Improvement of embryo production by the replacement of the last two doses of porcine follicle-stimulating hormone with equine chorionic gonadotropin in Sindhi donors

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Use of a Low Dose of Equine Purified FSH to Induce Multiple Ovulations in Mares

The effects of a low dose of equine purified FSH (eFSH) on incidence of multiple ovulations and embryo recovery rate in mares were studied. During the physiological breeding season in Brazil (19 degrees 45'45'S), 14 Mangalarga Marchador donor mares were used in a crossover study and another 25 mares of the same breed, between 3 years and 12 years of age were used as recipients for the embryo transfers. Donors were monitored during two consecutive oestrus cycles, an untreated control cycle followed by a treated cycle, when eFSH was administered. In both cycles, after an embryo collection attempt on day 8 post-ovulation all mares received 7.5 mg dinoprost and had their two largest follicles tracked daily by ultrasonography until the period of ovulation. Mares were inseminated every 48 h with extended fresh semen from a single stallion after the identification of a 35-mm follicle until the period of ovulation. Ovulations were induced by intravenous administration of 2.500 IU of human chorionic gonadotropin, upon detection of a 35- to 40-mm follicle. In the treated cycle, 5 mg eFSH was given intramuscularly once a day, from day 8 post previous ovulation until at least one follicle reached 35 mm in diameter. Embryo flushes were performed on day 8 of dioestrus (day 0 = ovulation). Treatment with eFSH resulted in higher (p < 0.05) ovulation rate and incidence of multiple ovulations compared to the control (1.6 vs 1.0 and 50% vs 0%, respectively - one mare had triple ovulation). However, embryo recovery rates in the control and treated cycles were similar (0.8 and 1.0, respectively; p > 0.05). Pregnancy rates in the recipient mares following embryo transfer were similar for the control and eFSH cycles (11/11 and 10/14, respectively). Additional studies are necessary in order to develop a low-dose protocol for the use of eFSH that brings a more consistent contribution to the efficiency of commercial equine embryo transfer programs.

Embryo Transfer Efficiency of Quarter Horse Athletic Mares

This study was designed to compare embryo recovery rates and pregnancy rates of athletic and breeding Quarter Horse mares in a tropical warm climate. Thirty-nine barrel racing mares in training and 135 breeding mares as control donors were included. During the training period, the ambient temperature ranged from 31 degrees C to 36 degrees C and the average humidity from 70% to 90%. After the detection of a 35-mm follicle by ultrasound, ovulation was induced with 1 mg of deslorelin acetate (i.m), and insemination was performed 24 hours later with cooled and fresh semen from different fertile stallions. Embryos were collected on day 8 postovulation. The body temperature (rectal) was evaluated from eight athletic donor mares randomly selected from the same studied group. A total of 138 and 657 embryo collections were carried out on training and breeding mares, respectively, with a total of 105 (76%) and 466 (71%) embryos collected (P > .05). Similarly, no differences (P > .05) were observed for the pregnancy rates on day 15 (82/105, 78% vs. 370/466,79%), and day 40 (73/105, 69% vs. 328/466,70%) between the training and breeding donor mares. Just after training, the body temperature increased to an average of 39.4 degrees C and the respiratory rate from 14.5 to 35.3 breaths per minute. The results of the present study showed that embryo production from appropriately trained donor mares in good condition were similar to non-athletic broodmares. (C) 2011 Published by Elsevier B.V.

Ovarian function in cows submitted to acute stress during proestrus

Studies in which ACTH was administrated in heifers after the occurrence of luteolysis showed a delay in the onset of estrus and the estrus duration was shortened. This study evaluated the effect of acute stress by road transportation on estrous behavior and ovulation, monitored by serum progesterone and cortisol concentrations in cows at the periovulatory period, using a crossover design. Eleven crossbred cows, divided into Control and Transport groups had their estrus cycle synchronized with GnRH, an intravaginal progesterone device, and cloprostenol. Thirty hours after withdrawal of the device, the animals of the Transport group were transported for 60 min by truck and those from the Control group remained at pasture. Ovarian ultrasound examination was performed every 12 h from device withdrawal until ovulation in every cow. From the day after removal of the device until ovulation estrous behavior was monitored 24 h a day. Blood samples for serum cortisol and progesterone concentrations were taken at -90, -60, 0, 30, 60 and 180 min in relation to the end of transportation. Transportation during the estrous period induced stress in cows as reflected by changes in serum concentrations of progesterone and cortisol. However, we did not detect impairment in estrus expression, estrus duration or ovulation (P>0.05). (C) 2010 Elsevier B.V. All rights reserved.

Immunohistochemical detection of receptors for oestrogen and progesterone in endometrial glands and stroma during the oestrous cycle in Nelore (Bos taurus indicus) cows

The aim of the present study was to monitor endometrial distribution and concentrations of oestrogen receptors alpha (ER alpha) and progesterone receptors (PR) by immunohistochemistry in Nelore cows (Bos taurus indicus) during the oestrous cycle. Blood samples were collected for progesterone measurement and endometrial samples were taken from the uterine horn contra lateral to the corpus luteum in 16 cows at days 0 (ovulation), 5, 9, 13 and 19 of the oestrous cycle. Immunostaining evaluation for ER alpha and PR in the glandular epithelium and uterine stroma was performed by two methods: positive nuclei counting and staining intensity of the nuclei. Specific positive staining reactions for both receptors were limited to cell nuclei and they were not identified in the cytoplasm. The proportion of ER alpha positive nuclei had a temporal variation throughout the oestrous cycle in both cell types evaluated and was higher in uterine stroma than the glandular epithelium (p < 0.05). The greatest proportion of ER alpha stained nuclei was observed at oestrus and during the initial and mid luteal phase (days 5, 9 and 13) (p < 0.05) in the glandular epithelium and at days 0, 5 and 9 in the uterine stroma (p < 0.01). The proportion of PR positive nuclei remained constant throughout the entire oestrous cycle for both cell types evaluated (p > 0.05). A higher proportion of PR positive nuclei was measured in the uterine stroma compared with the glandular epithelium (p < 0.05). Intensity of staining for ER alpha and PR varied throughout the oestrous cycle (p < 0.01). There was a higher staining intensity at days 0 and 5 in the stroma for ER alpha (p < 0.01) and PR (p < 0.01) and in the glandular epithelium at days 0, 5, 9 and 13 for ER alpha (p < 0.01) and at days 0, 5 and 9 for PR (p < 0.01) when compared with the other evaluated days. These data demonstrate that ER alpha and PR expression varied throughout the oestrous cycle in Nelore cows, in general with highest concentrations at oestrus and the lowest during the luteal phase. This is similar to patterns observed in Bos taurus taurus.

Follicular Development in Ewes During Natural and Prostaglandins Induced Estrous Cycle

The follicular development was evaluated in ovine females during natural and prostaglandin-F(2 alpha) (PG) induced estrous cycle. Ewes were randomly divided in two treatments (n=7/treatment): T1 with natural cycle and T2 synchronized with two injections of PG. From one day before PG injection until next ovulation, daily transrectal ultrasonography was done. All follicles >= 2 mm were assessed. During the interovulatory intervals, follicular growth and regression occurred in a wave like pattern (2-3 waves). The maximum diameter of the largest follicle of the first wave was greater in T1 (5.83 +/- 0.31 mm) compared with T2 (5.0 +/- 0.1 mm; P<0.01), but there was no significant difference among the emergency day of largest follicle, during the growth phase of the follicular waves. The duration of the plateau phase in wave 2 differed between the two treatments (P<0.05) showing 0.83 +/- 0.31 and 1.83 +/- 0.17 d, for natural and synchronized treatment, respectively. Growth rate did not differ between treatments. Presence of new luteal tissue was detected on day 3 after ovulation. In conclusion, the follicular development was similar in female ovine during natural and PG induced estrous cycle.

Reuse of norgestomet implants in an eCG-based superovulation protocol administered to Nelore (Bos taurus indicus) cows

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ovarian activity reversibility after the use of deslorelin acetate as a short-term contraceptive in domestic queens

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ultrasonographic and endocrine aspects of follicle deviation, and acquisition of ovulatory capacity in buffalo (Bubalus bubalis) heifers

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effects of two estradiol esters (benzoate and cypionate) on the induction of synchronized ovulations in Bos indicus cows submitted to a timed artificial insemination protocol

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Follicle deviation and ovulatory capacity in Bos indicus heifers

The objectives of Experiment I were to determine the interval from ovulation to deviation, and diameter of the dominant follicle (DF) and largest subordinate follicle (SF) at deviation in Nelore (Bos indicus) heifers by two methods (observed and calculated). Heifers (n = 12) were examined ultrasonographically every 12 h from ovulation (Day 0) to Day 5. The time of deviation and diameter of the DF and largest SF at deviation did not differ (P > 0.05) between observed and calculated methods. Overall, deviation occurred 2.5 +/- 0.2 d (mean +/- S.E.M.) after ovulation, and diameters for DF and largest SF at deviation were 6.2 +/- 0.2 and 5.9 +/- 0.2 mm, respectively. Experiment 2 was designed to determine the size at which the DF acquires ovulatory capacity in B. indicus heifers. Twenty-nine heifers were monitored every 24 h by ultrasonography, from ovulation until the DF reached diameters of 7.0-8.4 mm (n = 9), 8.5-10.0 mm (n = 10), or >10.0 mm (n = 10). At that time, heifers were treated with 25 mg of pLH and monitored by ultrasonography every 12 h for 48 h. Ovulation occurred in 3 of 9, 8 of 10, and 9 of 10 heifers, respectively (P < 0.05). In summary, there was no significant difference between observed and calculated methods of determining the beginning of follicle deviation. Deviation occurred 2.5 d after ovulation when the DF reached 6.2 mm, and ovulatory capacity was acquired by DF as small as 7.0 mm. (c) 2008 Elsevier B.V. All rights reserved.

Timed artificial insemination in beef cattle using GnRH agonist, PGF2alpha and estradiol benzoate (EB)

The present work evaluated low-cost protocols for timed artificial insemination (TAI) in beef cattle. In Experiment 1, cycling nonlactating Nelore cows (Bos indicus, n=98) were assigned to the following groups: GnRH-PGF (GP) and GnRH-PGF-GnRH (GPG), whereas cycling (n=328, Experiment 2) or anestrus (n = 225, Experiment 3) lactating (L) cows were divided into 3 groups: GP-L, GPG-L and GnRH-PCF-Estradiol benzoate (GPE-L). In Experiment 4, lactating cows (n=201) were separated into 3 groups: GP-L, GPE-L and G 1/2PE-L. Animals from Experiment 1, 3 and 4 were treated (Day 0), at random stages of the estrous cycle, with 8 mug of buserelin acetate (GnRH agonist) intramuscularly (im), whereas in Experiment 2 half of the cows received 8 and the other half 12 mug of GnRH (im). Seven days later (D 7) all animals were treated with 25 mg of dinoprost trometamine (PGF2 alpha, im) except those cows from the G 1/2PE-L group which received only 1/2 dose of PGF2 alpha (12.5 mg) via intravulvo-submucosa (ivsm). Alter PGF2 alpha injection the animals from the control groups (GP and GP-L) were observed twice daily to detect estrus and Al was performed 12 h afterwards. The cows from the other groups received a second GnRH injection (D 8 in GPG-L and d9 in GPG groups) or one injection of estradiol benzoate (EB, 1.0 mg, D 8 in GPE-L group). All cows from GPG and GPG-L or GPE-L groups were AI 20 to 24 or 30 to 34 h, respectively, after the last hormonal injection. Pregnancy was determined by ultrasonography or rectal palpation 30 to 50 days after AI. In the control groups (GP and GP-L) percentage of animals detected in heat (44.5 to 70.3%) and pregnancy rate (20 to 42%) varied according to the number of animals with corpus luteum (CL) at the beginning of treatment. The administration of a second dose of GnRH either 24 (Experiment 2) or 48 h (Experiment 1) after PGF2 alpha resulted in 47.7 and 44.9% pregnancy rates, respectively, after TAI in cycling animals. However, in anestrus cows the GPG treatment induced a much lower pregnancy rate (14.9%) after TAI. The replacement of the second dose of GnRH by EB (GPE-L) resulted in a pregnancy rate (43.3%) comparable to that obtained after GnRH treatment (GPG-L, 47.7%, Experiment 2). Furthermore, the use of 1/2 dose of PGF2 alpha (12.5 mg ivms, Experiment 4) resulted in pregnancy rate (43.5%) similar to that observed with the full dose (im). Both protocols GPG and GPE were effective in synchronizing ovulation in cycling Nelore cows and allowed approximately a 45% pregnancy rate after TAI. Additionally, the GPE treatment is a promising alternative to the use of GPG in timed Al of beef cattle, due to the low cost of EB when compared to GnRH agonists. (C) 2001 by Elsevier B.V.

Use of knowledge regarding LH receptors to improve superstimulatory treatments in cattle

Embryo transfer is a biotechnology that has been used worldwide to increase the production of offspring from female bovines. Treatments to induce multiple ovulations (superovulation) have evolved from superstimulatory protocols that depended upon detection of oestrus to treatments that synchronise follicle growth and ovulation, allowing for improved donor management and fixed-timed AI (FTAI). The protocols associated with FTAI facilitate animal handling and produce at least as many viably embryos as conventional treatment protocols that required detection of oestrus. Recent knowledge regarding LH receptors (LHR) and follicular development can be applied to improve embryo transfer protocols. In fact, improvements in the superstimulatory treatment called the 'P-36 protocol', which include hormones that stimulate LHR, indicate that adjustments related to LHR availability may increase bovine embryo yield compared with conventional protocols based on the detection of oestrus.

Expression of protease nexin-1 and plasminogen activators during follicular growth and the periovulatory period in cattle

Extracellular matrix remodeling occurs during ovarian follicular development, mediated by plasminogen activators (PAs) and PA inhibitors including protease nexin-1 (PN-1). In the present study we measured expression/activity of the PA system in bovine follicles at different stages of development by timed collection of ovaries during the first follicular wave and during the periovulatory period, and in follicles collected from an abattoir. The abundance of mRNA encoding PN-1, tissue-type PA (tPA), urokinase (uPA) and PA inhibitor-1 (PAI-1) were initially upregulated by human chorionic gonadotropin (hCG) in bovine preovulatory follicular wall homogenates. PN-1, PAI-1 and tPA mRNA expression then decreased near the expected time of ovulation, whereas uPA mRNA levels remained high. PN-1 concentration in follicular fluid (FF) decreased and reached the lowest level at the time of ovulation, whereas plasmin activity in FF increased significantly after hCG. Follicles collected from the abattoir were classified as non-atretic, early-atretic or atretic based on FF estradiol and progesterone content: PN-1 protein levels in FF were significantly higher in non-atretic than in atretic follicles, and plasmin activity was correspondingly higher in the atretic follicles. No changes in PN-1 levels in FF were observed during the growth of pre-deviation follicles early in a follicular wave. These results indicate that PN-1 may be involved in the process of atresia in non-ovulatory dominant follicles and the prevention of precocious proteolysis in periovulatory follicles.

The fibroblast growth factor family: involvement in the regulation of folliculogenesis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)