Biology of Reproduction, Vol 14, 381-397, Copyright © 1976 by Society for the Study of Reproduction

Use of Spin Labels and Electron Spin Resonance Spectroscopy to Characterize Membranes of Bovine Sperm: Effect of Butylated Hydroxytoluene and Cold Shock

¹ Departments of Biochemistry-Biophysics, Microbiology-Cell Biology, and Dairy Breeding Research Center, Pennsylvania State University, University Park, Pennsylvania

Spin label probes were used in conjunction with measurements of metabolic rate and electron microscopy to characterize bovine sperm membranes. Aqueous compartments, membrane hydrocarbon zones and lipid:water interfaces were studied separately using appropriate spin labels. For sperm suspended in aqueous medium, the cold shock associated with rapid cooling from room temperature to 0° increased membrane permeability. This membrane damage was readily detected using spin labels but was not detected using thin section electron microscopy. This change was prevented by the addition of butylated hydroxy toluene (BHT). BHT provided partial protection against further damage caused by freezing sperm on solid CO₂. ESR techniques provide a rapid means to quantify the changes in sperm membranes occurring during the epididymal maturation of sperm and subsequent events within the female tract leading to fertilization. The technique also could be used to assess damage to sperm, ova or embryos during preparation for storage in cryoprotective diluents.

Note:
ACKNOWLEDGMENTS Sandra Hay provided assistance with the metabolic measurements. Financial support was provided by NIH Grant HD-05859, ERDA Grant AT(11-1)-3419 and ADK is grateful to NIH award GMS 1-K4-GM70593-03.

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