Induction of polyspermy by treatment of sea urchin eggs with mercurials

Abstract

This study is intended to be a contribution to our knowledge of the mechanism protecting the sea-urchin egg against polyspermy. Various degrees of polyspermy were observed upon insemination of eggs that had been pretreated with the mercurials para-chloro mercuriphenylsulfonic acid (PCMPS) or salicyl-γ-hydroxymercuri-β-methoxypropylamide-0-acetic acid, mersalyl (salyrgan). The exposure of the unfertilized eggs to the reagents lasted usually for 15 min. Thereafter the eggs were washed and inseminated in sea water. The concentration range of the mercurials was approximately 10 −7−10 −4 M. The effect varied from batch to batch of eggs, indicating a variation of availability of sulfhydryl (SH-) groups in different egg materials. An attempt was made to define a 50 per cent value (−log Km) for the change in the surface that allows one supernumerary spermatozoon to enter. This value would serve the purpose of making experiments with different egg materials more directly comparable (see pp. 162–163). The degree of polyspermy was also dependent on sperm concentration, as exemplified on p. 163. The progress of polyspermy was followed by immersing the eggs for different periods after insemination in 0.001 per cent lauryl sulfate, which stops any further fertilization. Despite the mercurial pretreatment the eggs that remained monospermic developed normally. Pathological development of mercurial-pretreated eggs depended on a polyspermic state (p. 160). Merthiolate had a weaker polyspermy-promoting action than PCMPS and mersalyl. Whereas these latter compounds seemed to act only on the surface layers of the egg, merthiolate (as also p-chloromercuribenzoate and N-ethylmaleimide) also injured the internal processes (p. 167). The effect of mercurials could be removed by a subsequent exposure to cysteine. The concentrations of cysteine necessary were much higher (e.g. 1000 times) than that of mercurial. The mercurials are thus strongly linked, probably to proteins (pp. 167–168). On leaving the eggs in pure sea water after pretreatment with mercurials, their effect is lowered (pp. 168–169). Exposure of the eggs to ~10 −4 Mo-iodosobenzoate (IBA) decreased the effect of a following mercurial treatment. This confirms that SH groups are involved in the action of mercurials on the egg surface (pp. 164–165). Exposure to and insemination in the presence of ATP (1.8 × 10 −5 M) moderated the effect of pretreatment with mercurial (p. 166). Combination of treatment with IBA prior to the exposure to mercurial and after-treatment with ATP gave a higher moderating effect than that brought about by either IBA or ATP (p. 166). Treatment of the eggs with lower and higher doses of trypsin (see [36]) prior to exposure to mercurials did not modify the extent of polyspermy. The combination of low doses of trypsin and mercurials enhanced the suppression of cleavage beyond the effect of the trypsin doses alone (pp. 169–170). The rate of primary fertilization was not modified by the treatment with mercurials (p. 171). Neither does this treatment modify any microscopic changes accompanying fertilization in such a way that these modifications could be considered as the cause of polyspermy (p. 181). The effect of the mercurials must be at a submicroscopic, biochemical level. On the basis of available knowledge the mechanism of protection against polyspermy and changes in S H compounds upon fertilization are reviewed and discussed (pp. 186–190). The results of mercurial treatment applied soon after fertilization are in keeping with an activation of SH enzymes (pp. 185, 189). The site of attachment of supernumerary spermatozoa and their primary effect give further evidence (see [37]) that, prior to fertilization, a proximaldistal polarity is present in the egg (pp. 177, 189). This polarity also means that the eggs are more receptive to spermatozoa in one region, the proximal one. The cortical changes with ensuing protection evolve also more rapidly in the proximal region [37]. These circumstances are considered to be an important factor in the protection of the egg against polyspermy.