Amh secretion kinetic in doe kid

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Are artificial insemination centres ready to replace egg yolk by low density lipoproteins (LDL) in extenders for bull semen cryopreservation? : recent results review

Poster Abstract P50International audienceEgg yolk is widely used by artificial insemination (AI) centers for bull semen cryopreservation. Laboratory studies revealed that yolk contains granules that inhibit the respiration of spermatozoa and reduce their motility. Also, they interfere with microscopic observations. Egg yolk could also be a source of microbial contaminations. Therefore, there have been many attempts to find out which components in the egg yolk provide the cell protection with the aim to replace egg yolk with it’s cryoprotective fraction in order to prepare a clearly chemically defined extender without inconveniences. Many investigations showed that LDL is the cryoprotective fraction of egg yolk. However, prior to extending its use to AI centres, in vivo fertility studies were required. Semen was taken from three bulls and frozen-thawed in two extenders: the LDL extender and a standard tris-egg-yolk extender. The quality of the semen was assessed prior to AI: motility was assessed using Hamilton Thorne ceros 12, and the integrity of the plasma membrane was assessed using the hypo-osmotic swelling test. For the first time, pregnancies were obtained following the AI of cows in the field (n = 193) with semen that had been frozen-thawed in the LDL extender. No significant difference (p > 0.05) was detected between the success rates of AI between the semen that had been frozen-thawed in the LDL extender (59.2%) and the control extender, Tris- egg yolk (65.3%). In conclusion, the in vivo fertility of semen that has been frozen-thawed in the LDL extender is maintained since gestations are obtained following AI; the LDL extender is suitable for use by AI centres

Use of glutamine and low density lipoproteins isolated from egg yolk to improve buck semen freezing

International audienceTo improve the results obtained with a reference cryopreservation extender (control extender: Triladyl (R) + 20% (v/v) egg yolk + 6.4% (v/v) glycerol) for freezing caprine semen, glutamine was added to 18 split ejaculates at concentrations of 0, 20, 40, 80 and 120 mM (experiment 1). In experiment 2, glutamine was added to 18 split ejaculates at concentrations of 20, 25, 30, 35 and 40 mM. In the third experiment, the egg yolk was replaced with the low-density lipoprotein (LDL) fraction of egg yolk. The quality of frozen then thawed spermatozoa in each extender was compared using computer-assisted semen analysis. In experiment 1, glutamine at concentrations of 20 mM and 40 mM significantly improved sperm motility compared with the control extender. However, at 120 mM, a significant decrease in motility and velocity was observed. In experiment 2, motility, curvilinear velocity and amplitude of lateral head displacement (ALH) were improved in glutamine at 25 mM compared with the control. In experiment 3, 8% LDL and 25 mM glutamine significantly improved sperm motility, straight line velocity and ALH. In the fourth experiment, the quality of the previously defined freezing extender (Triladyl (R) + 8% (v/v) LDL + 25 mM glutamine + 6.4% (v/v) glycerol) was tested by comparing acrosome, tail membrane, plasma membrane and DNA integrity in 18 split ejaculates of frozen then thawed spermatozoa with spermatozoa that had been frozen then thawed in the control extender, and with spermatozoa from fresh, unfrozen sperm. The percentage of spermatozoa with intact acrosomes and tail membranes was significantly higher with the newly defined extender than that observed with the control extender. There was no significant difference in the percentage of spermatozoa with intact DNA between the frozen and fresh semen

Effect of equilibration time on the motility and functional integrity of canine spermatozoa frozen in three different extenders

International audienceThe present work aimed to assess the effect of equilibration time on post-thaw motility parameters of canine sperm frozen in three extenders: 6% low-density lipoproteins (LDL), 6% liposomes (LIPO), and 40% egg yolk plasma (EYP). A second experiment is aimed at evaluating the functional integrity of canine spermatozoa frozen in the three extenders at the best equilibration time found in the experiment one. In the first experiment, 20 ejaculates harvested from 7 dogs, were frozen in three extenders (LDL, LIPO, and EYP) after four equilibration times (30 min, 1 h, 3 h, and 6 h). The semen was evaluated after thawing using an image analyser (HT-IVOS 14.0). The 6 h equilibration time gave better results of motility and progressive motility in the three studied extenders. (LDL: 58.9% vs. 42.7%; LIPO: 54.4% vs. 31.9%; EYP: 55.4% vs 40.5% for motility 6 vs. 1 h). In the second experiment, 10 ejaculates taken from 6 dogs were frozen under the same conditions as the previous experiment, after 6 h equilibration time. The integrity parameters of the spermatozoal membrane (hypo-osmotic swelling test, and SYBR14/propidium Iodide staining), acrosome (FITC-Pisium sativum Aglutinin staining), and DNA (acridine orange staining) were evaluated at three different stages: post-dilution (TO), post-equilibration, and post-thawing. Post-thaw results were as follows: membrane integrity (HOSt: 62;6% vs 58% vs 64.4%; SYBR14/IP: 63.6% vs 57.9% vs 64.8%); acrosome integrity (FITC-PSA: 79.4% vs 74% vs 76.2%) and DNA integrity (Acridine-orange: 98.9% vs 98.5% vs 98.7%) respectively for LDL vs. LIPO vs. EYP. No significant difference existed between the extenders tested; thus 6%LIPO and 40%EYP could be good candidates for replacement of 6%LDL in the protection of canine sperm during the freeze-thaw process without altering motility and integrity parameters