Comparison of quick-frozen and chemically fixed sea-urchin eggs: structural evidence that cortical granule exocytosis is preceded by a local increase in membrane mobility

Abstract

Eggs of the purple sea-urchin, Strongylocentrotus purpuratus, were fertilized and fixed with 2% glutaraldehyde at various stages during cortical granule exocytosis. Fixation resulted in membrane blebs being formed precisely at the point of incipient granule fusion. These blebs pinched off to form the membranous vesicles frequently seen in exocytic pockets and in the perivitelline space. In contrast, eggs that were fixed with osmium tetroxide or were quick-frozen without chemical fixation, showed no signs of bleb or vesicle formation. Rather, fusion of each granule appeared to begin at a single minute pore, 30-50 nm in diameter, which then enlarged. We suggest that formation of blebs during glutaraldehyde fixation is an artifact that is caused by a highly localized and transient increase in membrane mobility. Normally, this increased mobility facilitates fusion of granule and plasma membranes, but in the presence of glutaraldehyde it leads to large-scale distortions of these fusing membranes.