Fatty acid composition and biophysical characteristics of the cell membrane of feline spermatozoa

Abstract

Sperm membrane composition and biophysical characteristics play a pivotal role in many physiological processes (i.e. sperm motility, capacitation, acrosome reaction and fusion with the oocyte) as well as in semen processing (e.g. cryopreservation). The aim of this study was to characterize the fatty acid content and biophysical characteristics (anisotropy, generalized polarization) of the cell membrane of domestic cat spermatozoa. Semen was collected from 34 adult male cats by urethral catheterization. After a basic semen evaluation, the fatty acid content of some of the samples (n = 11) was evaluated by gas chromatography. Samples from other individuals (n = 23) were subjected to biophysical analysis: membrane anisotropy (which is inversely proportional to membrane fluidity) and generalized polarization (describing lipid order); both measured by fluorimetry at three temperature points: 38 °C, 25 °C and 5 °C. Spermatozoa from some samples (n = 10) were cryopreserved in TRIS egg yolk-glycerol extender and underwent the same biophysical analysis after thawing. Most fatty acids in feline spermatozoa were saturated (69.76 ± 24.45%), whereas the polyunsaturated fatty acid (PUFA) content was relatively low (6.12 ± 5.80%). Lowering the temperature caused a significant decrease in membrane fluidity and an increase in generalized polarization in fresh spermatozoa, and these effects were even more pronounced following cryopreservation. Anisotropy at 38 °C in fresh samples showed strong positive correlations with viability and motility parameters after thawing. In summary, feline spermatozoa are characterized by a very low PUFA content and a low ratio of unsaturated:saturated fatty acids, which may contribute to low oxidative stress. Cryopreservation alters the structure of the sperm membrane, increasing the fluidity of the hydrophobic portion of the bilayer and the lipid order in the hydrophilic portion. Because lower membrane fluidity in fresh semen was linked with better viability and motility after cryopreservation, this parameter may be considered an important factor in determination of sperm cryoresistance.

Keywords: Domestic cat; Fatty acids; Membrane fluidity; Membrane generalized potential; Semen.

Figure 1

Anisotropy of sperm membranes measured at three temperature points: 38 °C (A 38C), 25 °C (A 25C), and 5 °C (A 5C), in fresh (n = 15) and frozen-thawed (n = 18) feline spermatozoa. The upper and lower sides of the box represent lower and upper quartiles, the vertical line represents the median, the cross represents the mean. The whiskers represent minimal and maximal value, circles represent outliers. Asterisks indicate significant differences: *p < 0.05, **p < 0.02, ***p < 0.001.

Figure 2

Generalized polarization of sperm membranes measured at three temperature points: 38 °C (GP 38C), 25 °C (GP 25C), and 5 °C (GP 5C), in fresh (n = 15) and frozen-thawed (n = 18) feline spermatozoa. The upper and lower sides of the box represent lower and upper quartiles, the vertical line represents the median, the cross represents the mean. The whiskers represent minimal and maximal value, circles represent outliers. Asterisks indicate significant differences: *p < 0.05, **p < 0.02, ***p < 0.001, ns—not significant.

Figure 3

Experimental design. *ten samples were both analyzed fresh and underwent cryopreservation, five were only analyzed fresh and 8 were only cryopreserved. CASA computer assisted sperm analysis.