Sperm concentration at freezing affects post-thaw quality and fertility of ram semen.

ITRA-ULE, INDEGSAL, University of León, León, Spain.
Theriogenology (Impact Factor: 1.85). 12/2011; 77(6):1111-8. DOI: 10.1016/j.theriogenology.2011.10.013
Source: PubMed

ABSTRACT We have investigated the effect of sperm concentration in the freezing doses 200, 400, 800, and 1600 × 10(6) mL(-1) on the post-thaw quality and fertility of ram semen. Semen was collected from seven adult Churra rams by artificial vagina during the breeding season. The semen was diluted in an extender (TES-Tris-fructose, 20% egg yolk, and 4% glycerol), to a final concentration of 200, 400, 800, or 1600 × 10(6) mL(-1) and frozen. Doses were analyzed post-thawing for motility (computer-assisted sperm analysis system [CASA]), viability, and acrosomal status (fluorescence probes propidium iodide [PI]/peanut agglutinin conjugated with fluorescein thiocyanate (PNA-FITC), SYBR-14/PI [Invitrogen; Barcelona, Spain] and YO-PRO-1/PI [Invitrogen; Barcelona, Spain]). Total motility and velocity were lower for 1600 × 10(6) mL(-1) doses, while progressive motility and viability were lower both for 800 and 1600 × 10(6) mL(-1). The proportion of viable spermatozoa showing increased membrane permeability (YO-PRO-1+) rose in 800 and 1200 × 10(6) mL(-1). Intrauterine inseminations were performed with the 200, 400, and 800 × 10(6) mL(-1) doses at a fixed sperm number (25 × 10(6) per uterine horn) in synchronized ewes. Fertility (lambing rate) was similar for semen frozen at 200 (57.5%) or 400 × 10(6) mL(-1) (54.4%), whereas it was significantly lower for 800 × 10(6) mL(-1) (45.5%). In conclusion, increasing sperm concentration in cryopreserved semen, at least at 800 × 10(6) mL(-1) and more, adversely affects the postthawing quality and fertility of ram semen.

1 Bookmark
  • [Show abstract] [Hide abstract]
    ABSTRACT: Variation among individuals is substantial for spermatozoa concentration in fresh milt in sea trout (Salmo trutta m. trutta L.). The objective of the present study was to examine effects of spermatozoa concentration in this species on subsequent cryopreservation success. Milt with high spermatozoa concentration was diluted with seminal plasma to obtain concentrations ranging between 6 and 24 × 10(9) mL(-1) with steps of 2 × 10(9) mL(-1). Diluted milts were cryopreserved in 0.25-mL straws with extender (0.3 M glucose) containing 10% methanol and 10 % (vol/vol) supplement of hen egg yolk. The dilution ratio was 1:3 (milt:cryomedium). Cryopreservation efficacies were assessed according to evaluation of motility of frozen/thawed spermatozoa and quantification of fertilizing ability. Percentage of motility of frozen/thawed spermatozoa was influenced by spermatozoa concentration in the cryomedium (P < 0.05). The highest motility was observed in samples with 3.0 to 4.0 × 10(9) spermatozoa per mL of cryomedium, which corresponds to 12 to 16 × 10(9) spermatozoa per mL in fresh milt. Higher sperm concentrations and lower sperm concentrations in cryomedium reduced the effectiveness of cryopreservation when compared with the optimum. Cryopreservation success measured according to fertilization rate was in agreement with results for motility of frozen/thawed spermatozoa, but the optimum could not be determined with statistical precision because of differences in fertilization rate among individual donor males. However, a significant positive correlation was found between postthaw motility and fertilization rate and between cryopreserved spermatozoa velocity and fertilization rate (P < 0.05). In sea trout, cryopreservation efficiency is influenced by spermatozoa concentration in cryomedium. Individual adjustment of the dilution ratio, based on initial spermatozoa density, is recommended in the freezing protocol. Maximum cryoresistance of the cell was obtained when spermatozoa concentration in cryomedium ranged from 3.0 to 4.0 × 10(9) mL(-1).
    Theriogenology 07/2013; · 1.85 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Seminal plasma purportedly plays a critical role in reproduction, but epididymal spermatozoa are capable of fertilisation following deposition in the uterus, calling into question the biological requirement of this substance. Through a combination of direct observation of spermatozoa in utero using probe-based Confocal Laser Endomicroscopy, in vivo assessment of sperm fertility and in vitro analysis of various sperm functional parameters, the present study investigated the role of seminal plasma in spermatozoa transit through the cervix of the ewe. Following deposition in the cervical os, epididymal spermatozoa previously exposed to seminal plasma displayed an enhanced ability to traverse the cervix as evidenced by both significantly higher pregnancy rates and numbers of spermatozoa observed at the utero-tubal junction when compared to epididymal spermatozoa not previously exposed to seminal plasma. The beneficial effect of seminal plasma on sperm transport was clearly localised to transit through the cervix as pregnancy rates of spermatozoa deposited directly into the uterus were unaffected by exposure to seminal plasma. This phenomenon was not explained by changes to sperm motion characteristics, as seminal plasma had no effect on the motility, kinematic parameters or mitochondrial membrane potential of spermatozoa. Rather, in vitro testing revealed that seminal plasma improved the ability of epididymal spermatozoa to penetrate cervical mucus recovered from ewes in oestrus. These results demonstrate that the survival and transport of ram spermatozoa through the cervix of the ewe is not linked to their motility or velocity but rather the presence of some cervical penetration trait conferred by exposure to seminal plasma.
    Reproduction (Cambridge, England) 08/2014; · 3.26 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Sperm is produced by the testis and mature in the epididymis. For having a successful conception, the fertilizing sperm should have functional competent membranes, intact acrosome, functional mitochondria and an intact haploid genome. The effects of genetic and environmental factors result in sperm vulnerability to damage in the process of spermatogenesis and maturation. In recent years, the feasibility of detecting sperm damage is enhanced through the advances in technologies like fluoscerent staining techniques assisted with fluorescence microscope, flow cytometry and computer analysis systems. Fluoscerent staining techniques involve the use of fluorescent dyes, either directly or indirectly for binding them with some ingredients of sperm and evaluating the damage of the structure or function of the sperm, i.e. membrane, acrosome, mitochondria, chromosome or DNA.
    Journal of reproduction & infertility. 01/2013; 14(3):120-125.


Available from
Jul 24, 2014