CELLULAR-LOCALIZATION OF POLYAMINES - CYTOCHEMICAL AND ULTRASTRUCTURAL METHODS PROVIDING NEW CLUES TO POLYAMINE FUNCTION IN RAM SPERMATOZOA

Polyamine binding sites have been localized in ram spermatozoa using b iochemical and cytochemical tools. Incubating the cells with C-14-sper mine and determining its distribution after sonication and differentia l centrifugation, revealed that 60% of the radioactive spermine was lo calized in the head, 21.5% in the tail and about 9% in the plasma memb rane. A polyamine specific cytochemical staining by the formaldehyde-f luorescamine method, revealed that most of the polyamines were localiz ed in the midpiece, where the cell mitochondria are located, and in th e acrosome region. Two additional studies used electron microscopy, em ploying polycationic colloidal gold and spermine-ferritin as cytochemi cal markers. The most sensitive and specific method was the staining o f the cells with ferritin-spermine whose synthesis is described in thi s study. The outer acrosomal membrane was the preferential site for sp ermine binding which was densely distributed in a highly orderly patte rn. There was a sparse distribution of spermine binding sites on the p lasma membrane surrounding the acrosome and none on the post acrosomal region. The role of spermine in the acrosome reaction and Ca2+ fluxes in sperm cells is discussed.