Dimethylacetamide in Rooster Semen Cryopreservation.

BACKGROUND: Sperm cryopreservation of cockerels is a major challenge, and so far there is no adequate information to enable commercial use of frozen semen. OBJECTIVE: To test the toxicity of dimethylacetamide (DMA). MATERIALS AND METHODS: DMA was added at 3%, 6%, 9% and 12% to the freezing diluent, and maintained for equilibration with the semen sample for 1 min, 3 min, 5 min, 7 min and 9 min prior to freezing. Thawed semen was evaluated for kinetic characteristics by computer-assisted semen analysis (CASA) and for structural and functional properties by flow cytometry (plasma membrane rupture, mitochondrial functionality and plasma membrane functionality). RESULTS AND CONCLUSION: The addition of 6% DMA for 3-min equilibration resulted in the highest total and progressive motility, 42.0% and 36.9%, respectively. The point of intersection between a good protection and low plasma membrane rupture was obtained with the addition of 6% of DMA for 3-min equilibration with the rooster semen.

Effect of egg yolk plasma on dog sperm cryopreservation.

This study evaluated the quality of frozen-thawed dog spermatozoon after the inclusion of egg yolk plasma (EYP) instead of whole egg yolk (EY) in the cryopreservation extender and after distinct periods of exposure to EYP. Seven mongrel dogs were used as sperm donors, and EYP was obtained by centrifugation. In Experiment 1, post-thawing sperm motility (MOT) and integrity of membrane (INT) and acrosome (ACR) were superior for spermatozoon extended with 20% EYP T2 than with 20% EY (P < 0.05), although normal sperm morphology (MOR) did not differ (P > 0.05). In Experiment 2, after ejaculates extended with 20% EYP were cooled at 5°C for 2, 6 and 10 h before freezing, MOT, INT and ACR were similar among periods (P > 0.05). Thus, dog spermatozoon extended with 20% EYP can be kept cooled for up to 10 h prior to freezing, achieving post-thawing quality greater than that obtained with the inclusion of EY in freezing extenders.

Produção e preservação do sêmen de galos / Production and preservation of rooster semen

A seleção de reprodutores capazes de transmitir características de valor econômico para a progênie é fundamental para o sucesso da indústria de aves. Nesse sentido, os machos devem produzir sêmen de boa qualidade, com volume e concentração adequados. O ejaculado de galos é composto por uma grande quantidade de células espermáticas suspensas em um pequeno volume de plasma seminal, proveniente dos túbulos seminíferos. Por esse motivo, a adição de diluentes ao sêmen é uma prática de rotina nos estabelecimentos que realizam inseminação artificial, com a finalidade de aumentar o aproveitamento do ejaculado. A diluição também permite manter a qualidade do sêmen por várias horas, quando resfriado, ou por períodos mais longos, quando congelado. Para a manutenção da capacidade fertilizante do sêmen resfriado, substratos energéticos e agentes tampões devem ser incluídos no diluente. Para diminuir os danos sofridos pelos espermatozoides durante o congelamento, crioprotetores são adicionados ao sêmen. Entre eles, a dimetilacetamida vem sendo o mais utilizado nos protocolos para o congelamento de sêmen de galos nos últimos anos. A modulação lipídica do espermatozoide, por meio de dietas ou por adição de gema de ovo ou de seus derivados ao diluente, também auxilia na preservação do sêmen. Os benefícios são ainda maiores quando antioxidantes são incluídos.

Selecting breeders capable of transmit characteristics of economic value to the offspring is fundamental to poultry industry success. In this sense, males must produce good quality semen, with appropriate volume and concentration. Rooster ejaculates are composed of a great amount of sperm cells suspended in a small volume of seminal plasma, originated in the seminiferous tubules. For this reason, adding diluents to semen is a common practice in artificial insemination establishments, aiming to increase ejaculate use. Dilution also allows semen quality maintenance for hours, when refrigerated, or for longer periods, when frozen. To keep fertilizing capacity of refrigerated sperm, energetic substrates and buffer agents must be included into the diluents. To decrease sperm damage during freezing, cryoprotectants are added to the semen. Among them, dimethylacetamide has been the most used in freezing protocols for rooster semen in the last years. Sperm lipid modulation, by dietary means or by adding egg yolk or its derivates to the diluents, also helps in semen preservation. Benefits are improved when antioxidants are included.

Produção e preservação do sêmen de galos / Production and preservation of rooster semen

A seleção de reprodutores capazes de transmitir características de valor econômico para a progênie é fundamental para o sucesso da indústria de aves. Nesse sentido, os machos devem produzir sêmen de boa qualidade, com volume e concentração adequados. O ejaculado de galos é composto por uma grande quantidade de células espermáticas suspensas em um pequeno volume de plasma seminal, proveniente dos túbulos seminíferos. Por esse motivo, a adição de diluentes ao sêmen é uma prática de rotina nos estabelecimentos que realizam inseminação artificial, com a finalidade de aumentar o aproveitamento do ejaculado. A diluição também permite manter a qualidade do sêmen por várias horas, quando resfriado, ou por períodos mais longos, quando congelado. Para a manutenção da capacidade fertilizante do sêmen resfriado, substratos energéticos e agentes tampões devem ser incluídos no diluente. Para diminuir os danos sofridos pelos espermatozoides durante o congelamento, crioprotetores são adicionados ao sêmen. Entre eles, a dimetilacetamida vem sendo o mais utilizado nos protocolos para o congelamento de sêmen de galos nos últimos anos. A modulação lipídica do espermatozoide, por meio de dietas ou por adição de gema de ovo ou de seus derivados ao diluente, também auxilia na preservação do sêmen. Os benefícios são ainda maiores quando antioxidantes são incluídos.(AU)

Selecting breeders capable of transmit characteristics of economic value to the offspring is fundamental to poultry industry success. In this sense, males must produce good quality semen, with appropriate volume and concentration. Rooster ejaculates are composed of a great amount of sperm cells suspended in a small volume of seminal plasma, originated in the seminiferous tubules. For this reason, adding diluents to semen is a common practice in artificial insemination establishments, aiming to increase ejaculate use. Dilution also allows semen quality maintenance for hours, when refrigerated, or for longer periods, when frozen. To keep fertilizing capacity of refrigerated sperm, energetic substrates and buffer agents must be included into the diluents. To decrease sperm damage during freezing, cryoprotectants are added to the semen. Among them, dimethylacetamide has been the most used in freezing protocols for rooster semen in the last years. Sperm lipid modulation, by dietary means or by adding egg yolk or its derivates to the diluents, also helps in semen preservation. Benefits are improved when antioxidants are included.(AU)

Vitrification of the inner perivitelline layer of chicken eggs for use in the sperm-egg interaction assay.

The sperm-egg interaction assay is a good predictor of the fertilizing potential of rooster semen; the ability of chicken sperm to interact with the egg can be assessed by counting the number of holes in the inner perivitelline layer (IPVL) of a freshly laid egg. Although isolated IPVL can be stored for up to 24h, preservation of IPVL for prolonged intervals in liquid nitrogen would facilitate the sperm-egg interaction assay. The objective of this study was to adapt the technique of vitrifying swine oocytes for use with the IPVL. Our hypothesis was that vitrification would not alter the ability of the membrane to bind sperm; therefore, there would be no difference between vitrified and fresh IVPL in the number of hydrolysis holes made by sperm. Our hypothesis was supported; there were no differences in the mean+/-SEM number of holes made by the same sample of sperm in vitrified and in fresh membranes (146.0+/-17.7 holes/mm(2) IPVL and 159.5+/-17.7 holes/mm(2) IPVL, respectively, P>0.05; n=123 IVPLs tested). Furthermore, 80% of frozen-thawed membranes were recovered intact. Because vitrification did not significantly change the ability of membranes to bind sperm, vitrified membranes can be safely used for the sperm-egg interaction assay. Vitrified IVPL would ensure availability for sperm evaluation and facilitate wide distribution of IPVL, enabling assays to be conducted even in the absence of facilities or expertise to prepare membranes.

Evaluation of amides and centrifugation temperature in boar semen cryopreservation.

Two experiments were conducted to evaluate the use of amides as cryoprotectants and two centrifugation temperatures (15 or 24 degrees C) in boar semen cryopreservation protocols. Semen was diluted in BTS, cooled centrifuged, added to cooling extenders, followed by the addition of various cryoprotectants. In experiment 1, mean (+/-S.E.M.) sperm motility for 5% dimethylformamide (DMF; 50.6+/-1.9%) and 5% dimethylacetamide (DMA; 53.8+/-1.7%) were superior (P<0.05) to 5% methylformamide (MF; 43.2+/-2.4%) and 3% glycerol (GLY; 38.1+/-2.3%), with no significant difference between MF and GLY. Sperm membrane integrity was higher (P<0.05) for DMA than for MF or GLY (50.9+/-1.9, 43.3+/-2.5, and 34.5+/-2.8%, respectively). Sperm membrane integrity was higher in DMF (47.9+/-2.1%) than in glycerol (34.5+/-2.8%, P<0.05), but was similar to other treatments (P>0.05). In experiment 2, we tested MF, DMF, and DMA at 3, 5, and 7%. Sperm motility and membrane integrity were higher for 5% DMA (53.8+/-1.7 and 50.9+/-1.9%) and 5% DMF (50.6+/-1.9 and 47.9+/-2.1%), in comparison with 7% DMF and all MF concentrations (P<0.05). For sperm motility and membrane integrity, 5% DMA exceeded (P<0.05) 3% DM, with greater membrane integrity than 3% DMF (P<0.05). In both experiments, sperm motility and membrane integrity were superior at 15 degrees C versus 24 degrees C (P<0.05), with no interaction between centrifugation temperature and treatments (P>0.05). In conclusion, boar semen was successfully cryopreserved by replacement of glycerol with amides (especially 5% DMA) and centrifugation at 15 degrees C, with benefits for post-thaw sperm motility and membrane integrity.