The nucleus of the mammalian spermatozoon is an exceptionally stable organelle requiring very rigorous chemical treatments to free its chromatin (Borenfreund et al., 1961). In spite of this, the nucleus swells and its chromatin begins to decondense very rapidly as the spermatozoon is incorporated into the egg cytoplasm (Austin, 1961;Yanagimachi & Noda, 1970;Bedford, 1972). As yet, the mechanism underlying this phenomenon has not been directly examined.Various methods have been used to decondense sperm nuclei and to isolate and characterize the sperm chromatin and nuclear proteins (Borenfreund et al., 1961;Henricks & Mayer, 1965;Lung, 1972). Sodium dodecyl sulphate (SDS, an anionic surfactant) and dithiothreitol (DTT, a reagent which specifically cleaves disulphide linkages) have been used to study nuclear stabilization during maturation of mammalian spermatozoa (Calvin & Bedford, 1971;Calvin et al., 1973). Nuclear stabilization by disulphide (-S-S-) bonding has been studied from an evolutionary standpoint by Bedford & Calvin (1974), who have found that this -S-S-bonding is prominent in the spermatozoa only of eutherian mammals. In the present study, the types of chemicals that can decondense mammalian sperm nuclei were determined, then the common characteristics of these chemicals were evaluated so that possible inferences could be made about the mechanism of nuclear decondensation during fertilization.Epididymal spermatozoa of the golden hamster, rat, guinea-pig and rabbit were obtained by puncturing the excised cauda epididymidis with a needle and squeezing the spermatozoa into a Petri dish containing 0-9% NaCl at 37°C. Pellets of frozen canine semen were prepared according to the techniques of Seager (1969) and were air-shipped from Oregon to Hawaii in liquid nitrogen (-196°C). For these experiments, pellets were thawed rapidly in 0-9% NaCl (37°C), and fresh canine ejaculates were obtained for comparison. Both fresh and frozen-thawed semen were washed by centrifugation two or four times in 0-9% NaCl and resuspended in 0-9% NaCl. The sperm suspension of each species (0-1 ml) was mixed with an equal volume of a test solution in a 5-ml glass test-tube. The final concentration of spermatozoa varied from 105 to 107 spermatozoa/ml. The tubes were sealed with Parafimi (American Can Co., Neenah, Wisconsin) and incubated at 37°C. At the end of incubation, the pH was measured and the degree of nuclear decondensation was determined by 293