Influence of Superovulatory Protocols on In Vitro Production of Nellore (Bos indicus) Embryos

There are indications in the literature that delaying the period between ovarian superestimulation and ovum pick up (OPU) would induce follicles to a condition of initial atresia, which could be beneficial to oocyte development. In this work, we compared three protocols for OPU and in vitro production (IVP) of embryos, in Nellore cattle. Nellore cows (n = 18) were randomly allocated in three groups: Group 1 (OPU), Group 2 [Follicle stimulating hormone (FSH) and OPU] and Group 3 (FSH deprivation and OPU). Three OPUs were performed, and the animals were switched to a different group each time (crossover), in such a way that at the end of the experiment all cows received the 3 protocols. At random stage of the oestrous cycle (D‐2), all follicles ≥ 6 mm were aspirated to induce a new follicular wave 2 days afterwards (D0). In Group 1, OPU was performed on D2 and oocytes were processed to IVP. In Group 2, starting on D0, cows were superstimulated (FSH, Folltropin^®, 30 mg administered daily, i.m., during three consecutive days, total dose = 180 mg), and 6 h after the last FSH dose, they received exogenous luteinizing hormone (LH) (12.5 mg, i.m., Lutropin^®, D3). The OPU was performed 6 h after LH administration, i.e. 12 h after the last dose of FSH. Animals in Group 3 received the same treatment as those in Group 2, except that LH was administered 42 h after the last dose of FSH, and OPU occurred 6 h later. Therefore, in this group, follicles were deprived of FSH at 48 h. Both cleavage and blastocyst rates were similar (p > 0.05, anova ) among oocytes from Groups 1, 2 and 3, respectively: 77.4% (144/185) and 42.70% (79/185); 75.54% (105/139) and 31.65% (44/139); 63.52% (101/159) and 33.33% (53/159). However, hatched blastocyst rate was higher (p < 0.01) in Group 1 (30.27%, 56/185) when compared with Group 2 (11.51%, 16/139) or 3 (15.72%, 25/159). It is concluded that, contrary to previous work on European breeds (Bos taurus), ovarian superstimulation associated with deprivation of FSH and OPU (Group 3) did not increase IVP of Nellore embryos (Bos indicus). On the contrary, the highest hatched blastocyst rates were observed in oocytes from non‐superstimulated cows.     

13,14‐Dihydro‐15‐Keto Prostaglandin F2α Release in Response to Oxytocin Challenge Early Post‐Partum in Anoestrous Nelore Cows Submitted to Temporary Calf Removal and Progesterone Priming

The study evaluated, in early post‐partum anoestrous Nelore cows, if the increase in plasma oestradiol (E2) concentrations in the pre‐ovulatory period and/or progesterone priming (P4 priming) preceding ovulation, induced by hormonal treatment, reduces the endogenous release of prostaglandin PGF₂αand prevents premature lysis of the corpus luteum (CL). Nelore cows were subjected to temporary calf removal for 48 h and divided into two groups: GPE/eCG group (n = 10) and GPG/eCG group (n = 10). Animals of the GPE/eCG group were treated with a GnRH agonist. Seven days later, they received 400 IU of eCG, immediately after PGF₂α treatment, and on day 0, 1.0 mg of oestradiol benzoate (EB). Cows of the GPG/eCG group were similarly treated as those of the GPE/eCG group, except that EB was replaced with a second dose of GnRH. All animals were challenged with oxytocin (OT) 9, 12, 15 and 18 days after EB or GnRH administration and blood samples were collected before and 30 min after OT. Irrespective of the treatments, a decline in P4 concentration on day 18 was observed for cows without P4 priming. However, animals exposed to P4 priming, treated with EB maintained high P4 concentrations (8.8 ± 1.2 ng/ml), whereas there was a decline in P4 on day 18 (2.1 ± 1.0 ng/ml) for cows that received GnRH to induce ovulation (p < 0.01). Production of 13,14‐dihydro‐15‐keto prostaglandin F₂α (PGFM) in response to OT increased between days 9 and 18 (p < 0.01), and this increase tended to be more evident in animals not exposed to P4 priming (p < 0.06). In conclusion, the increase in E2 during the pre‐ovulatory period was not effective in inhibiting PGFM release, which was lower in P4‐primed than in non‐primed animals. Treatment with EB promoted the maintenance of elevated P4 concentrations 18 days after ovulation in P4‐primed animals, indicating a possible beneficial effect of hormone protocols containing EB in animals with P4 priming.     

Closure of large patent ductus arteriosus with a self-expanding duct occluder in two dogs

Of the different catheterisation methods described for closure of patent ductus arteriosus (PDA), coil embolisation is most commonly used in dogs. However, for a PDA larger than 4 to 5 mm in diameter, coil implantation is difficult. For these cases, the Amplatzer duct occluder (ADO) offers an alternative method. This report describes the successful implantation of an ADO in two dogs with large PDAs of approximately 6 mm diameter. The self-expandible device attached to an implantation wire was advanced through a long sheath antegrade to the femoral vein through the right heart and pulmonary artery to the duct and delivered into the PDA. Thereafter the device was released by unscrewing it from the delivery cable. The large PDA in both dogs was totally occluded by these means without any residual shunt. Thus, the ADO is a controlled release implant that also allows occlusion of a large PDA. Its high costs limit its general use in veterinary medicine at the present time.     

Uptake and assimilation of atmospheric NO2 — N by spruce needles (Picea abies): A field study

NO₂ enters spruce needles by gas exchange through the slomata. Nitrate formed from NO₂ is reduced in the cytosol by nitrate reductase (NR), the rate limiting enzyme of the nitrogen assimilatory pathway. A linear relationship was found between the nitrate reductase activity (NRA), NO₂ concentration and the amount of N incorporated into amino acids and proteins, so that NRA was suggested as an estimate of NO₂-uptake. In the present field study, 50 spruce trees (Picea abies) have been selected, which grow in a natural habitat in a NO₂ concentration gradient in a forest crossed by a highway which is a major NO source. At part of the sites, the microclimatic conditions have been recorded, so that common models of local gas exchange of the needles could be used to estimate stomatal uptake of NO₂. NRA was investigated as a function of radiation and stomatal uptake on the day before needle sampling. Close to the highway NRA was permanently elevated with a maximum in summer. As with the laboratory results, a linear relationship between stomatal uptake and NRA was found. Total N — content of current year shoots was not affected by the additional N-source provided by airborne NO₂. The present study shows that the gas exchange models are consistent with the physiological reactions of spruce needles on a local level and therefore contribute to the validation of calculations of NO₂ dry deposition to spruce forests.     

Cryopreservation of Sheep Primordial Follicles

The aim of this study was to evaluate the efficiency of 1 M dimethylsulphoxide (DMSO), ethylene glycol (EG), propylene glycol (PROH) and glycerol (GLY) to cryopreserve primordial follicles. The first evaluation was performed soon after cryopreservation and the second evaluation after 4 days of in vitro culture, using the cryoprotectants that allowed the higher results (higher follicular survival rate) after cryopreservation. The results after follicular isolation (control) and cryopreservation using 1 M DMSO, EG, PROH and GLY showed that the mean number (+/- SEM) of live follicles per millilitre was 3204 (100%) +/- 319.27, 2798 (87%) +/- 239.14, 2492 (78%) +/- 345.8, 448 (14%) +/- 46.3 and 208 (7%) +/- 75.26, respectively. Higher follicular survival was reported when DMSO and EG were used. Control follicles and follicles cryopreserved with these two cryoprotectants were cultured and the percentage of follicular survival was 55% (control), 42% (EG) and 34% (DMSO). Similar results were found between control and follicles cryopreserved with EG. In conclusion, 1 M EG is the most effective cryoprotectant to preserve primordial follicles isolated from ovaries of sheep.