Evaluation of plasma membrane stability by detergent-induced rupture of osmotically swollen sperm

Abstract

A general assay for plasma membrane stability was developed and tested. Osmotically swollen spermatozoa were ruptured with detergents and their volume distribution was monitored with resistance pulse spectroscopy. The extent of cell breakage was determined and expressed as [D]50, the concentration of detergent necessary to lyse 50% of the initially intact cells. Preliminary experiments established the degree to which spermatozoa could be swollen without lysis (no detergent) and the ability of the method to detect known mixtures of intact and membrane disrupted spermatozoa. [D]50 values were determined for caput (immature) and cauda (mature) ram epididymal spermatozoa with four detergents (cetyltrimethylammonium bromide, sodium dodecylsulfate, Zwittergent 3–14, and sodium deoxycholate). [D]50 values for caput spermatozoa were higher than those for cauda spermatozoa (P < 0.05) for all detergents but cetyltrimethylammonium bromide. Theese changes are consistent with a qualitative model of membrane structure and stability based on lipid shape and composition and with the compositional changes known to occur during epididymal maturation. Additional studies using rooster spermatozoa established that a typical cryopreservation protocol leaves the surviving spermatozoa with membranes with greater sensitivity to detergent-induced stress. Since osmotic swelling has been microscopically localized to the tail plasma membrane, the changes in membrane stability can be assigned specifically to that region.