Comparison of efficiency between two artificial insemination methods using fresh semen in domestic cat (Felis Catus) / Comparação da eficiência entre dois métodos de inseminação artificial utilizando sêmen fresco em gata doméstica (Felis Catus)

This study aimed to compare domestic cats' pregnancy rates using fresh semen for the bilateral intrauterine insemination (BIUI) method and the novel uterine body insemination (UBI) method. Queens received a single injection of equine chorionic gonadotropin (eCG) (200 IU; IM) to induce ovarian follicular development and, after 83 h, an injection of human chorionic gonadotropin (hCG) (100 IU; IM) for final oocyte maturation and ovulation induction. Thirty-four hours after hCG administration, 3 × 106fresh spermatozoa were used for insemination by the BIUI (n = 8 queens) or by the UBI (n = 7 queens) techniques,respectively. Pregnancy rates were 75.00% (6/8) by BIUI and 42.85% (3/7) by the UBI method. Themean litter size was 3.0 ± 0.86 for the BIUI, and 2.0 ± 1.0 for the UBI method. Spontaneous abortion occurred on day 35 of pregnancy in one queen following the UBI method. Our findings showed that the BIUI of queens with fresh semen resulted in higher pregnancy rates than the novel UBI method; also, acceptable pregnancy rates were achieved following BIUI with fresh semen in the domestic cat.(AU)

O objetivo deste estudo foi comparar as taxas de prenhez em gatas domésticas usando sêmen fresco para o método de inseminação intrauterina bilateral (BIUI) e o novo método de inseminação do corpo uterino (UBI). As gatas receberam uma única injeção de gonadotrofina coriônica equina (eCG) (200 UI; IM) para induzir o desenvolvimento folicular ovariano e, após 83 h, uma injeção de gonadotrofina coriônica humana (hCG) (100 UI; IM) para maturação final do oócito e ovulação indução. Trinta e quatro horas após a administração de hCG, 3 × 106 espermatozoides frescos foram utilizados para inseminação pelas técnicas de BIUI (n = 8 gatas) ou UBI (n = 7 gatas), respectivamente. As taxas de gravidez foram de 75,00% (6/8) pela BIUI e 42,85% (3/7) pelo método UBI. O tamanho médio da ninhada foi de 3,0 ± 0,86 para o método BIUI e 2,0 ± 1,0 para o método UBI. Aborto espontâneo ocorreu no dia 35 de gestação em uma gata seguindo o método UBI. Nossos achados mostraram que a BIUI de gatas com sêmen fresco resultou em maiores taxas de prenhez do que o novo método UBI; também, taxas de prenhez aceitáveis foram alcançadas após BIUI com sêmen fresco no gato doméstico.(AU)

Use of new recombinant proteins for ovarian stimulation in ruminants

Currently, gonadotropin products (follicle stimulating hormone, FSH, and luteinizing hormone, LH) used in animal reproduction are produced by extraction and purification from abattoir-derived pituitary glands. This method, relying on animal-derived materials, carries the potential risk of hormone contamination and pathogen transmission. Additionally, chorionic gonadotropins are extracted from the blood of pregnant mares (equine chorionic gonadotropin; eCG) or the urine of pregnant women (human chorionic gonadotropin; hCG). However, recent advancements have introduced recombinant gonadotropins for assisted animal reproduction therapies. The traditional use of FSH for superovulation has limitations, including labor requirements and variability in superovulation response, affecting the success of in vivo (SOV) and in vitro (OPU/IVEP) embryo production. FSH treatment for superstimulation before OPU can promote the growth of a homogenous follicular population and the recovery of competent oocytes suitable for IVEP procedures. At present, a single injection of a preparation of long-acting bovine recombinant FSH (rFSH) produced similar superovulation responses resulting in the production of good-quality in vivo and in vitro embryos. Furthermore, the treatment with eCG at FTAI protocol has demonstrated its efficacy in promoting follicular growth, ovulation, and P/AI, mainly in heifers and anestrous cows. Currently, treatment with recombinant glycoproteins with eCG-like activity (r-eCG) have shown promising results in increasing follicular growth, ovulation, and P/AI in cows submitted to P4/E2 -based protocols. Bovine somatotropin (bST) is a naturally occurring hormone found in cows. Recombinant bovine somatotropin (rbST), produced through genetic engineering techniques, has shown potential in enhancing reproductive outcomes in ruminants. Treatment with rbST has been found to improve P/IA, increase donor embryo production, and enhance P/ET in recipients. The use of recombinant hormones allows to produce non-animal-derived products, offering several advantages in assisted reproductive technologies for ruminants. This advancement opens up new possibilities for improving reproductive efficiency and success rates in the field of animal reproduction.(AU)

Regulation of progesterone during follicular development by FSH and LH in sheep

Progesterone (P4) can participate in the development of female mammalian antral follicles through nuclear receptor (PGR). In this experiment, the differences of P4 synthesis and PGR expression in different developmental stages of sheep antral follicles (large > 5mm, medium 2-5mm, small < 2mm) were detected by enzyme-linked immunosorbent assay, immunohistochemistry, qRT-PCR and Western blotting. Secondly, sheep follicular granulosa cells were cultured in vitro. The effects of different concentrations of FSH and LH on P4 synthesis and PGR expression were studied. The results showed that acute steroid regulatory protein (StAR), cholesterol side chain lyase (P450scc) and 3β Hydroxysteroid dehydrogenase (3β-HSD) and PGR were expressed in antral follicles, and with the development of antral follicles in sheep, StAR, P450scc and the expression of 3β-HSD and PGR increased significantly. In vitro experiments showed that FSH and LH alone or together treatment could regulate P4 secretion and PGR expression in sheep follicular granulosa cells to varying degrees, hint P4 and PGR by FSH and LH, and LH was the main factor. Our results supplement the effects of FSH and LH on the regulation of P4 synthesis during follicular development, which provides new data for further study of steroid synthesis and function in follicular development.(AU)

Effect of different gonadotropins on in vitro maturation of sheep oocytes / Efeito de diferentes gonadotrofinas na maturação in vitro de oócitos ovinos

The aim of this study was to examine the effect of replacing the use of follicle-stimulating hormone (FSH) with equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG) on the in vitro maturation (IVM) of sheep oocytes. After sheep ovaries were collected (n=300), the cumulus-oocyte complexes were aspirated, selected, and divided into four groups according to the IVM medium: CON group, in which the basic IVM medium was used; and eCG, hCG, and FSH groups, in which the oocytes were immersed in basic IVM medium with 10 IU/mL eCG, 10 IU/mL hCG, and 10 µg/mL FSH-p, respectively. In vitro maturation of the oocytes was performed at 38.5 °C, in a humidified atmosphere of 5% CO2 in air, for 24 h. Subsequently, the oocytes were evaluated for the degree of cumulus-cell expansion, chromatin configuration, GSH levels, and active mitochondria. There were no significant differences for the rate of cumulus cell expansion. The percentage of oocytes in MII was higher in the eCG group than in the CON and hCG groups (P<0.05) and similar to that of the FSH group. In conclusion, eCG can be used as a substitute for FSH in IVM of sheep oocytes.

O objetivo deste estudo foi avaliar o efeito da gonadotrofina coriônica equina (eCG) e da gonadotrofina coriônica humana (hCG), em substituição ao uso de hormônio folículo estimulante (FSH) na maturação in vitro (MIV) de oócitos ovinos. Após a coleta de ovários (n=300) ovinos, os complexos cúmulus-oócitos (CCOs) foram aspirados, selecionados e divididos em quatro grupos de acordo com o meio de MIV: grupo CON, em que foi utilizado o meio MIV base; e grupos ECG, HCG e FSH, em que os oócitos foram imersos em meio MIV base adicionado de 10 UI/mL de eCG, 10 UI/mL de hCG e 10 µg/mL de FSH-p, respectivamente. A MIV dos oócitos foi realizada a 38,5°C, em atmosfera umidificada de 5% de CO2 em ar, durante 24 horas. Posteriormente, os oócitos foram avaliados, quanto grau de expansão das células do cumulus, configuração da cromatina, níveis de GSH e mitocôndrias ativas. Não foram observadas diferenças significativas com relação à taxa de expansão de células do cumulus. A percentagem de oócitos em MII foi maior no grupo ECG do que no grupo CON e HCG (P<0,05) e semelhante ao grupo FSH. Em conclusão, a eCG pode ser utilizada em substituição ao FSH na MIV de oócitos ovinos.

Hormonal induction and synchronization in the reproduction of Nile tilapia (Oreochromis niloticus) / Sincronização e indução hormonal na reprodução de tilápia do nilo (Oreochromis niloticus)

The present study aims to evaluate the sexual synchronization of Nile tilapia (Oreochromisniloticus) breeding herds under different distributions and days. This study was carried out during, a period of 21 experimental days. It was conducted in a randomized block design (DBC), in a factorial scheme 2 × 2 × 3, (male and female, with and without hormone induction, and three times blocks of 7, 14 and 21 days). A total of 324 fish were used, 180 females and 144 males, previously microchipped with approximately 250 ± 12.25 g of body weight.. Each tank contained 36 animals, with every three tanks comprising a block of which were repeated in time. Each of the tanks was composed of only males, another with 12 males and 24 females separated by a glass and a screen on the bottom and the third tank with only females. In each treatment, half of the animals (male and female) were applied the hCG hormone, in a single dose at a concentration of 5 IU / gram fish live weight. The animals were evaluated at 7, 14, and 21 days. After 667 hours-degree after application of the hormone was made extrusion and spermatozoa by celomatic massage. It was observed females that showed ease of extrusion of gametes. For the females, spawning index, absolute and relative fecundity, oocyte weight per 1 g, diameters, and germinal vesicle peripheral position were evaluated; the sperm concentration, volume (ml), motility (%), duration of sperm motility (min) and integrity were evaluated in males. The data were submitted to analysis of variance at the 5% level of significance. For females at 14 days there was greater ease of extrusion when separated from males, with hormone application . The distribution influenced the spawning weight, the larger diameter, and for the diameters smaller the days and induction presented interference. In males, the days interfered in the increase in volume and duration of semen motility independent of induction and environment, and finally, the environment interfered significantly in semen concentration.

O presente estudo tem como objetivo avaliar a sincronização sexual de reprodutores de tilápias do Nilo (Oreochromis niloticus) em cativeiro sob diferentes distribuições e dias. Este estudo foi realizado por um período de 21 dias experimentais. Foi conduzido num delineamento de blocos casualizados (DBC), num esquema fatorial 2×2×3, (sendo macho e fêmea, com e sem indução hormonal) e três tempos, 7, 14 e 21 dias (blocos). Foram utilizados 324 peixes, sendo 180 fêmeas e 144 machos, com aproximadamente 250 ± 12,25g de peso corporal, previamente microchipados. Cada tanque continha 36 animais, sendo que a cada três tanques compreendia um bloco do qual foram repetidos no tempo. Cada um dos tanques esteve composto por: somente machos, outro com 12 machos e 24 fêmeas separados por um vidro e uma tela na parte de baixo e o terceiro tanque com apenas fêmeas. Em cada tanque, metade dos peixes (macho e fêmeas) recebeu indução hormonal com hCG (CHORULON®), na concentração de 5000UI/grama de peso vivo por peixe. Os animais foram avaliados aos 7, 14, e 21 dias. Passado 667 horas-grau após aplicação do hormônio foi feita extrusão e espermeação mediante massagem celomática. Foram observadas fêmeas que apresentaram facilidade de extrusão de gametas. Para asfêmeas foram avaliadas as seguintes variáveis: índice de desova, fecundidade absoluta e relativa, o peso dos ovócitos por cada 1g, os diâmetros, e posição periférica da vesícula germinativa; nos machos, foram avaliados a concentração espermática, o volume (ml), motilidade (%), tempo de duração de motilidade espermática (min) e integridade. Os dados obtidos foram submetidos a análise de variância ao nível de 5% de significância. Observou-se para fêmeas aos 14 dias, mantidas separada dos machos e com aplicação de hormônios, maior facilidade de extrusão, peso da desova diâmetro dos ovócitos. Nos machos, os dias interferiram no aumento dovolume e na duração de motilidade do sêmen independente da indução e ambiente, e por fim, o ambiente interferiu significativamente na concentração do sêmen.

Influence of eCG and reproductive management in the resynchronization of ovulation in dairy goats

Resynchronization protocols have been proposed as a way of shortening females' unproductive time in the flock, with good results in cattle and sheep. In goats, initial studies have shown that a second progestogen device inserted before luteolysis and pregnancy diagnosis does not interfere with the corpus luteum lifespan or functionality. This study aimed to evaluate the follicular growth, ovulation pattern and pregnancy rate after insertion of a second and new progestogen device for resynchronizing, with or without equine Chorionic Gonadotrophin (eCG), submitted to natural mating (NM) or artificial insemination (AI) to propose a viable resynchronization protocol for dairy goats. A total of 38 multiparous Saanen goats underwent a short-term progesterone protocol [six days exposed to medroxyprogesterone acetate (MAP) intravaginal sponges + 200 IU eCG and 0.12 mg of cloprostenol sodium on the 5th day + 0.025 mg of lecirelin 34 hours after sponge withdrawal] and, on day 16th after the ovulation, received a new MAP device which was retained until day 21. At this moment females were split into four groups: GeCG+NM ­ 100 IU eCG with NM; GSal+NM ­ saline solution with NM; GeCG+AI ­ 100 IU eCG with AI; and GSal+AI ­ saline solution with AI. Ultrasound scans were performed every 12 h from sponge withdrawal (day 21) until 108 h after sponge withdrawal (day 25) for follicular dynamics evaluation, at 240 h (day 31) for assessing the presence of active corpus luteum, and on day 60 for pregnancy diagnosis. No differences were found regarding ovulation time, synchronization and follicle size. However, GeCG+NM presented a greater estrus manifestation rate (100%) and pregnancy rate (62.5%) when compared to GSal+AI. In conclusion, resynchronization protocols in dairy goats may present satisfactory results.(AU)

Induction of accessory corpus luteum in goats and sheep: From physiological basis to reproductive effects / Indução de corpo lúteo acessório em caprinos e ovinos: Das bases fisiológicas aos efeitos reprodutivos

A identificação das ondas foliculares ovarianas e de seu padrão hormonal revelou que os folículos ovarianos dominantes da onda ovulatória (FDOO) crescem em ambiente hormonal com predominância crescente de estradiol, diferentemente daqueles da primeira (FDPO) e das ondas foliculares intermediárias (FDOI), que crescem sob forte impacto da progesterona (P4). O hormônio luteinizante (LH) é considerado o hormônio gonadotrófico decisivo para direcionar se um folículo dominante ovulará (↑ LH) ou não (↓ LH). Estratégias foram desenvolvidas para aumentar o LH endógeno (administração de GnRH) ou fornecer LH exógeno de origem suína (pLH) ou, ainda, gonadotrofinas semelhantes ao LH, como gonadotrofina coriônica humana (hCG). Estas medidas são capazes de disponibilizar LH para maturação final, ovulação e/ou luteinização de FDPO ou FDOI, formando corpos lúteos acessórios (CLa). Como consequência, a P4 aumenta e favorece o estabelecimento da gestação, sobretudo em condições em que a P4 for o fator limitante para a implantação e manutenção embrionária. Em ovelhas e cabras, em diferentes estudos, a hCG foi administrada de cinco a sete dias após o início do estro e revelou que o FDPO responde positivamente à administração de hCG, formando CLa e/ou promovendo a hipertrofia do CL formado originalmente, aumentando a área luteal. O incremento da P4 normalmente acompanha o aumento de área do tecido luteal. Como efeito final e mais desejável, a gestação e o nascimento de cordeiros/cabritos também aumentam. Esses conceitos serão discutidos na presente revisão sobre indução de CLa em ovinos e caprinos.

The identification of ovarian follicular waves and associated hormonal milieux has revealed that dominant follicles of the ovulatory wave (OWDF) grow in a hormonal environment where there is an increasing predominance of estradiol, unlike first-wave dominant follicles (FWDF) and intermediatewave dominant follicles (IWDF), which grow under increasing progesterone (P4) concentrations. The luteinizing hormone (LH) is considered the decisive gonadotropic hormone to direct whether a dominant follicle will (↑ LH) or will not (↓ LH) ovulate. Based on this, strategies have been developed to either increase endogenous LH (GnRH administration) or provide exogenous LH of porcine origin (pLH) or LH-like gonadotropins, such as human chorionic gonadotropin (hCG). Such strategies are able to provide LH for final maturation, ovulation, and/or luteinization of the FWDF or IWDF, forming accessory corpora lutea (aCL). As a consequence, P4 increases and favors the establishment of pregnancy, particularly when P4 is the limiting factor for the success of the conceptus implantation and maintenance. In sheep and goats, previous studies have administered hCG five to seven days after the onset of estrus and revealed that FWDF positively respond to hCG administration by either forming aCLs and/or promoting hypertrofia of the original CL which, in turn, increases its luteal tissue area. Normally, P4 synthesis increases along with the increase in luteal tissue area. As a final and most desirable outcome, pregnancy and the birth of lambs/kids also increase. These concepts will be discussed in this review, focusing on aCL induction in sheep and goats.

Estrous synchronization in sheep with reused progesterone devices and eCG

The objective of the present research was to determine the effect of long synchronization protocols based on reused progesterone devices (controlled internal drug release [CIDR]) associated with different doses of equine chorionic gonadotropin (eCG) on reproductive variables in crossbred sheep (Suffolk × Kathadin × Dorset). The CIDR were used for eleven days in a previous study in sheep from the same herd and were washed and disinfected before reusing. Sixty-four sheep, in the reproductive season, were randomly assigned to four experimental groups (n = 16). Treatments consisted of a group with 10 d CIDR and 300 IU eCG; a group with 10 d CIDR and 400 IU of eCG; a group with 12 d CIDR and 300 IU of eCG; and a group with 12 d CIDR and 400 IU of eCG. A completely randomized design was used. There was an estrous presentation rate of 100% in all treatments. The begining of estrous, gestation rate, fertility rate, type of parturition, and prolificacy index were equal between groups. Progesterone serum concentration was higher in sheep from the 10 d CIDR groups. The CIDR, reused for the second time, associated with 300 or 400 IU of eCG for estrous synchronization in sheep, are effective to obtain good pregnancy rates and ensures higher prolificacy rates.

Application of hormonal subdoses at the Bai Hui acupoint for estrus synchronization in sheep / Aplicação de subdoses hormonais no ponto de acupuntura Bai Hui para sincronização de estro em ovelhas

Here, the efficiency of applying subdoses of equine chorionic gonadotropin (eCG) and prostaglandin F2α (PGF2α) at the Bai Hui (BH) acupoint for estrus synchronization in sheep was evaluated. Thirty Santa Inês ewes received intravaginal sponges containing 60 mg medroxyprogesterone acetate for 7 days. The animals were divided into five treatments (T) (n=6): T1 (control), 132.5 µg PGF2α (100% of dose) and 300 IU eCG (100% of dose), both intramuscularly (IM); T2, 39.75 µg PGF2α (30% of dose) at the BH acupoint and 300 IU eCG (100% of dose) IM; T3, 132.5 µg PGF2α (100% of dose) IM and 90 IU eCG (30% of dose) at the BH acupoint; T4, 39.75 µg PGF2α (30% of dose) and 90 IU eCG (30% of dose), both at the BH acupoint; and T5, 39.75 µg PGF2α (30% of dose) and 90 IU eCG (30% of dose) both at a false acupoint (IM, at the same location as in T1). After 12 h of sponge removal and hormone application, the ewes were subjected to monitoring for estrus and mating and ultrasound assessments of follicular growth and ovulation time every 12 h. The data were evaluated for normality and subjected to analysis of variance, considering a 5% probability. There were no differences in the percentage of animals in estrus (93.3±9.1%), interval from sponge removal to estrus onset (48.1±6.0 h), interval from sponge removal to estrus end (69.6±5.4 h), duration of estrus (24.0±4.7 h), interval from sponge removal to ovulation (73.7±5.9 h), interval from estrus onset to ovulation (26.5±2.3 h), size of the largest follicle (6.7±0.4 mm), follicular growth rate (0.8±0.1 mm•day-1), number of ovulations (1.3±0.1), and plasma progesterone levels 7 days after ovulation (9.6±2.0 ng•mL-1). The cost of the synchronization protocol per animal was US$ 4.37, 4.12, 2.96, 2.71, and 2.71 for T1, T2, T3, T4, and T5, respectively, with 37.99% cost reduction using PGF2α and eCG at 30% of the conventional doses. Therefore, the application of reduced PGF2α (39.75 µg) and eCG (90 IU) doses at the BH or false acupoint effectively synchronized estrus and reduced protocol cost in sheep, indicating that the conventional doses are overestimated.(AU)

Aqui, foi avaliada a eficiência da aplicação de subdoses de gonadotrofina coriônica equina (eCG) e prostaglandina F2α (PGF2α) no ponto de acupuntura Bai Hui (BH) para sincronização de estro em ovelhas. Trinta ovelhas Santa Inês receberam esponjas intravaginais contendo 60 mg de acetato de medroxiprogesterona por 7 dias. Os animais foram divididos em cinco tratamentos (T) (n=6): T1 (controle), 132,5 µg PGF2α (100% da dose) e 300 UI de eCG (100% da dose), ambos por via intramuscular (IM); T2, 39,75 µg de PGF2α (30% da dose) no ponto de acupuntura BH e 300 UI eCG (100% da dose) IM; T3, 132,5 µg PGF2α (100% da dose) IM e 90 UI eCG (30% da dose) no ponto de acupuntura BH; T4, 39,75 µg PGF2α (30% da dose) e 90 UI eCG (30% da dose), ambos no ponto de acupuntura BH; e T5, 39,75 µg PGF2α (30% da dose) e 90 UI eCG (30% da dose) ambos em um falso acuponto (IM, no mesmo local que em T1). Após 12 horas da retirada da esponja e aplicação do hormônio, as ovelhas foram submetidas ao monitoramento do estro e acasalamento, e avaliação ultrassonográfica do crescimento folicular e tempo de ovulação a cada 12 horas. Os dados foram avaliados quanto à normalidade e submetidos à análise de variância, considerando 5% de probabilidade. Não houve diferenças na porcentagem de animais em estro (93,3 ± 9,1%), intervalo da remoção da esponja ao início do estro (48,1 ± 6,0 h), intervalo da remoção da esponja ao final do estro (69,6 ± 5,4 h), duração do estro (24,0 ± 4,7 h), intervalo da remoção da esponja à ovulação (73,7 ± 5,9 h), intervalo do início do estro à ovulação (26,5 ± 2,3 h), tamanho do maior folículo (6,7 ± 0,4 mm), taxa de crescimento folicular (0,8 ± 0,1 mm•dia-1), número de ovulações (1,3 ± 0,1) e níveis de progesterona plasmática 7 dias após a ovulação (9,6 ± 2,0 ng•mL-1). O custo do protocolo de sincronização por animal foi de US $ 4,37, 4,12, 2,96, 2,71 e 2,71 para T1, T2, T3, T4 e T5, respectivamente, com redução de custo de 37,99% usando PGF2α e eCG a 30% das doses convencionais. Portanto, a aplicação de PGF2α (39,75 µg) e doses reduzidas de eCG (90 UI) no BH ou falso acuponto sincronizaram efetivamente o estro e reduziram o custo do protocolo em ovelhas, indicando que as doses convencionais estão superestimadas.(AU)

Comparison of cattle embryo collection between Ringer's lactate solution and Dulbecco's phosphatebuffered saline

The objective of this study was to determine the efficiency of Ringer's lactate solution (RL) and RL + 1% fetal bovine serum (FBS) and compare them with the efficiency of Dulbecco's phosphate-buffered saline (D-PBS). Twenty-two Wagyu female cattle were subjected to superovulation and were randomly distributed to form three groups: group 1 ­ uterine flushing with RL (n = 8), group 2 ­ uterine flushing with RL + 1% FBS (n = 7), and group 3 ­ uterine flushing with D-PBS (n = 7, control group). Cows received a CIDR® device containing 1.9 g of progesterone at random stages of the estrous cycle (day 0). Progesterone withdrawal occurred on day 8 in the morning. For heifers, 160 mg of porcine follicle-stimulating hormone (FSH-P) was used and for cows, 200 mg. Prostaglandin F2α was also injected on the eighth day of FSH-P administration. On day 9, in the morning, hCG was administered. Females were superovulated and inseminated twice in a fixed time for embryo transfer. On the 16th day, females were subjected to uterine flushing for embryo collection. We collected 76 embryos from 22 females subjected to superovulation, of which 52 were transferable and 24 had degenerated. The total of embryos collected was 23, 16, and 23 for groups 1, 2, and 3, respectively. The embryo recovery rates per group were 13.86±4.23, 15.39±4.61 and 27.16±13.33%, in groups 1, 2, and 3, respectively. The means for the total structures collected per female were 2.88±0.85, 3.00±1.23, and 4.57±1.72 in groups flushed with RL, RL + 1% FBS, and D-PBS, respectively. We conclude that Ringer's lactate solution and Ringer's lactate solution + 1% of FBS and Dulbecco's phosphate-buffered saline showed no significant differences in terms of embryo quality or quantity, suggesting that Ringer's lactate solution is an alternative for collecting embryos in cattle.

Subdose of human chorionic gonadotropin applied at the Hou Hai acupoint on follicular dynamics and luteal development in donkeys

The objective of this study was to evaluate the effects of an hCG subdose applied at the Hou Hai acupoint as an ovulation inducer in donkeys. Eleven donkeys were distributed in randomized blocks in T1 = application of 1,500 IU of hCG intravenous (IV); T2 = 450 IU of hCG applied at the false acupoint (IV), and T3 = 450 IU of hCG applied at the Hou Hai acupoint. There was no difference (P > 0.05) between the treatments regarding the mean diameter of the pre-ovulatory follicle (34.5 ± 1.3 mm), the ovulation rate (96.97%), the interval between induction and ovulation (58.07 ± 16.82 h), the mean diameter of the CL (D0 = 23.0 ± 0.6; D2 = 27.7 ± 1.9 and D8 = 28.2 ± 0.8mm), and serum P4 concentrations (10.50 ± 2.99 ng.mL-1). The application of 450 IU of hCG at the Hou Hai acupoint increased ovulation rate (72.73%) more than 48 h after induction (P = 0.03) and a larger diameter of the CL on D4 (30.7 ± 5.1 mm) (P = 0.04). The vascularization area of the CL on D8, obtained by minimum number of colored pixel (NCP), was greater (P 0.05) in the donkeys that received 1,500 IU of IV hCG (T1, 41.91 ± 1.17), and we found a positive correlation (P 0.05) between mean NCP and P4 concentration in the donkeys that received 450 IU of hCG IV at the false acupoint (T2) or at the Hou Hai acupoint (T3). The application of 450 IU of hCG by IV route at the false acupoint or the Hou Hai acupoint was sufficient to induce ovulation in donkeys, demonstrating that the average dosage commonly used for this species is too high.

Subdose of human chorionic gonadotropin applied at the Hou Hai acupoint on follicular dynamics and luteal development in donkeys

The objective of this study was to evaluate the effects of an hCG subdose applied at the Hou Hai acupoint as an ovulation inducer in donkeys. Eleven donkeys were distributed in randomized blocks in T1 = application of 1,500 IU of hCG intravenous (IV); T2 = 450 IU of hCG applied at the false acupoint (IV), and T3 = 450 IU of hCG applied at the Hou Hai acupoint. There was no difference (P > 0.05) between the treatments regarding the mean diameter of the pre-ovulatory follicle (34.5 ± 1.3 mm), the ovulation rate (96.97%), the interval between induction and ovulation (58.07 ± 16.82 h), the mean diameter of the CL (D0 = 23.0 ± 0.6; D2 = 27.7 ± 1.9 and D8 = 28.2 ± 0.8mm), and serum P4 concentrations (10.50 ± 2.99 ng.mL-1). The application of 450 IU of hCG at the Hou Hai acupoint increased ovulation rate (72.73%) more than 48 h after induction (P = 0.03) and a larger diameter of the CL on D4 (30.7 ± 5.1 mm) (P = 0.04). The vascularization area of the CL on D8, obtained by minimum number of colored pixel (NCP), was greater (P 0.05) in the donkeys that received 1,500 IU of IV hCG (T1, 41.91 ± 1.17), and we found a positive correlation (P 0.05) between mean NCP and P4 concentration in the donkeys that received 450 IU of hCG IV at the false acupoint (T2) or at the Hou Hai acupoint (T3). The application of 450 IU of hCG by IV route at the false acupoint or the Hou Hai acupoint was sufficient to induce ovulation in donkeys, demonstrating that the average dosage commonly used for this species is too high.(AU)

Ultrassonografia testicular no modo color Doppler∕ espectral e avaliação andrológica de garanhões tratados com gonadotrofina coriônica humana (hCG) em diferentes estações do ano / [Testicular ultrasonography in the doppler / spectral color mode and andrological evaluation of stallions treated with human corionic gonadotrofin (hCG) in different seasons of the year]

Utilizaram-se quatro garanhões nos meses de janeiro, abril, julho e outubro de 2016, em dois protocolos, em que: GI (n=4; 5mL de solução salina, i.v.) e GII (n=4; 5000UI de hCG, i.v.) e subdividiram-se esses protocolos em ciclos (C1 e C2), seguindo o esquema crossover, sendo: CI=animais 1 (GI) e 2 (GII), avaliados nos dias D1, D3 e D5, e animais 3 (GI) e 4 (GII), em D2, D4 e D6; CII= animais 1 (GII) e 2 (GI), em D1, D3, D5, e animais 3 (GII) e 4 (GI), em D2, D4 e D6. Realizou-se o tratamento em D1 e D2 de cada ciclo e a ultrassonografia testicular no modo color Doppler e Doppler espectral, uma hora antes de cada coleta de sêmen e imediatamente após. Avaliou-se: número de reflexo de Flehmen, de montas sem ereção, início da monta, tempo de reação à ereção e total da monta e análises de qualidade seminal. Estatisticamente foram utilizados os testes de qui-quadrado e ANOVA. Não houve diferenças estatísticas (P>0,05) entre os parâmetros analisados. Concluiu-se que uma única dose de 5000UI de hCG em garanhões não causou alterações significativas nos parâmetros avaliados em diferentes estações do ano.(AU)

Four stallions were used in January, April, July and October 2016 in two protocols: GI (n=4; 5ml saline, iv) and GII (n=4; 5000 hCG, iv), and these protocols were subdivided into cycles (C1 and C2) following the cross over scheme, as follows: CI=animal 1 (GI) and 2 (GII) evaluated on days D1, D3 and D5 and animal 3 (GI) and 4 (GII) at D 2, D 4 and D 6; CII=animal 1 (GII) and 2 (GI) at D1, D3, D5 and animal 3 (GII) and 4 (GI) at D2, D4 and D6. Treatment was performed on D1 and D2 of each cycle and testicular ultrasound in color Doppler and spectral Doppler mode, one hour before each semen collection and immediately after. We evaluated: Flehmen's reflex number, mounts without erection, start of the mount, reaction time to erection and total mount and seminal quality analyzes. Statistically, the Chi-square and ANOVA tests were used. There were no statistical differences (P>0.05) between the analyzed parameters. It was concluded that a single dose of 5000IU hCG in stallions did not cause significant changes in the parameters evaluated in different seasons of the year.(AU)

Ultrassonografia testicular no modo color Doppler∕ espectral e avaliação andrológica de garanhões tratados com gonadotrofina coriônica humana (hCG) em diferentes estações do ano / [Testicular ultrasonography in the doppler / spectral color mode and andrological evaluation of stallions treated with human corionic gonadotrofin (hCG) in different seasons of the year]

Utilizaram-se quatro garanhões nos meses de janeiro, abril, julho e outubro de 2016, em dois protocolos, em que: GI (n=4; 5mL de solução salina, i.v.) e GII (n=4; 5000UI de hCG, i.v.) e subdividiram-se esses protocolos em ciclos (C1 e C2), seguindo o esquema crossover, sendo: CI=animais 1 (GI) e 2 (GII), avaliados nos dias D1, D3 e D5, e animais 3 (GI) e 4 (GII), em D2, D4 e D6; CII= animais 1 (GII) e 2 (GI), em D1, D3, D5, e animais 3 (GII) e 4 (GI), em D2, D4 e D6. Realizou-se o tratamento em D1 e D2 de cada ciclo e a ultrassonografia testicular no modo color Doppler e Doppler espectral, uma hora antes de cada coleta de sêmen e imediatamente após. Avaliou-se: número de reflexo de Flehmen, de montas sem ereção, início da monta, tempo de reação à ereção e total da monta e análises de qualidade seminal. Estatisticamente foram utilizados os testes de qui-quadrado e ANOVA. Não houve diferenças estatísticas (P>0,05) entre os parâmetros analisados. Concluiu-se que uma única dose de 5000UI de hCG em garanhões não causou alterações significativas nos parâmetros avaliados em diferentes estações do ano.(AU)

Four stallions were used in January, April, July and October 2016 in two protocols: GI (n=4; 5ml saline, iv) and GII (n=4; 5000 hCG, iv), and these protocols were subdivided into cycles (C1 and C2) following the cross over scheme, as follows: CI=animal 1 (GI) and 2 (GII) evaluated on days D1, D3 and D5 and animal 3 (GI) and 4 (GII) at D 2, D 4 and D 6; CII=animal 1 (GII) and 2 (GI) at D1, D3, D5 and animal 3 (GII) and 4 (GI) at D2, D4 and D6. Treatment was performed on D1 and D2 of each cycle and testicular ultrasound in color Doppler and spectral Doppler mode, one hour before each semen collection and immediately after. We evaluated: Flehmen's reflex number, mounts without erection, start of the mount, reaction time to erection and total mount and seminal quality analyzes. Statistically, the Chi-square and ANOVA tests were used. There were no statistical differences (P>0.05) between the analyzed parameters. It was concluded that a single dose of 5000IU hCG in stallions did not cause significant changes in the parameters evaluated in different seasons of the year.(AU)

Effect of the subdose of human chorionic gonadotropin applied in the Hou Hai acupoint on ovulation induction in mares / [Efeito da subdose de gonadotrofina coriônica humana aplicada no acuponto Hou Hai sobre a indução da ovulação em éguas]

The objective of this study was to evaluate the effects of an hCG sub dose applied at the Hou Hai acupoint on corpus luteum (CL) quality and ovulation induction in mares. Fifteen crossbred mares were distributed in randomized blocks and used in three periods with each period employed as the blocking factor in three treatments: T1 = 1500 IU of hCG via intravenous (IV); T2 = 450 IU of hCG applied at the false acupoint (IV); and T3 = 450 IU of hCG applied at the Hou Hai acupoint. Mean diameter of the CL, serum concentration of progesterone (P4), vascularization of the pre-ovulatory follicle and CL were evaluated. Females administered 450 IU of hCG at the Hou Hai acupoint exhibited greater ovulation rates (33.33%) 48h after induction; The minimum number of colored pixel (NCP) of the pre-ovulatory follicle of control females was superior (40.33) to that of mares administered 450 IU of hCG IV at the false acupoint (36.84) and similar to that of those administered hCG at the Hou Hai acupoint (39.31). Further, moderately positive correlations were found between the CL diameter and the P4 concentration on D8 (P<0.05). IV administration of 450 IU of hCG or at the Hou Hai acupoint was efficient at inducing ovulation and ensuring the quality of CL in mares.(AU)

O objetivo foi avaliar os efeitos de uma subdose de hCG aplicada no acuponto Hou Hai na qualidade do corpo lúteo (CL) e na indução da ovulação em éguas. Quinze éguas mestiças foram distribuídas em blocos ao acaso, sendo o período utilizado como fator de blocagem, em: T1 = 1500 UI de hCG por via intravenosa (IV); T2 = 450 UI de hCG aplicado no falso acuponto (IV) e T3 = 450 UI de hCG aplicada no acuponto Hou Hai. Avaliou-se diâmetro médio do CL, concentração sérica de progesterona (P4), vascularização do folículo pré-ovulatório e do CL. As fêmeas que receberam 450 UI de hCG no acuponto Hou Hai apresentaram maiores taxas de ovulação (33,33%) 48h após a indução. O número de pixels coloridos (NPC) mínimo do folículo pré-ovulatório das fêmeas do grupo controle foi superior (40,33) ao das éguas que receberam 450 UI de hCG IV no falso acuponto (36,84) e semelhante ao das éguas que receberam hCG no acuponto Hou Hai (39,31); correlações moderadamente positivas foram encontradas entre o diâmetro do CL e a concentração de P4, ambos no D8 (P <0,05). A administração IV de 450 UI de hCG ou no acuponto Hou Hai foi eficiente na indução da ovulação e na garantia da qualidade do CL nas éguas.(AU)

Efeito de protocolo hormonal e programa de luz artificial na gestação de éguas receptoras de embrião durante o período de transição de primavera / Effect of hormonal protocol and artificial light program on the pregnancy of embryo recipient mares during spring transition period

O objetivo do estudo foi comparar o efeito das técnicas hormonais e de luz artificial nas éguas receptoras de embrião acíclicas avaliando as taxas de gestação aos 14 e 28 dias durante a fase de transição de primavera. Os 48 animais foram distribuídos aleatoriamente nos grupos: controle (CONT, n=16), éguas cíclicas na fase ovulatória; luz artificial (LUZ, n=16), éguas acíclicas submetidas ao tratamento de luz artificial; e hormônio (HORM, n= 16), éguas acíclicas submetidas ao protocolo hormonal na fase de transição. As éguas do grupo LUZ foram estimuladas por 60 dias com luz artificial durante cinco horas por dia. Nos grupos CONT e LUZ, quando observada a presença de folículo 35 mm de diâmetro e edema uterino grau II, foram administrados 1,5 mg de acetato de deslorelina e 1500 UI de hCG para induzir a ovulação. As éguas do grupo HORM foram tratadas com três doses de 1,5 mg de benzoato de estradiol e seguiram os mesmos protocolos dos Grupos CONT e LUZ. Foi avaliada a taxa de gestação por ultrassonografia aos 14 dias e confirmação aos 28 em todos os grupos experimentais. Foi realizada análise descritiva e teste Qui-quadrado (significância de 5%). Taxas de gestação aos 14 e 28 dias foram semelhantes (p>0,05) entre todos os grupos. Os tratamentos HORM e LUZ durante o período de transição inverno-primavera mostraram-se eficazes para atender ao programa de transferência de embr

The aim of this study was to compare the effect of hormonal and artificial light techniques on acyclic embryo recipient mares by assessing pregnancy rates at 14 and 28 days during the spring transition period. The 48 animals were randomly assigned to the groups: control (CONT, n = 16), cyclic mares in the ovulatory phase; artificial light (LIGHT, n = 16), acyclic mares subjected to artificial light treatment; and hormone (HORM, n = 16), acyclic mares submitted to hormonal protocol in transition phase. In the LIGHT group, mares were stimulated with artificial light for five hours a day, for 60 days. In CONT and LIGHT groups, when a follicle ≥ 35 mm in diameter and uterine edema ≥ grade II were observed, 1.5 mg of deslorelin acetate and 1500 IU hCG were administered to induce ovulation. In the HORM group, mares were treated with three doses of 1.5 mg of estradiol benzoate and followed the same protocols as the CONT and LIGHT groups. Pregnancy rate was assessed by ultrasound at 14 days and confirmation at 28 days in all experimental groups. Descriptive analysis and chi-square test (5% significance) were performed. Pregnancy rates at 14 and 28 days were similar (p> 0.05) among all groups. The HORM and LIGHT treatments during the winter-spring transition period proved to be effective during the embryo transfer programs. As it is a more natural method, the LIGHT protocol has the potential to be one more biotechnological tool in equine reproduction.(AU)

Efeito de protocolo hormonal e programa de luz artificial na gestação de éguas receptoras de embrião durante o período de transição de primavera / Effect of hormonal protocol and artificial light program on the pregnancy of embryo recipient mares during spring transition period

O objetivo do estudo foi comparar o efeito das técnicas hormonais e de luz artificial nas éguas receptoras de embrião acíclicas avaliando as taxas de gestação aos 14 e 28 dias durante a fase de transição de primavera. Os 48 animais foram distribuídos aleatoriamente nos grupos: controle (CONT, n=16), éguas cíclicas na fase ovulatória; luz artificial (LUZ, n=16), éguas acíclicas submetidas ao tratamento de luz artificial; e hormônio (HORM, n= 16), éguas acíclicas submetidas ao protocolo hormonal na fase de transição. As éguas do grupo LUZ foram estimuladas por 60 dias com luz artificial durante cinco horas por dia. Nos grupos CONT e LUZ, quando observada a presença de folículo 35 mm de diâmetro e edema uterino grau II, foram administrados 1,5 mg de acetato de deslorelina e 1500 UI de hCG para induzir a ovulação. As éguas do grupo HORM foram tratadas com três doses de 1,5 mg de benzoato de estradiol e seguiram os mesmos protocolos dos Grupos CONT e LUZ. Foi avaliada a taxa de gestação por ultrassonografia aos 14 dias e confirmação aos 28 em todos os grupos experimentais. Foi realizada análise descritiva e teste Qui-quadrado (significância de 5%). Taxas de gestação aos 14 e 28 dias foram semelhantes (p>0,05) entre todos os grupos. Os tratamentos HORM e LUZ durante o período de transição inverno-primavera mostraram-se eficazes para atender ao programa de transferência de embr

The aim of this study was to compare the effect of hormonal and artificial light techniques on acyclic embryo recipient mares by assessing pregnancy rates at 14 and 28 days during the spring transition period. The 48 animals were randomly assigned to the groups: control (CONT, n = 16), cyclic mares in the ovulatory phase; artificial light (LIGHT, n = 16), acyclic mares subjected to artificial light treatment; and hormone (HORM, n = 16), acyclic mares submitted to hormonal protocol in transition phase. In the LIGHT group, mares were stimulated with artificial light for five hours a day, for 60 days. In CONT and LIGHT groups, when a follicle ≥ 35 mm in diameter and uterine edema ≥ grade II were observed, 1.5 mg of deslorelin acetate and 1500 IU hCG were administered to induce ovulation. In the HORM group, mares were treated with three doses of 1.5 mg of estradiol benzoate and followed the same protocols as the CONT and LIGHT groups. Pregnancy rate was assessed by ultrasound at 14 days and confirmation at 28 days in all experimental groups. Descriptive analysis and chi-square test (5% significance) were performed. Pregnancy rates at 14 and 28 days were similar (p> 0.05) among all groups. The HORM and LIGHT treatments during the winter-spring transition period proved to be effective during the embryo transfer programs. As it is a more natural method, the LIGHT protocol has the potential to be one more biotechnological tool in equine reproduction.

Paeonia lactiflora improves ovarian function and oocyte quality in aged female mice

Although ovarian aging is a key cause of decreased ovarian function and oocyte quality, it remains a problem in infertility treatment. Therefore, this study is aimed to investigate whether Paeonia lactiflora (PL), a herb improves ovarian function and oocyte quality using aged female mice. C57BL/6 female mice aged 8 months were treated orally every day with PL of 26.5 mg/kg (n=7) and 53 mg/kg (n=7) of body weight for 4 weeks using an oral zoned needle. The control group (n=7) was treated with normal saline. Ovaries and serum were collected for the H&E stain and the evaluation of reactive oxygen species (ROS) levels, respectively. In the second experiment, female mice were orally administered with PL (26.5 mg/kg: n=12, 53 mg/kg: n=12, control: n=12) and then superovulated with PMSG and hCG, and mated with male mice. Zygotes were retrieved and cultured for 4 days. Ovaries were provided for examination of expressions of genes associated with angiogenesis (VEGF and visfatin), anti-aging (Sirt1 and Sirt2), and follicular development (c-Kit, BMP-15, and GDF-9). PL significantly increased numbers of surviving follicles (primordial, primary, secondary, and antral), numbers of zygotes retrieved, embryo development rate, and ovarian expression of VEGF, visfatin, c-Kit, BMP-15, and GDF-9 at both doses. However, ovarian expression of Sirt1 and Sirt2 was increased at 53.0 mg/kg of PL. ROS levels were not affected by PL. These results suggest that PL may possess beneficial effects regarding ovarian function and oocyte quality, possibly by activation of ovarian angiogenesis and follicular development.

Paeonia lactiflora improves ovarian function and oocyte quality in aged female mice

Although ovarian aging is a key cause of decreased ovarian function and oocyte quality, it remains a problem in infertility treatment. Therefore, this study is aimed to investigate whether Paeonia lactiflora (PL), a herb improves ovarian function and oocyte quality using aged female mice. C57BL/6 female mice aged 8 months were treated orally every day with PL of 26.5 mg/kg (n=7) and 53 mg/kg (n=7) of body weight for 4 weeks using an oral zoned needle. The control group (n=7) was treated with normal saline. Ovaries and serum were collected for the H&E stain and the evaluation of reactive oxygen species (ROS) levels, respectively. In the second experiment, female mice were orally administered with PL (26.5 mg/kg: n=12, 53 mg/kg: n=12, control: n=12) and then superovulated with PMSG and hCG, and mated with male mice. Zygotes were retrieved and cultured for 4 days. Ovaries were provided for examination of expressions of genes associated with angiogenesis (VEGF and visfatin), anti-aging (Sirt1 and Sirt2), and follicular development (c-Kit, BMP-15, and GDF-9). PL significantly increased numbers of surviving follicles (primordial, primary, secondary, and antral), numbers of zygotes retrieved, embryo development rate, and ovarian expression of VEGF, visfatin, c-Kit, BMP-15, and GDF-9 at both doses. However, ovarian expression of Sirt1 and Sirt2 was increased at 53.0 mg/kg of PL. ROS levels were not affected by PL. These results suggest that PL may possess beneficial effects regarding ovarian function and oocyte quality, possibly by activation of ovarian angiogenesis and follicular development.(AU)

Efeito de protocolo hormonal e programa de luz artificial na gestação de éguas receptoras de embrião durante o período de transição de primavera / Effect of hormonal protocol and artificial light program on the pregnancy of embryo recipient mares during spring transition period

O objetivo do estudo foi comparar o efeito das técnicas hormonais e de luz artificial nas éguas receptoras de embrião acíclicas avaliando as taxas de gestação aos 14 e 28 dias durante a fase de transição de primavera. Os 48 animais foram distribuídos aleatoriamente nos grupos: controle (CONT, n=16), éguas cíclicas na fase ovulatória; luz artificial (LUZ, n=16), éguas acíclicas submetidas ao tratamento de luz artificial; e hormônio (HORM, n= 16), éguas acíclicas submetidas ao protocolo hormonal na fase de transição. As éguas do grupo LUZ foram estimuladas por 60 dias com luz artificial durante cinco horas por dia. Nos grupos CONT e LUZ, quando observada a presença de folículo ≥ 35 mm de diâmetro e edema uterino ≥ grau II, foram administrados 1,5 mg de acetato de deslorelina e 1500 UI de hCG para induzir a ovulação. As éguas do grupo HORM foram tratadas com três doses de 1,5 mg de benzoato de estradiol e seguiram os mesmos protocolos dos Grupos CONT e LUZ. Foi avaliada a taxa de gestação por ultrassonografia aos 14 dias e confirmação aos 28 em todos os grupos experimentais. Foi realizada análise descritiva e teste Qui-quadrado (significância de 5%). Taxas de gestação aos 14 e 28 dias foram semelhantes (p>0,05) entre todos os grupos. Os tratamentos HORM e LUZ durante o período de transição inverno-primavera mostraram-se eficazes para atender ao programa de transferência de embrião. Por ser um método mais natural, o protocolo LUZ tem potencial como mais uma ferramenta biotecnológica na reprodução de equinos.

The aim of this study was to compare the effect of hormonal and artificial light techniques on acyclic embryo recipient mares by assessing pregnancy rates at 14 and 28 days during the spring transition period. The 48 animals were randomly assigned to the groups: control (CONT, n = 16), cyclic mares in the ovulatory phase; artificial light (LIGHT, n = 16), acyclic mares subjected to artificial light treatment; and hormone (HORM, n = 16), acyclic mares submitted to hormonal protocol in transition phase. In the LIGHT group, mares were stimulated with artificial light for five hours a day, for 60 days. In CONT and LIGHT groups, when a follicle ≥ 35 mm in diameter and uterine edema ≥ grade II were observed, 1.5 mg of deslorelin acetate and 1500 IU hCG were administered to induce ovulation. In the HORM group, mares were treated with three doses of 1.5 mg of estradiol benzoate and followed the same protocols as the CONT and LIGHT groups. Pregnancy rate was assessed by ultrasound at 14 days and confirmation at 28 days in all experimental groups. Descriptive analysis and chi-square test (5% significance) were performed. Pregnancy rates at 14 and 28 days were similar (p> 0.05) among all groups. The HORM and LIGHT treatments during the winter-spring transition period proved to be effective during the embryo transfer programs. As it is a more natural method, the LIGHT protocol has the potential to be one more biotechnological tool in equine reproduction.