Protamine sulfate induces enzyme secretion from rabbit neutrophils. Enzyme secretion is mainly due to exocytosis but, depending on the experimental conditions, a small amount of cytolysis may occur. As compared with stimulation of neutrophil functions by other activators, protamine sulfate-induced enzyme release by exocytosis is a relatively slow process and is not accompanied by a marked activation of the metabolic burst. For optimal exocytosis, extracellular Ca2+ is required, but there is still some enzyme release in its absence, and other metal ions (Sr2+, Ba2+, Mg2+) can partly mimic the effect of Ca2+. Positive charges on protamine are of primary importance because the polyanion heparin completely inhibits protamine sulfate-induced enzyme release. Protamine linked to agarose beads is able to induce enzyme release; thus the induction of exocytosis is due to an interaction of the positive charges on protamine with the plasma membrane. Sialic acid residues on the membrane, however, seem not to play an important role in this process.