Nuclear chromatin decondensation of spermatozoa in vitro: a method for evaluating the fertilizing ability of ovine semen

Abstract

Spermatozoa obtained from testes, epididimydes and complete ejaculates of healthy rams during the breeding and non-breeding seasons were induced to show nuclear chromatin decondensation by controlled exposure to dithiotreitol (DTT) and sodium dodecyl sulphate (SDS) in vitro. A gradual resistance to decondensation was shown by sperm during epididymal transit, confirming a progressive increase in the prevalence of chromatinic disulphide bonds during sperm maturation in this species. A high % of stable (non-decondensed) sperm nuclei after treatment (79%) was found in semen from rams with normal fertility (64% non-return rate at first oestrus). Opposite changes were found in the semen from rams having low fertility rates (37%), as these showed only 31% of stable sperm nuclei. There were no differences in the spermiograms of these two groups. When semen from the same rams was tested during the non-breeding season, a similar relationship was found, although in both groups there was a higher % of sperm with stable nuclei than during the breeding season. The possible role of seminal plasma and of some of its constituents (e.g., zinc) on the decondensation of ram sperm nuclear chromatin was also studied. The presence of seminal plasma and the addition of zinc largely or completely inhibited the decondensation of ram sperm nuclear chromatin whilst the reverse situation was seen following the addition of chelating agents (e.g. EDTA) to the semen samples. The present results suggest that the induction of sperm nuclear decondensation by exposure to DTT and SDS under controlled conditions may provide a simple but reliable method for predicting in vitro the fertilizing ability of a ram semen sample.

Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art

“…133 Methods for assessing decondensation must therefore be considered useful for male fertility. Several groups have measured sperm chromatin decondensation by observing changes in size and shape of the sperm head by microscopic examinations when exposed to detergents (mostly sodium dodecylsulphate) and S-S bond reduction (using dithiothreitol), 134,135,136 methods that can predict fertility in vitro. FC offers an excellent possibility to quantity events associated with chromatin decondensation.…”

Distribution of DNA, nuclear micro-heterogeneities and compaction of the chromatin in rabbit epididymal spermatozoa. Ultrastructural evaluation of the Feulgen-like technique using osmium ammine

“…Therefore, in sperm nuclei, the Feulgenlike (Gledhill, 1975;Gledhill et al, 1966;Hunter ef al, 1976). This can be evaluated by measuring the decondensation ability of the chromatin using, for example, swelling of sperm heads by detergents (Rodriguez et al, 1985;Rosenborg ef al, 1990), or quantification of accessible protein side groups such as sulfhydryl groups (Bedford and Calvin, 1974), amino groups , or epitopes of the main sperm nucleoprotein, the protamine (Rodriguez et al, 1990). The accessibility of DNA is also one such criterion (Gledhill et al, 1971 This can be evaluated using, for example, intercalating dyes Kasten, 1960) and has been widely used in light microscopy.…”

A Method for Reactivation of the Movement of Demembranated Ram Sperm Models with Decondensed Nuclei and a Description of their Ultrastructure

“…Human spermatozoa have been reported to be more variable in this respect in both the cauda [16] and in ejaculates [17] even when demembranated with SDS and when treated with EDTA or otherwise rendered deficient in zinc [18]. The percentage of spermatozoa decondensing in ejaculates has been used as a diagnostic technique for evaluation of fertility [17,19]. Another technique utilizing decondensed spermatozoa is in situ hybridization, e.g., in assaying Y-bearing sperm for use in monitoring sperm separation techniques [20].…”