Membrane fusion events in the Ca2+/ionophore-induced acrosome reaction of ram spermatozoa

Abstract

An acrosome reaction was induced in ejaculated ram spermatozoa by treatment with calcium and the ionophore A23187. Samples were fixed at different times after initiation of induction, and the morphological changes within the head membranes that took place as exocytosis occurred were studied in freeze-fracture replicas. Reacted acrosomes appeared in individual spermatozoa within the calcium/ionophore-treated population at different times after the start of treatment; the first cells had reacted by 10 min, whereas some took more than 40 min to react. No changes were observed in control populations. An early effect of treatment (seen in most cells within 10 min) was the appearance of particle-free ‘clearings’ in the plasma membrane over the entire acrosomal region, with aggregation of intramembranous particles between and around these ‘clearings’. At the same time, there was an increase in the number of large particles (greater than or equal to 10 nm) within the plasma membrane over the ‘lunula’ of the equatorial segment and the anterior part of the post-acrosomal region. Fusion of the plasma and outer acrosomal membranes began in a limited area at the border between the anterior and equatorial segments of the acrosome. It then spread, following arborescent pathways, sideways along this border and forwards towards the apex of the head. This labyrinthic propagation resulted in an ‘acrosomal cap’ increasingly fenestrated towards its posterior margin. Fusion propagation over the equatorial segment was inhibited, apparently as a result of the highly ordered structure of the membranes in this region.