Maria Amélia Zogno

Effect of two ram sperm capacitating media on acrosome reaction and zona-free hamster oocyte penetration test

An in vitro zona-free hamster oocyte penetration test was utilized in 24 trials in order to evaluate the capacitation media used for ram sperm. A pool of fresh semen was collected from three crossed breed rams. Two semen drops were washed by centrifugation and incubated in high ionic strength treatment (HIS) or in a defined medium with HEPES, on heat ewe serum and heparin. After the incubation to promote capacitation, simplified triple-stain technique was used to evaluate the spontaneous acrosome reaction of the capacitated sperm. Superovulation in 96 golden hamsters was induced by PMSG and hCG. The oocytes were treated with hyaluronidase and trypsin to remove, respectively, the cumulus cells and the zona pellucida. Oocytes and capacitated sperm were incubated during 3 hours for further penetration. Then, oocytes were fixed and stained, being evaluated under phase contrast microscope. No significant statistical difference (p >; 0.05) was found between media, concerning the penetration rate of the capacitated sperm and between number of sperm viable with acrosome reaction after the capacitation treatment using two different media. It was concluded that both media utilized were effective in capacitating ram sperm.

Análise bioquímica dos líquidos fetais e citologia do fluido amniótico da fêmea de Mocó (Kerodon rupestris)

The aim of the present work was to study certain biochemical characteristics of fetal fluids of the female moco (Kerodon rupestris), by assesment of the following parameters present in the amniotic and allantoic fetal fluids: osmolarity, pH, calcium, phosphorous, total protein, urea, creatinina and glycose. Ten genital systems of mocos were sampled (gestation = 30-45 and 65-70 days). The amniotic and allantoic sacs were punctured and the fetal fluid obtained for posterior analysis. It was observed the following concentrations (mg/ml) in the amniotic fluid: glycose= 45,87 ± 22,38; calcium = 6,31 ± 1,24; phosphorus = 1,64 ± 0,72; creatinine = 0,45 ± 0,12; urea = 34,03 ± 5,96; total protein = 31,24 ± 16,67. In the allantoic fluid the concentrations were: : glycose = 59,17 ± 10,85; calcium = 5,58 ± 0,59; phosphorous = 1,27 ± 0,73; creatine = 0,38 ± 0,38; urea = 31,49 ± 2,28; total protein = 30,70 ± 18,39. Results obtained from the analisys of the composition of fetal fluids showed small oscillations between the amniotic and the allantoic fluids. Their concentrations, probably, are determined by the materno-fetal metabolic activity. Amniotic fluid cytology showed four cells types.

Comparação da eficácia de dois métodos de extração para determinação do perfil lipídico de ácidos graxos do sêmen ovino por meio da Cromatografia Líquida de Alta Eficiência

The sheep industry for wool, milk and meat production is of increasing importance and new technologies for assessment of semen are in course for the elucidation of male infertility causes. It is well known that damage to the sperm membrane decreases semen quality. Considering that sperm membranes are composed by a phospholipid bilayer, and that lipid peroxidation is a major cause of cell damage, studieson the lipid components of semen are relevant. Lipid peroxidation is a consequence of the reaction between lipids and reactive oxygen species. This event may be reduced in the presence of antioxidants in semen. Semen was collected with an artificial vagina and, after sperm evaluation, samples were centrifuged to separate the sample into two fractions: seminal plasma and spermatozoa pellet. Both had their lipids extracted by two different methods based on Folch, Less& Stanley method modified, using chloroform e methanol as solvents. After the extraction, some esterified fatty acids were qualified and quantified for the sensitivity and specificity to the high performance liquid chromatography in order to determine the most efficient extraction technique in quantitative and qualitative aspects. Statistical analysis was performed using SAS software for Windows. The predominant saturated fatty acid in sperm under these experimental conditions was the myristic and the most abundant insaturated fatty acid in both extractions was DHA. In seminal plasma, in both methods, the prevailing fatty acid is the saturated palmitic and the unsaturated oleic. Among the methods evaluated, we obtained the best results of identification and quantification of fatty acids in Method 1.

Influência do nível de uréia plasmático sobre parâmetros citogenéticos de machos ovinos deslanados

The experiment was planned to verify the effects of increasing blood urea levels resulting from increasing urea in the diet on cytogenetic parameters of Santa Ines Brazilian breed rams (Ovis aries), aged 13 to 18 months. Eighteen animals were divided into three experimental treatments consisted of: diet A ration with nitrogen levels, according to NRC (1985); diet B - ration with a 133% higher level of nitrogen than diet A; and C- ration with a 166% higher level of nitrogen than diet A. The increase in nitrogen level was obtained by adding 20, 40 g of feed grade urea in treatments B and C, respectively. Blood was collected on a weekly basis to determine urea concentration. Blood was also collected from each animal, 60 days after desired diet urea levels were reached, to obtain a lymphocyte culture in order to count chromosome number and determine mitotic index. Increases in plasma urea level were observed (p < 0.0001), but did not affect the observed traits.0 experiment° foi planejado para verificar o efeito do aumento dos niveis plasmaticos de ureia, resultantes do acrescimo de ureia a dieta, sobre parametros citogeneticos de ovinos (Ovis aries) deslanados brasileiros, da raga Santa Ines, corn idade entre 13 e 18 meses. Dezoito animais foram distribuidos a tres dietas que consistiam de: dieta A racdo contendo niveis de nitrogenio de acordo corn NRC (1985); dieta B racdo contendo nivel de nitrogenio 133% superior ao da rag-do A; e dieta C racdo contendo nivel de nitrogenio 166% superior ao da racdo A. 0 aumento no nivel de nitrogenio foi obtido corn acrescimo de 20 e 40 g de ureia grau alimento, para os tratamentos B e C, respectivamente. A colheita de sangue para determinacdo do nivel de ureia no plasma foi semanal. Para a realizacdo da cultura de linfOcitos, para a verificacdo do ntimero de cromossomos e indice mitOtico, 60 dias apOs terem sido alcancados os niveis de ureia desejados na dieta, foi coletado sangue de cada animal. 0 aumento dos niveis de ureia no plasma (p < 0,0001) ndo afetou os pardmetros observados.Estudou-se o efeito de niveis aumentados de ureia plasmatica sobre parâmetros reprodutivos de machos ovinos (Ovis aries) deslanados da raca Santa Ines, em duas fases de 60 dias cada. Durante a primeira fase, 18 animais divididos em tres blocos de acordo com perimetro escrotal foram submetidos aos seguintes tratamentos: dieta A1 - com niveis de nitrogenio de acordo com ARC(1980); dieta B1 - com niveis de nitrogenio 133% maiores que dieta A1; e dieta C1 - niveis de nitrogenio 166% maiores que dieta A1. Durante a segunda fase, 12 animais divididos em dois grupos receberam as dietas: A2 - com niveis de nitrogenio de acordo com ARC (1980) e dieta B2 - niveis de nitrogenio 200% maiores que dieta A2. O aumento no equivalente proteico foi obtido com adicao de 20, 40 e 60 g de ureia aos tratamentos B1, C1 e B2, respectivamente. Foi coletado semen diariamente com vagina artificial e duas vezes por semana foram analisados: motilidade retilinea e progressiva, vigor, turbilhonamento, densidade, pH, concentracao, morfologia espermatica, volume ejaculado e total de espermatozoides por ejaculado. A concentracao de ureia no plasma e no semen foi analisada semanalmente. O aumento (p < 0,0001) de ureia no semen e no plasma durante ambas as fases nao afetou os parâmetros estudados.