Importance of sperm genotype (indicus versus taurus) for fertility and embryonic development at elevated temperatures

Heat stress has negative effects on bovine reproduction, particularly for European breeds (Bos taurus taurus) that are less thermotolerant than zebu cattle (Bos taurus indicus). Here, the evidence that spermatozoa and oocyte both contribute to early embryonic resistance to heat shock is demonstrated. In addition, the use of reproductive biotechnologies to improve bovine thermotolerance, are outlined by comparing data from taurus, indicus and crossbred genotypes. (c) 2005 Published by Elsevier B.V.

Morphological and functional characteristics of chilled semen obtained from domestic feline epididymides (Felis catus)

The objective of this study was to determine morphological and functional characteristics of semen retrieved from the feline epididymis before and after cooling. Sixteen adult male cats were orchiectomized. The distal portion of the epididymis and proximal part of the deferent ducts were dissected and squeezed to obtain their content. After centrifugation, the supernatant was removed, sperm were resuspended in a 0.9 mL Tris-fructose-citric acid extender containing 20% egg yolk, aliquoted into three 0.3 mL samples, placed in a refrigerator (4.8 degrees C) and cooled (0.5 degrees C/min). Semen evaluations were performed on four occassions: immediately after epididymal sperm retrieval (TO), and at 24 h (T-1), 48 h (T-2) and 72 h (T-3) after cooling. on each occasion, progressive motility, vigor and sperm morphology were determined. Mean motility and vigor decreased (P < 0.05) between each successive examination. Although the majority of sperm cell damage occurred within the first 24 h, there was a decrease (P < 0.05) in mean percentage of morphologically normal sperm between To and each evaluated time (T-1, T-2, T-3) after cooling, due to an increase in coiled and bent sperm tails. Further studies are needed to evaluate the effects of cooling on the fertilizing capacity of cat epididymal spermatozoa in assisted reproduction programs. (c) 2006 Elsevier B.V. All rights reserved.

Influence of semen storage and cryoprotectant on post-thaw viability and fertility of stallion spermatozoa

The aim of the current study was to verify that stallion, spermatoza could be cooled for 24 hours and then frozen. In experiment I, one ejaculate from each of 13 stallions was used. Semen was collected and split into two parts; one part immediately frozen using standard cryo-preservation techniques and the other diluted, stored in an Equitainer for 24 hours, and then frozen. In experiment II, one ejaculate from each of 12 stallions was collected, diluted with Botu-Semen, and split into two parts: one cooled in an Equitainer and the other in Max-Semen Express without prior centrifugation. After 24 hours of cooling, the samples were centrifuged to remove seminal plasma and concentrate the sperm, and resuspended in Botu-Crio (R) extender containing on e of three cryoprotectant treatments (1% glycerol + 4% dimethylformamide, 1% glycerol + 4% dimethylacetamide and 1% glycerol + 4% methylformamide), maintained at 5 degrees C for 20 minutes, then frozen in nitrogen vapour. No difference was observed between the two cooling systems. The association of 1% glycerol and 4% methylformamide provided the best post-thaw progressive motility. For experiment III, two stallions were used for a fertility trial. Forty three inseminations were performed using 22 mares. No differences were seen in semen parameters and pregnancy rates when comparing the two freezing protocols (conventional and cooled/frozen). Pregnancy rates for conventional and cooled/frozen semen were, respectively, 72.7% and 82.3% (stallion A), and 40.0% and 50.0% (stallion B). We concluded that cooling equine-semen for 24 hours before freezing while maintaining sperm viability and fertility is possible.

Acrosomal ultrastructure of stallion spermatozoa cryopreserved with ethylene glycol using two packaging systems

The present experiments aimed to examine the substitution of glycerol (G) by ethylene glycol (E) as a cryoprotective agent for stallion spermatozoa. Two different ethylene glycol concentrations (5% and 10%) and also the association of glycerol (2%) and ethylene glycol (3%) (E/G) were studied (Experiment 1). In Experiment 2, two packing systems (0.5 x 4.0 ml) were evaluated using both cryoprotectors. In both experiments, the sperm membrane integrity after freezing was evaluated using transmission electron microscopy. The mean post-thaw motility was 34.25, 36.5, 29.25 and 34.75% for G5%, E5%, E10% and E/G, respectively. It was observed that the percentage of motile spermatozoa was significantly smaller (P<0.05) when semen was processed with E10%. A decrease in the acrosome integrity was observed in frozen thawed spermatozoa from all treated groups. It was observed that 28.0, 22.5, 25.5 and 22.5 % of the sperm cells had a normal acrosome following freezing with G5%, E5%, E10% and E/G, respectively. Undulation of the outer acrosomal membrane, acrosomal swelling and loss of acrosomal content density and homogeneity were the most evident ultrastructural alterations observed. In Experiment 2, the post-thaw motility was higher (P<0.05) for sperm frozen in 0.5 ml straws than in 4.0 mi straws, regardless of the cryoprotector used. The ultrastructural evaluation showed 26.7 and 16.0% of intact acrosomes for sperm frozen in 0.5 ml and 4.0 ml straws, respectively. We concluded that ethylene glycol has similar cryoprotective properties to glycerol and that utilisation of 0.5 ml straws improved the ability of horse sperm cells to withstand damage after the cryopreservation process.

Testicular thermoregulation in Bos indicus, crossbred and Bos taurus bulls: relationship with scrotal, testicular vascular cone and testicular morphology, and effects on semen quality and sperm production

Mechanisms of testicular thermoregulation, the relationship of scrotal, testicular vascular cone (TVC), and testicular morphology with thermoregulatory capability, and their effects on semen quality and sperm production were studied in 20 Bos indicus, 28 crossbred, and 26 Bos taurus bulls. The ratio of testicular artery length and volume to testicular volume were larger (P < 0.05) in B. indicus and crossbred bulls than in B. taurus bulls (1.03 and 0.94 cm/cm(2). versus 0.48 cm/cm(3); 0.034 and 0.047 ml/cm(3) versus 0.017 ml/cm(3), respectively). Testicular artery wall thickness (average 192.5, 229.0, and 290.0 mum, respectively) and arterial-venous blood distance in the TVC (average 330.5, 373.7, and 609.4 pm, respectively) were smallest in B. indicus, intermediary in crossbred, and greatest in B. taurus bulls (P < 0.05); the proximity between arterial and venous blood was consistent with the estimated decrease in arterial blood temperature after passage through the TVC (5.9, 5.0, and 2.9 degreesC, in B. indicus, crossbred, and B. taurus bulls, respectively). In crossbred and B. taurus bulls, there was a positive top-to-bottom scrotal temperature gradient and a negative testicular subtunic temperature gradient. However, in B. indicus bulls, both scrotal and testicular subtunic temperatures gradients were positive. Differences in the vascular arrangement, characteristics of the artery (e.g. wall thickness) or thickness of the tunica albuginea may have affected the testicular arterial blood and subtunic temperatures in B. indicus bulls. Better testicular thermoregulatory capability was associated with increased scrotal shape (pendulosity), testicular artery length and volume, and top-to-bottom gradient of the distance between the artery wall and the veins in the TVC. Increased semen quality was associated with increased testicular volume and scrotal subcutaneous (SQT) temperature gradient, and with decreased scrotal surface and testicular temperatures. Increased sperm production was associated with increased testicular artery volume, testicular volume, and SQT temperature gradient, and with decreased testicular artery wall thickness, scrotal circumference (SC), and scrotal surface, testicular subtunic, and epididymal temperatures. In conclusion, morphology of the TVC may contribute to the greater resistance of B. indicus bulls to high ambient temperatures by conferring a better testicular blood supply and by facilitating heat transfer between the testicular artery and veins. Testicular thermoregulation was associated with opposing scrotal and testicular subtunic temperatures gradients only in crossbred and B. taurus bulls. Scrotal, TVC, and testicular morphology influence testicular thermoregulatory capability and were associated with differences in semen quality and sperm production. (C) 2003 Elsevier B.V. All rights reserved.

Gonadic Sperm Reserve in Buffaloes Created under Tropical Conditions

The objective of the present study was quantifying the spermatic production per gram and daily rate of the testicular parenchyma. Testis of 12 crossbred Murrah buffaloes aged 24 to 48 months created under extensive conditions in the State of São Paulo/Brazil were analyzed. Animals were divided into groups based on the testicular shape (long and long-moderate) and testicular sides (right and left) and the studied parameters were: length and testicular width, weight and volume of the albuginea and mediastinum and net weight of the testicular parenchyma, gonadic sperm reserves, spermatic production/g of testicular parenchyma, daily spermatic production/g, total daily spermatic production, resulting into the values: 8.16 +/- 0.87 and 4.29 +/- 0.50 cm; 9.09 +/- 1.91 g and 8.77 +/- 1.88 mL; 0.97 +/- 0.39 g and 0.90 +/- 0.38 mL; 112.91 +/- 18.85 g; 14.32 x 10(9) +/- 0.15; 13.42 x 10(6) +/- 0.17; 27.40 x 10(6) +/- 0.35 and 2.92 x 10(9) +/- 0.30, registering no difference between right and left testis in relation to the parameters (P>0.05). There was no relation between testicular biometry and spermatic production (P>0.05). According to the obtained values, all animals were considered sexually mature and presented an efficient spermatic production per gram of testicular parenchyma and total daily production.

Osteopontin in Seminal Plasma and Sperm Membrane of Dogs

The objective of this work was to describe the presence of osteopontin (OPN) in canine seminal plasma and sperm membranes. A pool of seminal plasma and sperm membrane extract from 30 dogs was used. Polyacrylamide electrophoresis gels were performed and the bands were transferred to nitrocellulose paper and Western blot was undertaken using an antibody anti-OPN. Two and 12 bands were marked in the seminal plasma (77.2 and 15.6 kDa) and sperm membrane extracts (70.6-26.6 kDa), respectively. However, from 12 marked bands in the sperm membrane extract, only three (46.4, 37.7 and 36.5 kDa) were strongly marked. We conclude that, seminal plasma and sperm membranes from dogs contain different isoforms of OPN; yet, further studies will be necessary to determine their function in this species.