Ultrastructure evaluation of goat spermatozoa after freezing in a skim milk-based extender with Trolox supplementation

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Abstract

The aim of this work was to evaluate the in vitro effect of adding Trolox in freezing extender for goat semen. Ejaculates from five bucks were evaluated, and when approved, the samples were pooled, diluted according to experimental groups [Trolox 0 (control), 30, 60 and 120 nmol ml−1] and frozen in an automated system. Thawed samples (37 °C/30 s) were evaluated for plasma membrane (PMi) and acrosome integrity (Aci), mitochondrial membrane potential (MMP) and sperm kinematics by CASA system. Spermatozoa ultrastructure was evaluated in fresh and post-thawed semen. No significant difference (P > 0.05) was observed among control and Trolox groups in the analyses of PMi, Aci, MMP and CASA in goat spermatozoa after thawing. Samples of 60 and 120 nmol ml−1 Trolox groups had a higher percentage of cells that had intact plasma membranes in spermatozoa head than in the other groups, although they did not differ (P > 0.05) before being frozen. A higher percentage (P < 0.05) of spermatozoa with intact mitochondria was observed in fresh semen, control and Trolox 60 nmol ml−1 groups than in the other groups. Addition of Trolox to skim milk extender at 60 nmol ml−1 ultrastructurally preserves the plasma membrane and mitochondrial sheath integrity in goat spermatozoa after cryopreservation.

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