Cell Subpopulation‐related Volumetric Parameters: a Complementary Tool of the Modified Hypo‐osmotic Swelling Test on Model of Boar Spermatozoa

It is a general property of the intact animal cell to swell rapidly in response to hypo‐osmotic conditions. The modified hypo‐osmotic swelling test (HOS‐test) is an indicative test to evaluate the integrity of the plasma membrane by means of an electronic cell counter, based on the relative increase of the cell volume in response to hypo‐osmotic conditions. In this study the relationships between the osmotically induced changes of the cell volume of boar spermatozoa as determined by cell counter and the integrity of the membrane as determined by propidium iodide staining (PI) were studied. Boar sperm cell volume distributions were measured under iso‐osmotic (300 mosmolar) conditions and after a hypo‐osmotic stress (150 mosmolar). The relative volume shift of mean and modal volume were calculated as a proportion coefficient of modal and mean values of the cell volume distributions by transition from iso‐osmotic to hypo‐osmotic conditions. The volumetric parameters related to the different cell subpopulations were derived from the different peaks of cell volume distributions. PI‐staining techniques were used for comparison. The values of the volume shift and of derived percentages of the osmotically inactive cells were correlated negatively and positively, respectively (p < 0.05) with the percentage of the PI‐stained cells. This correlation indicates that a relationship exists between membrane functions of the different cell compartments (sperm head and tail) due to the circumstance that the increase of the cell volume in the HOS‐test is associated with the morphological changes in the tail and the PI‐staining is associated with the membrane integrity and permeability of the head region. The advantage of computer‐assisted volume measurement is that a large number of cells (5000–50 000 spermatozoa) can be measured and evaluated during one procedure and in a very short time. The relative volume shift is a quantitative continuous parameter characterizing the osmotic reactivity and membrane functional competence of a cell population and of subpopulations within one ejaculate. This parameter could be useful to evaluate membrane functional competence rapidly and sensitively.