scholarly journals Effect of taurine on turkey (Meleagris gallopavo) spermatozoa viability and motility

2018 ◽  
Vol 63 (No. 4) ◽  
pp. 127-135 ◽  
Author(s):  
T. Slanina ◽  
M. Miškeje ◽  
F. Tirpák ◽  
M. Błaszczyk ◽  
R. Stawarz ◽  
...  
Keyword(s):  
Meleagris Gallopavo ◽  
Lower Percentage ◽  
Computer Assisted ◽  
Viability Assessment ◽  
Spermatozoa Motility ◽  
In Vitro Incubation ◽  
Time Periods ◽  
Casa System ◽  
Motility Parameters

The effect of taurine on the turkey spermatozoa motility and viability during the in vitro incubation was assessed. Experimental samples were prepared by diluting the raw semen in nine different concentrations of taurine – from 10 mg/ml to 0.078125 mg/ml. The motility parameters were evaluated by the CASA system (Computer Assisted Semen Analyser) using the program Sperm Vision<sup>®</sup> and for spermatozoa viability assessment the eosin-nigrosin staining was performed. Selected parameters were evaluated at six time periods: 0, 1, 2, 3, 4, and 5 h at 5°C and 41°C. At 5°C, a significantly lower percentage of motility and progressive motility was detected only in the samples with the highest concentration of taurine (10 mg/ml) at time 0 and 1. After 2 h of incubation a significant preventive effect of taurine on spermatozoa parameters was observed. The tendency of the taurine effect on motility parameters was different during the in vitro incubation at 41°C. Significantly lower values of motility parameters were detected in all experimental samples in comparison to the control after 5 h. The analysed concentrations of taurine did not significantly affect viability of turkey spermatozoa during all time periods. A higher percentage of dead spermatozoa were observed at 41°C (4.87–9.90%) if compared to 5°C (2.12–4.88%). The results indicated that the addition of taurine (from 2.5 to 7.5 mg/ml) to turkey spermatozoa positively affected the monitored spermatozoa parameters incubated at 5°C.

scholarly journals In vitro effect of ferrous sulphate on bovine spermatozoa motility parameters, viability and Annexin V-labeled membrane changes

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0257766
Author(s):  
Zuzana Knazicka ◽  
Hana Duranova ◽  
Veronika Fialkova ◽  
Michal Miskeje ◽  
Tomas Jambor ◽  
...  
Keyword(s):  
Cell Viability ◽  
Ferrous Sulphate ◽  
Annexin V ◽  
Computer Assisted ◽  
Spermatozoa Motility ◽  
Bovine Spermatozoa ◽  
High Concentrations ◽  
Vitro Effect ◽  
Motility Parameters

The aim of this study was to assess the dose- and time-dependent in vitro effects of ferrous sulphate (FeSO4.7H2O) on the motility parameters, viability, structural and functional activity of bovine spermatozoa. Spermatozoa motility parameters were determined after exposure to concentrations (3.90, 7.80, 15.60, 31.20, 62.50, 125, 250, 500 and 1000 μM) of FeSO4.7H2O using the SpermVisionTM CASA (Computer Assisted Semen Analyzer) system in different time periods. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay, and the Annexin V-Fluos was applied to detect the membrane integrity of spermatozoa. The initial spermatozoa motility showed increased average values at all experimental concentrations compared to the control group (culture medium without FeSO4.7H2O). After 2 h, FeSO4.7H2O stimulated the overall percentage of spermatozoa motility at the concentrations of ≤ 125 μM. However, experimental administration of 250 μM of FeSO4.7H2O significantly (P < 0.001) decreased the spermatozoa motility but had no negative effect on the cell viability (P < 0.05) (Time 2 h). The lowest viability was noted after the addition of ≥ 500 μM of FeSO4.7H2O (P < 0.001). The concentrations of ≤ 62.50 μM of FeSO4.7H2O markedly stimulated (P < 0.001) spermatozoa activity after 24 h of exposure, while at high concentrations of ≥ 500 μM of FeSO4.7H2O the overall percentage of spermatozoa motility was significantly inhibited (P < 0.001) and it elicited cytotoxic action. Fluorescence analysis confirmed that spermatozoa incubated with higher concentrations (≥ 500 μM) of FeSO4.7H2O displayed apoptotic changes, as detected in head membrane (acrosomal part) and mitochondrial portion of spermatozoa. Moreover, the highest concentration and the longest time of exposure (1000 μM of FeSO4.7H2O; Time 6 h) induced even necrotic alterations to spermatozoa. These results suggest that high concentrations of FeSO4.7H2O are able to induce toxic effects on the structure and function of spermatozoa, while low concentrations may have the positive effect on the fertilization potential of spermatozoa.

scholarly journals Caffeine strongly improves motility parameters of turkey spermatozoa with no effect on cell viability

2018 ◽  
Vol 66 (1) ◽  
pp. 137-150 ◽  
Author(s):  
Tomáš Slanina ◽  
Michal Miškeje ◽  
Filip Tirpák ◽  
Martyna Błaszczyk ◽  
Grzegorz Formicki ◽  
...  
Keyword(s):  
Incubation Temperature ◽  
Control Sample ◽  
Computer Assisted ◽  
Time Period ◽  
The Individual ◽  
The Impact ◽  
Casa System ◽  
Positive Effect ◽  
Motility Parameters

The purpose of this study was to evaluate the impact of caffeine on turkey spermatozoa during in vitro incubation. Experimental samples were prepared by diluting the raw semen with nine different concentrations of caffeine – from 0.078125 mg/mL to 10 mg/mL. The individual motility parameters were evaluated by the Computer Assisted Semen Analyser (CASA) system, and the viability of spermatozoa was evaluated using eosin-nigrosin staining. Selected parameters were recorded at six time periods: 0, 1, 2, 3, 4 and 5 h at 5 °C and 41 °C. A significantly higher motility and progressive motility of spermatozoa (P < 0.01 and P < 0.001, respectively) was detected in the samples containing caffeine ranging from 0.15625 to 7.5 mg/mL as compared to the control sample at 5 °C. At an incubation temperature of 41 °C the positive effect of caffeine on motility parameters was observed only at the beginning of incubation (at times 0 and 1). The tested caffeine concentrations showed no significant effect on the viability of turkey spermatozoa at any time period of incubation. A higher percentage of dead spermatozoa was observed for incubation at 41 °C (from 5.96% to 11.1%) in comparison to 5 °C (from 1.62% to 5.79%). The results suggest that caffeine can be used as a suitable component of turkey semen extenders and has the potential to improve fertility.

In vitroeffect of various cryoprotectants on the semen quality of endangered Oravka chicken

Zygote ◽  
2017 ◽  
Vol 26 (1) ◽  
pp. 33-39 ◽  
Author(s):  
Andrea Svoradová ◽  
Lenka Kuželová ◽  
Jaromír Vašíček ◽  
Andrej Baláži ◽  
Emília Hanusová ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Semen Quality ◽  
Semen Analysis ◽  
Lower Percentage ◽  
Computer Assisted ◽  
Spermatozoa Motility ◽  
Progressive Movement ◽  
Animal Genetic Resources

SummaryWe aimed to compare the effect of three different permeating cryoprotectants on the post-thaw spermatozoa quality. Pooled semen from Oravka cock line (n= 6) was diluted inKobidil+extender and frozen in cryoprotectant solutions containing 8% dimethylsulfoxide (DMSO), 8% ethylene glycol (EG) or 8% glycerol (GL) in liquid nitrogen vapours before being plunged into the liquid nitrogen. Spermatozoa motility parameters were assessedin vitroafter freezing–thawing by a computer-assisted semen analysis (CASA) system and viability status was examined using fluorescent probes. The lower percentage (P <0.05) of motile and progressively moving spermatozoa immediately after thawing were obtained in all experimental groups (DMSO, EG, GL) compared with the control. Significant (P< 0.05) differences in total motility and progressive movement between GL and DMSO, EG groups were observed. However, the higher number (P <0.05) of acrosome damaged spermatozoa was found in the DMSO and EG groups and no significant differences were observed in the GL group compared with the control. Differences (P< 0.05) between experimental groups and the control in the results of spermatozoa necrosis were observed. No significant differences in the percentage of apoptotic spermatozoa were found between control and experimental groups. However, significant differences (P< 0.05) in number of live and necrotic spermatozoa between GL and DMSO, EG groups were examined. The findings of the present study indicate that glycerol seems to be suitable for semen cryopreservation in the gene banks. In addition, fertility evaluationin vivois needed in order to evaluate the possible contribution for the bank of animal genetic resources.

scholarly journals The effect of resorcinol on bovine spermatozoa parameters in vitro

2020 ◽  
pp. 675-686
Author(s):  
M Massányi ◽  
M Halo ◽  
L Strapáková ◽  
T Slanina ◽  
P Ivanič ◽  
...  
Keyword(s):  
Cultivation Temperature ◽  
Negative Effects ◽  
Mtt Test ◽  
Progressive Motility ◽  
Time Period ◽  
Entire Duration ◽  
Bovine Spermatozoa ◽  
Time Periods ◽  
Motility Parameters

The goal of this study was to observe the effect of resorcinol on motility, viability and morphology of bovine spermatozoa. The semen was used from six randomly chosen breeding bulls. Ejaculate was diluted by different solutions of resorcinol in 1:40 ratio. Samples were divided into 7 groups with different concentrations of resorcinol (Control, RES1 – 4 mg/ml, RES2 – 2 mg/ml, RES3 – 1 mg/ml, RES4 – 0.5 mg/ml, RES5 – 0.25 mg/ml and RES6 – 0.125 mg/ml). Motility of spermatozoa was detected using CASA method at temperature of 37 °C in time periods 0, 1, 2, 3, 4 hours from the start of the experiment. Significant motility differences between all groups except control and RES6 with difference of 5.58 %, as well as between RES1 and RES2 groups with difference of 2.17 % were found. Progressive motility had the same significant differences. Spermatozoa viability (MTT test) decreased compared to control in all experimental groups during the entire duration of experiment. Observing morphologically changed spermatozoa, no significant changes were observed and a higher percentage of spermatozoa with separated flagellum in all experimental resorcinol groups compared to control were detected. Also, increased number of spermatozoa with broken flagellum, acrosomal changes and other morphological forms in the group with the highest concentration of resorcinol (RES1) were found. Results of our study clearly show negative effects on motility parameters of spermatozoa which depend on concentration, cultivation temperature and time period.

Effect of transfection and co-incubation of bovine sperm with exogenous DNA on sperm quality and functional parameters for its use in sperm-mediated gene transfer

Zygote ◽  
2016 ◽  
Vol 25 (1) ◽  
pp. 85-97 ◽  
Author(s):  
María Elena Arias ◽  
Esther Sánchez-Villalba ◽  
Andrea Delgado ◽  
Ricardo Felmer
Keyword(s):  
Gene Transfer ◽  
Sperm Quality ◽  
Computer Assisted ◽  
Exogenous Dna ◽  
Sperm Analysis ◽  
Functional Parameters ◽  
Transgenic Embryos ◽  
Fertilization Capacity ◽  
Motility Parameters

SummarySperm-mediated gene transfer (SMGT) is based on the capacity of sperm to bind exogenous DNA and transfer it into the oocyte during fertilization. In bovines, the progress of this technology has been slow due to the poor reproducibility and efficiency of the production of transgenic embryos. The aim of the present study was to evaluate the effects of different sperm transfection systems on the quality and functional parameters of sperm. Additionally, the ability of sperm to bind and incorporate exogenous DNA was assessed. These analyses were carried out by flow cytometry and confocal fluorescence microscopy, and motility parameters were also evaluated by computer-assisted sperm analysis (CASA). Transfection was carried out using complexes of plasmid DNA with Lipofectamine, SuperFect and TurboFect for 0.5, 1, 2 or 4 h. The results showed that all of the transfection treatments promoted sperm binding and incorporation of exogenous DNA, similar to sperm incorporation of DNA alone, without affecting the viability. Nevertheless, the treatments and incubation times significantly affected the motility parameters, although no effect on the integrity of DNA or the levels of reactive oxygen species (ROS) was observed. Additionally, we observed that transfection using SuperFect and TurboFect negatively affected the acrosome integrity, and TurboFect affected the mitochondrial membrane potential of sperm. In conclusion, we demonstrated binding and incorporation of exogenous DNA by sperm after transfection and confirmed the capacity of sperm to spontaneously incorporate exogenous DNA. These findings will allow the establishment of the most appropriate method [intracytoplasmic sperm injection (ICSI) orin vitrofertilization (IVF)] of generating transgenic embryos via SMGT based on the fertilization capacity of transfected sperm.

94 ADDITION OF REDUCED GLUTATHIONE TO THAWING MEDIUM IMPROVED THE SPERM MOTILITY AND REDUCED ROS GENERATION IN FROZEN OVINE AND CAPRINE SPERMATOZOA

2006 ◽  
Vol 18 (2) ◽  
pp. 155 ◽  
Author(s):  
J. C. Gardón ◽  
J. A. Rodriquez ◽  
J. Gadea
Keyword(s):  
Sperm Motility ◽  
Reduced Glutathione ◽  
Ovis Aries ◽  
Capra Hircus ◽  
Ros Generation ◽  
Computer Assisted ◽  
Sperm Analysis ◽  
Spermatozoa Motility ◽  
Curvilinear Trajectory ◽  
Motility Parameters

The processes of cooling and freezing/thawing produce physical and chemical stress on the sperm membrane, and this stress is associated with oxidative stress and reactive oxygen species (ROS) generation that further reduce sperm viability and fertilizing ability. It is known that the process of freezing is associated with a significant reduction of the intracellular reduced glutathione (GSH) content. The aim of these experiments was to investigate the effects of addition of GSH to thawing extenders on motility parameters and ROS generation in frozen-thawed ovine and caprine spermatozoa. Frozen spermatozoa from eight rams (Ovis aries) and eight bucks (Capra hircus) (generously provided by Ovigen, Zamora, Spain) were thawed in a water bath at 37�C for 30 s and resuspended in sperm-TALP medium (Parrish et al. 1986 Theriogenology 25, 591-600) without (control) and with addition of 1 mM or 5 mM GSH. After 30 min of incubation at 37�C, sperm motility was evaluated using a computer-assisted sperm analysis (CASA) system (SCA, Microptic, Barcelona, Spain). The recorded parameters of motility were: % total, % progressive, curvilinear velocity, straight-line velocity, average path velocity, linearity of the curvilinear trajectory, straightness, amplitude of lateral head displacement, wobble of the curvilinear trajectory and beat cross frequency. Another set of sperm samples was incubated in the presence of (0.7 �M) 22,72-dichlorodihydrofluorescein diacetate (Gadea et al. 2005 J. Androl. 26, 396-404) to estimate production of ROS by flow cytometry. Data were analyzed by two-way ANOVA, considering the specific sperm treatment (GSH addition) and the males as the main variables. In ram frozen spermatozoa, all of the motility parameters were significantly improved when the medium was supplemented with GSH (P < 0.01) with even better results when 5 mM GSH was used. As an example, progresive motility increased from 31.16% (control) to 39.17 and 43.97%, respectively, for 1 and 5 mM GSH. Despite of the male effect detected (P < 0.01), all eight rams studied presented a similar pattern (interaction P > 0.05). The generation of ROS was significantly reduced when GSH was added (6.23a for control vs. 5.32b and 3.85c for 1 and 5 mM, respectively; P < 0.01). In buck frozen spermatozoa, % motility and progressive motility were significantly higher in GSH groups than in the control (P < 0.01), with no differences between 1 and 5 mM GSH. However, for the other motility parameters, the differences were not significant, which probably could be related to differences in the pattern shown by different animals (interaction of buck by treatment P < 0.05). ROS generation was significantly reduced when GSH was added (7.50a for control vs. 4.32b and 2.70b for 1 and 5 mM, respectively; P < 0.01). The addition of GSH to the thawing medium had a positive influence on the parameters studied in both species, increasing the motility patterns and reducing the ROS generation. In conclusion, we can assume that the addition of reduced glutathione to the thawing medium exerts a protective effect on spermatozoa functionality. This work was supported by AGL-2003-03144.

scholarly journals IN VITRO EFFECTS OF ISOQUERCITRIN ON SELECTED VITALITY MARKERS OF BOVINE SPERMATOZOA

2019 ◽  
Vol 1 (2) ◽  
pp. 18-23
Author(s):  
Filip Benko ◽  
Hana Greifová ◽  
Eva Tvrdá
Keyword(s):  
Physiological Saline ◽  
Biological Properties ◽  
Computer Assisted ◽  
Movement Activity ◽  
Spermatozoa Motility ◽  
Physiological Saline Solution ◽  
Bovine Spermatozoa ◽  
In Vitro Effects

The aim of this study was to evaluate the dose- and time-dependent in vitroeffects of isoquercitrin (ISO), a natural flavonoid with numerous biological properties on bovine spermatozoa during three different time periods (0 h, 2 h, 24 h). Bovine semen samples were diluted and cultivated in physiological saline solution containing 0.5% DMSO together with 200, 100, 50, 10, 5 and 1 μmol/L ISO. Spermatozoa motility was measured using the HTM IVOS CASA (Computer Assisted Semen Analyzer) system. The viability of spermatozoa was assessed by the metabolic (MTT) assay, production of superoxide radicals was quantified using the nitroblue tetrazolium (NBT) test, and chemiluminescence was used to evaluate the generation of reactive oxygen species (ROS). The results of the movement activity showed a significant increase in the motility during long term cultivation in case of concentrationsranging between 5 and 50 μmol/L ISO (P<0.05; 24 h). At the same time, supplementation of several concentrations of ISO led to a significant preservation of the cell viability (P<0.05 in the case of 50 μmol/L, P<0.01 with respect to 1 and 5 μmol/L, and P<0.001 in relation to 10 μmol/L; 24 h). ISO addition at 10 and 50 μmol/L also provided a significantly higher protection against superoxide (P<0.05) and ROS (P<0.001) overgeneration after a 24 h cultivation. We may suggest that supplementation of ISO to bovine spermatozoa, particularly at concentrations ranging between 10 and 50 μmol/L, may offer protection to the motility, viability and oxidative status of the sperm cells, particularly notable at 24 h.

12 COMPUTER-ASSISTED SPERM ANALYSIS OF FRESH EPIDIDYMAL CAT SPERMATOZOA AND THE IMPACT OF COOLED STORAGE (4°C) ON SPERM QUALITY

2008 ◽  
Vol 20 (1) ◽  
pp. 86
Author(s):  
M. Filliers ◽  
T. Rijsselaere ◽  
P. Bossaert ◽  
V. De Causmaecker ◽  
J. Dewulf ◽  
...  
Keyword(s):  
Reference Values ◽  
Sperm Quality ◽  
Membrane Integrity ◽  
Computer Assisted ◽  
Epididymal Sperm ◽  
Sperm Analysis ◽  
Computer Assisted Sperm Analysis ◽  
The Impact ◽  
Motility Parameters

Feline epididymal sperm is commonly used for in vitro fertilization. It also yields the opportunity to conserve genetic material from valuable males that suddenly die. Epididymal sperm quality parameters vary considerably among laboratories, implicating the need for objective evaluation methods. The aim of the present study was to describe reference values of computer-assisted sperm analysis (CASA) parameters of fresh epididymal cat sperm and to assess the effect of prolonged cooled storage (4�C) on various sample characteristics. Epididymides obtained from tomcats after routine orchiectomy (2–4 pairs/replicate) were sliced to release spermatozoa. The sperm suspension was placed on a 2-layer gradient and, after centrifugation, the sperm pellet was recovered. In Experiment 1 (20 replicates), sperm motility parameters were assessed immediately after retrieval (T0) using the Hamilton Thorne analyzer Ceros 12.1 (HTR; Hamilton Thorne Biosciences, Beverly, MA, USA). In Experiment 2, fresh (T0) sperm samples (4 replicates) were evaluated for motility parameters (HTR), acrosomal status (FITC-Pisum sativum agglutinin staining), morphology (eosin/nigrosin (E/N) staining), and membrane integrity (E/N and SYBR�-14-propidium iodide staining; Molecular Probes, Inc., Eugene, OR, USA). After addition (1:2) of a Tris-glucose-citrate diluent containing 20% egg yolk, samples were cooled and reassessed on Days 1 (T1), 3 (T3), 5 (T5), 7 (T7), and 10 (T10). Results were analyzed in a mixed linear model, with replicate as random factor and time as fixed effect (S-PLUS 7.0; Insightful Corp., Seattle, WA, USA). Results of Experiment 1 were as follows (mean � SD): motility (MOT): 80.8% � 23.5; progressive motility (PMOT): 69.9% � 23.2; velocity average pathway (VAP): 98.7 µm s–1 � 24.2; velocity straight line (VSL): 89.3 µm s–1 � 25.4; velocity curved line (VCL): 134.8 µm s–1 � 31.9; amplitude lateral head (ALH): 4.3 µm � 2.0; beat cross frequency (BCF): 34.6 Hz � 7.0; and straightness (STR): 89.6% � 6.6. In Experiment 2, MOT, PMOT, VAP, VSL, VCL, BCF, and the percentage of normal spermatozoa showed a decrease over time (P < 0.05) compared to fresh samples, starting from T1, T3, T5, T7, T5, T3, and T1, respectively. In contrast, STR, ALH, membrane integrity, and the percentage of acrosome-intact spermatozoa were not affected (P > 0.05) by cooled storage. To summarize, we have presented a set of reference values for CASA-parameters of fresh, epididymal cat spermatozoa. Cooled storage impaired most motility parameters and lowered the percentage of normal spermatozoa, but did not influence membrane integrity or acrosomal status. The effect of cooled storage on DNA fragmentation of sperm and its subsequent influence on in vitro embryo development require further investigation.

88 SELECTION OF A CRYOSURVIVAL SPERM POPULATION DOES NOT IMPROVE THE IN VITRO PENETRATING ABILITY OF FROZEN - THAWED BOAR SPERMATOZOA

2008 ◽  
Vol 20 (1) ◽  
pp. 124
Author(s):  
M. L. Perals ◽  
M. A. Gil ◽  
E. M. Garcia ◽  
J. Sanchez-Osorio ◽  
J. M. Vázquez ◽  
...  
Keyword(s):  
Sperm Quality ◽  
Membrane Integrity ◽  
Molecular Probes ◽  
Computer Assisted ◽  
Immature Oocytes ◽  
Sperm Analysis ◽  
Boar Spermatozoa ◽  
Casa System ◽  
Selection Of

Boars can be classified as good or bad sperm freezers according to their sperm cryosurvival. Different sperm selection techniques, such as PureSperm� (PS; MidAtlantic Diagnostics, Inc., Mount Laurel, NJ, USA), have been developed to improve functional competence of spermatozoa. The aim of this experimental study was to assess the ability of PS for improving the in vitro penetrating ability of frozen–thawed boar spermatozoa from good and bad sperm freezers. The sperm-rich fractions from two boars, good (Boar A) and bad (Boar B) freezers, were extended in a lactose/eggyolk/ glycerol/Equex Stem (Noba Chemical Sales, Inc., Scituate, ME, USA) mixture (1 � 109 sperm mL–1), dispensed into 0.5-mL straws, and frozen using a programmable cell freezer. After thawing (1.200�C min–1), semen from each boar was split into two aliquots of 500 µL. One aliquot was used as the control. The second was placed into a tube of PS gradient (90%/45%) and centrifuged at 425g for 20 min; the pellet re-suspended in 1 mL of BTS and re-centrifuged at 320g for 10 min (PS sample). Control and PS samples were diluted in supplemented TCM-199 (TCMm; Roca et al. 1998 Reprod. Fertil. Dev. 10, 479–485) at 200 � 106 sperm mL–1. Sperm survival (SV) was assessed afterTCMm dilution according to progressive sperm motility (PSM, %) using a computer-assisted sperm analysis (CASA) system (ISAS�), and plasma and acrosome membrane integrity (PMI; %) by flow cytometry (SYBR�-14/PE-PNA/PI; Molecular Probes, Leiden, The Netherlands). A homologous in vitro penetration (hIVP) assay, using immature oocytes (20 oocytes/2 mL TCMm supplemented with caffeine and calcium lactate), was used to assess sperm penetrating ability (Martinez et al. 1993 Theriogenology 40, 547–557). A total of 960 immature oocytes were inseminated (200 � 103 sperm/oocyte) in 3 batches. After 18 h of co-incubation at 39�C under 5% CO2 in air, the oocytes were washed, mounted on slides, fixed with ethanol:acetic acid (3:1, v/v) for 48 h, stained with 1% lacmoid, and examined under a phase contrast microscope (�400). Oocytes with swollen or unswollen heads of sperm found in the vitellus were considered as penetrated. Sperm penetrability ability (SPA) was assessed according to penetration rate (PR) and the mean number of sperm per oocyte (S/O). Data were analyzed using a PROMIXED model and expressed as mean � SEM. Boar A showed better (P ≤ 0.01) results for both SV and SPA parameters than boar B, independent of sperm treatment. PureSperm improved (P ≤ 0.05) PSM and PMI in both boar A (control v. PS: 48.0 � 5.8 v. 66.5 � 3.6 and 63.1 � 7.7 v. 88.4 � 1.3, respectively) and boar B (12.3 � 1.2 v. 22.2 � 3.7 and 44.3 � 3.5 v. 58.7 � 7.0, respectively). However, no differences (P ≥ 0.05) were observed in PR and S/O in either boar A (71.2 � 3.4 v. 78.3 � 3.1 and 5.0 � 0.4 v. 5.2 � 0.4, respectively) or boar B (34.3 � 3.6 v. 37.3 � 3.9 and 1.5 � 0.1 v. 1.5 � 0.1, respectively). In conclusion, under our laboratory conditions, PureSperm selection improves sperm quality but not in vitro penetrating ability of frozen–thawed spermatozoa of both good and bad sperm freezers. This work was supported by CICYT (AGF2005-00760), Madrid, Spain.

84 COMPUTER-ASSISTED SPERM ANALYSIS OF FROZEN SPERM MOTILITY AFTER REPEATED EXPOSURES TO ROOM TEMPERATURE

2010 ◽  
Vol 22 (1) ◽  
pp. 200
Author(s):  
A. Alvaro Garcia Guerra ◽  
G. M. Brogliatti
Keyword(s):  
Liquid Nitrogen ◽  
Sperm Motility ◽  
Room Temperature ◽  
Cell Damage ◽  
Beat Frequency ◽  
Computer Assisted ◽  
Initial Exposure ◽  
Frozen Semen ◽  
Casa System ◽  
Motility Parameters

The key factorin long-term cryopreservation is the very low temperature of liquid nitrogen. Several studies suggest temperatures should be maintained at -130°C or less to avoid cell damage. Damage due to initial exposure may not be overt; however, after repeated exposures a reduction in postthaw viability may become evident (Barth A 1991 Proc. 10th Annu. Conv. Am. ET Assoc, 20-26). The CASA system provides an opportunity to assess multiple motility characteristics on a semen sample objectively and with high repeatability. An experiment was designed to evaluate the effect that repeated exposure of frozen semen in 0.5-mL straws during 15 s to room temperature produces on motility characteristics assessed by CASA system. Groups were formed according to the number of exposures per straw; groups were as follows: 0, 3, 5, and 10 times of exposure during 15 s. Thirty-two ejaculates from different bulls (15 Angus, 3 Hereford, 8 Brangus, 3 others) were diluted using a chemically semi-defined media (Andromed, Minitub, Germany) and frozen in an automatic freezer (Digicool, IMV, Paillette Crista, France). Four frozen straws per bull were used, one for each group. Straws were exposed to a room temperature (15°C ± 1.28) and then placed back into liquid nitrogen. Semen thawing was conduced in a water bath at 37°C during 1 min. Motility characteristics were evaluated by the IVOS Sperm Analyzer (Hamilton Thorne Research). Two chambers of 20 μm depth and 5 fields per chamber were analyzed (30 frames/0.5 s for each field). Seven motility parameters were evaluated: % of motile sperm; % of progressive sperm; VAP (path velocity, μms-1); VCL (track speed, μm/s); ALH (lateral amplitude, μm); BCF (beat frequency, Hz); and LIN (linearity, %). The Kruskal Wallis test was used to compare variables among groups, and results are shown in Table 1. The average temperature inside the straw after 15 s of exposure was of -122.6°C. No difference (P > 0.05) was found among the groups for any of the 7 motility parameters. In conclusion, sperm motility seems not to be affected if straws are exposed up to 10 times during 15 s to room temperature. More research should be done to test higher room temperatures and pregnancy rates after AI. Table 1.CASA parameters of frozen sperm after different numbers of exposures at 15°C

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