Regulated exocytosis is controlled by internal and external signals. The molecular machinery controlling the sorting from the newly synthesized vesicles from the Golgi apparatus to the plasma membrane play a key role in the regulation of both the number and spatial location of the vesicles. In this context the mammalian acrosome is a unique vesicle since it is the only secretory vesicle attached to the nucleus. In this work we have studied the membrane trafficking between the Golgi apparatus and the acrosome during mammalian spermiogenesis. During bovine spermiogenesis, Golgi antigens (mannosidase II) were detected in the acrosome until the late cap-phase spermatids, but are not found in testicular spermatozoa (maturation-phase spermatids). This suggests that Golgi-acrosome flow may be relatively unselective, with Golgi residents retrieved before spermiation is complete. Surprisingly, rab7, a protein involved in lysosome/endosome trafficking was also found associated with the acrosomal vesicle during mouse spermiogenesis. Our results suggest that the acrosome biogenesis is associated with membrane flow from both the Golgi apparatus and the endosome/lysosome system in mammalian spermatids.