Inhibition of FGF receptor impairs primitive endoderm differentiation in bovine embryos.

The first cellular differentiation event in the pre-implantation embryo results in the trophectoderm (TE) and the inner cell mass (ICM). A second event occurs in the latter, resulting in the epiblast and the primitive endoderm (PE). This second differentiation is still not fully characterized in bovine development, although it is likely to involve FGF signalling. Thus, in this study, we tested the hypothesis that stimulation or inhibition of the FGF pathway during bovine embryo in vitro culture would only interfere with PE differentiation if maintained until later blastocyst stages. At first, we characterized the expression of PE marker SOX17 at different blastocyst stages. Then, we treated in vitro produced embryos during different windows of time: days 5.0-7.0 (D5-D7), D7-D9, and D5-D9 with 1 µg/ml FGF4 and 1 µg/ml heparin or 1 mM FGFR inhibitor, AZD4547. We observed that the SOX17-positive cell number only increases in late-stage blastocysts compared to early stages. Treatment of embryos with FGF4 did not change the number of SOX17-positive cells, while inhibition of FGFR signalling reduced SOX17-positive cells from D5-D7 and completely ablated SOX17 expression when kept until D9. In conclusion, FGFR inhibition repressed PE differentiation in bovine embryos at all time points, although stimulation with FGF4 did not interfere with PE cell numbers.

Low invasive estrous synchronization protocol for wild animals: an example with melengestrol acetate in brown brocket deer (Mazama gouazoubira).

Deer are sensitive to stressful stimuli by handling and their reproductive physiology could be altered by these procedures, making it necessary to develop less invasive protocols for ART. Melengestrol acetate (MGA), a synthetic progestin administered orally, appears as an alternative for estrous synchronization protocols (ESP), such as reported in cattle. Firstly, we compared two MGA doses (0.5 and 1.0 mg/day/animal), which would have suppression effect in estrous behavior (EB). Eight females were randomly and equally distributed in Group 1 (G1) and Group 2 (G2), which received 0.5 and 1.0 mg/day/animal respectively for 15 days (D1 to D15). Two cloprostenol (CP) applications were performed on D0 and D11. Estrus detection (ED) was performed every day. All females from G1 displayed estrus during treatment period, whereas all females from G2 displayed estrus after treatment, suggesting a suppressive effect of 1.0 mg in the EB. Once the suppressive MGA dose (1.0 mg) was defined, we used this dose for assessing ESP. The same eight females received 1.0 mg/animal for eight days (D-8 to D-1), followed by 0.25 mg of estradiol benzoate on D-8 and 265 µg of CP on D0. Feces for fecal progesterone metabolites (FPM) measurement were collected from D0 until seven days after the last day of estrus. Seven females displayed estrus between 12 and 72 h after CP application, which was followed by a significant increase in FPM levels (except female MG6), suggesting the formation of corpus luteum. After ED, females were placed with a fertile male to assess the fertility of the protocol. Pregnancy was confirmed by ultrasound 30 days after mating in 3/6 individuals. Although the low effectiveness of MGA protocol, it should be considered as a promising alternative in deer ESP since this protocol has less stressful effect on the animal during reproductive management when compared to other ESP.

Low invasive estrous synchronization protocol for wild animals: an example with melengestrol acetate in brown brocket deer (Mazama gouazoubira)

Deer are sensitive to stressful stimuli by handling and their reproductive physiology could be altered by these procedures, making it necessary to develop less invasive protocols for ART. Melengestrol acetate (MGA), a synthetic progestin administered orally, appears as an alternative for estrous synchronization protocols (ESP), such as reported in cattle. Firstly, we compared two MGA doses (0.5 and 1.0 mg/day/animal), which would have suppression effect in estrous behavior (EB). Eight females were randomly and equally distributed in Group 1 (G1) and Group 2 (G2), which received 0.5 and 1.0 mg/day/animal respectively for 15 days (D1 to D15). Two cloprostenol (CP) applications were performed on D0 and D11. Estrus detection (ED) was performed every day. All females from G1 displayed estrus during treatment period, whereas all females from G2 displayed estrus after treatment, suggesting a suppressive effect of 1.0 mg in the EB. Once the suppressive MGA dose (1.0 mg) was defined, we used this dose for assessing ESP. The same eight females received 1.0 mg/animal for eight days (D-8 to D-1), followed by 0.25 mg of estradiol benzoate on D-8 and 265 g of CP on D0. Feces for fecal progesterone metabolites (FPM) measurement were collected from D0 until seven days after the last day of estrus. Seven females displayed estrus between 12 and 72 h after CP application, which was followed by a significant increase in FPM levels (except female MG6), suggesting the formation of corpus luteum. After ED, females were placed with a fertile male to assess the fertility of the protocol. Pregnancy was confirmed by ultrasound 30 days after mating in 3/6 individuals. Although the low effectiveness of MGA protocol, it should be considered as a promising alternative in deer ESP since this protocol has less stressful effect on the animal during reproductive management when compared to other ESP.(AU)

Low invasive estrous synchronization protocol for wild animals: an example with melengestrol acetate in brown brocket deer (Mazama gouazoubira)

Deer are sensitive to stressful stimuli by handling and their reproductive physiology could be altered by these procedures, making it necessary to develop less invasive protocols for ART. Melengestrol acetate (MGA), a synthetic progestin administered orally, appears as an alternative for estrous synchronization protocols (ESP), such as reported in cattle. Firstly, we compared two MGA doses (0.5 and 1.0 mg/day/animal), which would have suppression effect in estrous behavior (EB). Eight females were randomly and equally distributed in Group 1 (G1) and Group 2 (G2), which received 0.5 and 1.0 mg/day/animal respectively for 15 days (D1 to D15). Two cloprostenol (CP) applications were performed on D0 and D11. Estrus detection (ED) was performed every day. All females from G1 displayed estrus during treatment period, whereas all females from G2 displayed estrus after treatment, suggesting a suppressive effect of 1.0 mg in the EB. Once the suppressive MGA dose (1.0 mg) was defined, we used this dose for assessing ESP. The same eight females received 1.0 mg/animal for eight days (D-8 to D-1), followed by 0.25 mg of estradiol benzoate on D-8 and 265 g of CP on D0. Feces for fecal progesterone metabolites (FPM) measurement were collected from D0 until seven days after the last day of estrus. Seven females displayed estrus between 12 and 72 h after CP application, which was followed by a significant increase in FPM levels (except female MG6), suggesting the formation of corpus luteum. After ED, females were placed with a fertile male to assess the fertility of the protocol. Pregnancy was confirmed by ultrasound 30 days after mating in 3/6 individuals. Although the low effectiveness of MGA protocol, it should be considered as a promising alternative in deer ESP since this protocol has less stressful effect on the animal during reproductive management when compared to other ESP.

Stress in captive Blue-fronted parrots (Amazona aestiva): the animalists' tale.

Understanding stress physiology is crucial for species management because high levels of stress can reduce reproduction and the individual's ability to face threats to survive. One of the most popular methods for non-invasive monitoring of animal endocrine status is the glucocorticoid (GC) metabolite measurements, which can provide important information about how animals are affected by their surrounding environment. Here, we carried out the biological validation of corticosterone enzyme immunoassays (EIAs), which together with a cortisol EIA was used to quantified the concentrations of urofaecal GC metabolites (uGCMs) in wild and captive Blue-fronted amazon parrots (Amazona aestiva). Urofaecal GC concentrations were significantly higher (P < 0.05) in free-living parrots (157.9 ± 18.5 ng cortisol/g and 61.14 ± 23.5 ng corticosterone/g dry urofaecal sample) than in those kept in captivity, which showed the comparable levels of GC metabolites independently of the management system applied. The higher uGCM levels obtained in the wild population point to an adaptive response for survival and species propagation in a more challenging environment, in comparison with captive animals. Furthermore, the lower uGCM concentrations in captive parrots may indicate an adaptive capacity of the species A. aestiva to captivity and its potential as a legal pet. The corticosterone EIA applied in this study proved to be an effective technique for the adrenocortical activity monitoring in this species. We discuss our findings considering the management and destiny given to wild-caught birds that are kept in confinement or returned to nature.

Non-homogeneous distribution of steroids in fecal pellets: An example in brown brocket deer (Mazama gouazoubira) with progesterone metabolites.

Measuring reproductive hormones in feces has become an important tool in the endocrine characterization of wild animals' reproduction. However, several factors may influence its success, such as fecal collection and storage techniques, knowledge of steroid hormone metabolism, the extraction procedure, immunoassay selection, inherent factors, and the distribution of steroid hormones in the feces. It is known that the distribution of these hormones in the feces is not homogeneous, and prior to the extraction of the steroidal metabolites, homogenization of the feces is recommended. Hormonal analysis is based on only a small fraction of the feces, which in theory should be representative of the total. In the case of cervids and other ruminants, feces consist of pellets. Here, the concentration of the steroid metabolites of each pellet was measured in order to evaluate the distribution of the fecal progesterone metabolites concentration in 10 pellets/fecal mass from five female Mazama gouazoubira. There were large variations in fecal progesterone metabolites concentrations between the pellets of the same feces/female, showing the following amplitude variations: Animal A: 112%; Animal B: 164%; Animal C: 115%; Animal D: 62%; Animal E: 108%. These results show the importance of adequate homogenization prior to steroid metabolite extraction.