Maintenance of Motility in Mouse Sperm Permeabilized with Streptolysin O1

Abstract

One approach to studying the mechanisms governing sperm motility is to permeabilize sperm and examine the regulation of motility by manipulating the intracellular milieu of the cell. The most common method of sperm permeabilization, detergent treatment, has the disadvantage that the membranes and many proteins are extracted from the cell. To avoid this problem, we have developed a method that uses streptolysin O to create stable pores within the plasma membrane while leaving internal membranes intact. Sperm were permeabilized, preincubated, and then treated with 0.6 U/ml of streptolysin O. Permeabilization was assessed by fluorescent dye technologies and endogenous protein phosphorylation using exogenously added [gamma-32P]ATP. Streptolysin O-induced permeabilization rendered the sperm immotile, and the effect was Ca2+-dependent. When the cells were treated simultaneously with a medium containing ATP, streptolysin O-treated sperm maintained flagellar movement. These results demonstrate that the streptolysin O permeabilization model system is a useful experimental method for studying the mechanisms that regulate sperm motility since it allows the flagellar apparatus to be exposed to various exogenously added molecules.

F‐actin involvement in guinea pig sperm motility

“…In nonSLO permeabilized spermatozoa neither 5 mM gelsolin in calcium medium nor 2 mM gelsolin in calcium-free medium altered motility, that is, permeabilization was necessary for the gelsolin effect on sperm motility to occur. In mouse spermatozoa, SLO treatment rendered the sperm immotile (Johnson et al, 1999). Contrary to these results, in guinea pig spermatozoa, we did not observe changes in the motility of control versus SLOpermeabilized cells up to 4.5 hr after permeabilization.…”

Haprin, a Novel Haploid Germ Cell-specific RING Finger Protein Involved in the Acrosome Reaction

“…The anti-Haprin antibodies were able to pass through the pores made by SLO in the sperm plasma membrane, whereas the acrosome itself and the outer acrosomal membrane remained intact (17,18). The antibody against the N-terminal region, but not the antibody against the C-terminal region of Haprin, inhibited the acrosome reaction of sperm (Fig.…”

“…Haprin protein is expressed specifically in testis and is localized in the acrosomal region of testicular germ cells and mature sperm. In order to investigate the role of Haprin in the acrosome reaction, streptolysin O (SLO) 1 was used to permeabilize sperm (17,18). SLO is a bacterial toxin that binds cholesterol in the plasma membrane, aggregates, and forms stable, relativity large pores, while leaving the inner organelle membranes intact (19).…”

Roles of the Na,K‐ATPase α4 isoform and the Na+/H+ exchanger in sperm motility

“…The percentage of motile sperm was determined using a hemocytometer to count duplicate samples in varied order. Motility was assessed by counting at least 200 sperm/sample, and sperm with flagellar movement and forward motion were scored as motile (Johnson et al, 1999;Wadi and Ahmad, 1999). The time of addition of nigericin, monensin, valinomycin, or the appropriate vehicle is indicated on each graph.…”