Improvement in Semen Conservation of the Indigenous Czech Endangered Old Kladruber Horse: Special Focus on the Type of Extender and Packaging System

Abstract The aim of our study was to investigate the effect of two freezing extenders and two packaging systems on motility, plasma membrane (PM) integrity, and the apoptotic status of frozen-thawed (F-T) spermatozoa of the endangered Old Kladruber stallions. The collected semen (n = 6 stallions, three collections each) was diluted either with Gent or Lactose-EDTA (Lact) extender. Two aliquots of semen from each collection diluted in this way were prepared and then loaded into 5-mL aluminum tubes or 0.5-mL plastic straws. After thawing and then at 15 minutes intervals within 1 hour, the samples were analyzed for motility (CASA), PM integrity (CFDA/PI), and apoptotic changes of the spermatozoa (Yo-Pro-1/PI). Using Gent, the samples exhibited higher motility, kinematic parameters, higher representation of spermatozoa in medium, and fast subpopulation, as well as more spermatozoa with intact PMs and fewer spermatozoa with apoptotic changes compared with Lact extender ( P P P