology. Aust. J. Zool. 24, 499–511.

The establishment of a functional genome resource bank for the genetic management and future proofing of Australian native mammals sounds great in theory, but what is the reality of this idea. In order to understand the current rate of progress in this area, we will present an overview of the inherent structural and physiological limitations of non-eutherian mammalian reproduction in terms of gamete biology and ART. For the male, these include (1) an unique mode of spermatid condensation that imparts the need for major structural changes to sperm morphology during epididymal transit, (2) a lack of cysteine protamines and disulphide bonds in the sperm chromatin that predisposes the nucleus to post-thaw chromatin relaxation, (3) an extremely stable acrosome, which to date, has not been possible to experimentally react in vitro, (4) unusual lipid composition in the plasma membrane that potentially makes the sperm cell resistant to cold shock trauma and (5) the need, in some species, for extremely high concentrations of cryoprotectant, that paradoxically, appear to be cytotoxic to the spermatozoon. Female limitations include, (1) the production of a large yolky oocyte and resulting embryo, making it difficult to cryopreserve, (2) a small and technically challenging complex reproductive tract that makes gamete recovery and artificial insemination problematic and (3) a general lack of information on marsupial reproductive physiology and behaviour that has hindered the development of protocols for timed induction of oestrus and ovulation. We shall also identify, socio-political and ethical limitations holding back the application of assisted breeding technology in these species.

Variation in localization and distribution of saccharides on the sperm surface of a marsupial, the brushtail possum, Trichosurus vulpecula, was compared between spermatozoa from the caput and cauda epididymides. Spermatozoa were subjected to the following treatments: (i) unfixed and fixed spermatozoa were stained with fluorescein-labelled lectins; (ii) unfixed spermatozoa were incubated with lectins for determination of agglutination; and (iii) spermatozoa were incubated with detergent to remove the plasmalemma, the glycoproteins were separated on SDS-PAGE and western blots were stained with biotinylated lectins. Many of the fluorescein isothiocyanate (FITC)-labelled lectins bound selectively to the sperm surface, and marked differences were found in lectin staining affinity between caput and cauda epididymal spermatozoa. Incubation of spermatozoa from the cauda epididymidis with neuraminidase reversed many of the differences in staining of the cauda epididymal spermatozoa, indicating masking of some terminal saccharides by sialic acid. Agglutination of spermatozoa from the caput epididymidis occurred after incubation with Concanavalin A (ConA) and soybean agglutinin (SBA), but agglutination was less extensive for spermatozoa from the cauda epididymidis. Western blot analysis indicated several ConA-positive bands in caput sperm extracts, but fewer positive bands in the cauda sperm extracts, whereas SBA stained four bands from caput but none from the cauda epididymal spermatozoa. These results demonstrate extensive glycosylation of the surface proteins of spermatozoa from the caput epididymidis and significant differences in spermatozoa from the cauda epididymidis. In general, the findings indicate similar glycosylation of the surface of marsupial spermatozoa to those from eutherian mammals despite marked differences in their morphology and early divergence of marsupials from eutherian mammals. It would appear that this situation differs markedly from that in sub-mammalian vertebrates.