Sperm Nuclear Transformations in Cytoplasmic Extracts from Surf Clam (Spisula solidissima) Oocytes

Abstract

Following their incorporation into oocytes, sperm nuclei (SN) of the surf clam, Spisula solidissima, undergo an initial expansion, followed by condensation and then a dramatic enlargement during their development into male pronuclei. These changes are temporally correlated with alterations in the maternal chromatin: germinal vesicle breakdown (GVBD), meiotic maturation, and female pronuclear development, respectively. To analyze possible changes occurring in SN at fertilization, surf clam oocyte extracts; prepared before and after parthenogenetic activation, were examined for their ability to affect SN in vitro. Sperm heads were incubated in extracts for variable periods up to 5 hr. Extracts prepared from oocytes following GVBD (15 min postactivation) induced an expansion in ∼90% of SN by 60 min incubation. However, when SN were incubated in extracts from unactivated or 4-min-activated oocytes only ∼30% underwent expansion. Ultrastructural examination of specimens taken at increasing periods of incubation in oocyte extracts revealed that SN expansion in vitro resembled chromatin decondensation in vivo. SN incubated 1 to 5 hr in extracts prepared from oocytes following GVBD consisted of decondensed chromatin surrounded to varying degrees by membranous cisternae. Staining with anti-lamin antibody was variable: some specimens (60-70%) were positive while others (30-40%) were weak to negative. In contrast, all decondensed SN incubated in extracts from postmeiotic oocytes (65 min postactivation) were delimited by an intact nuclear envelope possessing nuclear pores and reactive to anti-lamin antibody. Decondensation of SN in 15- or 65-min extracts was blocked by EDTA, 2,6-dimethylami-nopurine, histone, and protamine. The presence (65-min extract) and absence (unactivated, 4- and 15-min extracts) of sperm nuclear envelope assembly in vitro is consistent with events in vivo, where such a structure forms after meiotic maturation in concert with the development of the female pronucleus.