Germoplasm banking is an important tool for the preservation of genetic material from Neotropical primates in captivity, and from free living species, especially the endangered ones like Saimiri vanzolinii (Black-headed squirrel monkey), a primate with a low incidence area (870 km(2) of floodplains) in the southern part of the Mamirauá Sustainable Development Reserve, Brazil. Therefore, in the present study we aimed to develop a sperm cryopreservation protocol comparing sperm cooling in presence (T1) and absence (T2) of egg yolk, and to test freezing protocols to preserve semen from captive (Saimiri collinsi), and free-living (Saimiri vanzolinii, Saimiri cassiquiarensis and Saimiri macrodon) New World primates. Cooling preserved sperm of S. collinsi in all evaluated microscopic parameters, except for sperm motility. No differences were observed among the treatments, indicating that semen of this species can be cooled without egg yolk. Freezing did not affect sperm quality of S. collinsi, except plasma membrane integrity that was negatively affected. Generally, a good maintenance rate was observed between cooling and thawing of semen for the four species, showing the positive translational application of protocols from S. collinsi to the free-living species. Developed freezing protocol proved to be useful for sperm cryopreservation of S. collinsi and in field conditions.
RESUMO Comparou-se a eficiência de protocolos para indução de estro em cutias. Em cinco fêmeas, foram administradas duas doses de cloprostenol (5µg) com intervalo de nove dias, via intraperitoneal; em outras cinco, administraram-se 30µg de análogo do hormônio liberador de gonadotrofinas (GnRH), via intravulvar, seguidos de 5µg de cloprostenol, via intraperitoneal, após sete dias e, após mais dois dias, nova dose do análogo de GnRH. A cada três dias, a ciclicidade reprodutiva dos animais foi monitorada, por meio de coleta de sangue, para dosagem hormonal, ultrassonografia ovariana e citologia vaginal. Duas das fêmeas que receberam apenas prostaglandina, as quais estavam em fase luteal no início do tratamento, manifestaram o estro aos três e seis dias após a segunda administração da droga. Já nas fêmeas que receberam a prostaglandina associada ao análogo do GnRH, duas que originalmente estavam em fase luteal apresentaram estro aos quatro dias após o tratamento, e uma outra apenas após 10 dias. Não foram evidenciadas diferenças estatísticas quanto à eficiência dos tratamentos (P>0,05). Conclui-se que, de acordo com os protocolos utilizados, o uso da prostaglandina isolada ou em associação com análogo do GnRH para a indução do estro em cutias D. leporina apresenta eficiência limitada às fêmeas que estejam em fase luteal por ocasião do início do tratamento.
Avaliou-se 31 cães saudáveis, sem raça definida, sendo 10 machos e 21 fêmeas, com 8 meses a 7 anos de idade e peso de1,5-28 kg. Inicialmente foram mensurados os diâmetros fronto-occiptal (DFO) e bizigomático (DBZ) do crânio com o auxílio de um paquímetro. A ultra-sonografia transpalpebral em modo-B foi realizada para mensurar as estruturas do bulbo ocular, conforme se segue: D1- espessura da córnea; D2- distância entre o ponto central da imagem da córnea e a da cápsula anterior do cristalino (câmara anterior); D3- distância entre o ponto central da imagem da córnea e a da cápsula posterior do cristalino; D4- espessura do cristalino, que corresponde a distância entre a imagem da cápsula anterior e a cápsula posterior do cristalino; D5- diâmetro do cristalino, distância entre as imagens dos pólos do cristalino; D6- área do cristalino; D7- câmara vítrea, distância entre a imagem da cápsula posterior do cristalino e a retina; D8- distância entre a cápsula anterior do cristalino e a retina; D9- distância entre a imagem da córnea e a retina. Com exceção da D4, houve efeito dos DFO e DBZ sobre as medidas das estruturas internas do BO. A análise de regressão linear entre as medidas das estruturas do bulbo ocular e os DFO e DBZ foram significativas para D1, D2, D3, D4, D5, D6, D7, D8 e D9.
The aim of this study was to evaluate whether an in vitro culture (IVC) medium containing either or not b-mercaptoethanol (BME), bone morphogenetic protein 4 (BMP4), or pregnant mare serum gonadotrophin (PMSG) could be able to promote the development of capuchin monkeys' preantral follicles enclosed in ovarian cortical strips. Follicular viability after IVC was similar to control (89.32%). Primordial follicle recruitment to primary stage was not reached with IVC, but the rate of secondary follicle formation was increased in the medium supplemented with BME, BMP4, and PMSG (44.86%) when compared to IVC control (9.20%). In the medium supplemented with BME, BMP4, and PMSG, contrary to other media, anti-müllerian hormone-messenger RNA (mRNA) expression in ovarian tissue was upregulated (3.4-fold), while that of growth differentiation factor-9 was maintained. The BMP4-mRNA expression, however, appeared downregulated in all cultured tissues. Our findings show a favorable effect of BME, BMP4, and PMSG on the in vitro development of secondary follicles from capuchin monkeys.
Ovarian fragments were exposed to 0.5 M sucrose and 1 M ethylene glycol (freezing solution; FS) with or without selenium or Trolox. Histological and ultrastructural analyses showed that the percentages of normal follicles in control tissue and in tissue after exposure to FS+50 μM Trolox were similar. Trolox prevented endoplasmic reticulum (ER)-related vacuolization, which is commonly observed in oocytes and stromal tissue after exposure to FS. From the evaluated stress markers, superoxide dismutase 1 (SOD1) was up-regulated in ovarian tissue exposed to FS+10 ng/ml selenium. Ovarian fragments were subsequently frozenthawed in the presence of FS with or without 50 μM Trolox, followed by in vitro culture (IVC). Antioxidant capacity in ovarian fragments decreased after freeze-thawing in Troloxfree FS compared with FS+50 μM Trolox. Although freezing itself minimized the percentage of viable follicles in each solution, Trolox supplementation resulted in higher rates of viable follicles (67 %), even after IVC (61 %). Furthermore, stress markers SOD1 and ERp29 were up-regulated in ovarian tissue frozen-thawed in Trolox-free medium. Relative mRNA expression of growth factors markers was evaluated after freeze-thawing followed by IVC. BMP4, BMP5, CTGF, GDF9 and KL were down-regulated independently of the presence of Trolox in FS but down-regulation was less pronounced in the presence of Trolox. Thus, medium supplementation with 50 μM Trolox prevents ER stress and, consequently, protects ovarian tissue from ER-derived cytoplasmic vacuolization. ERp29 but not ERp60, appears to be a key marker linking stress caused by freezing-thawing and cell vacuolization.
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