Follicular dynamics and production of oocytes in young Nellore heifers with energetic supplementation

To verify the effects of energy supplementation and fat on follicular dynamics, metabolic profile and the production of oocytes, 15 young heifers (Bos taurus indicus) with an average age of 14 months and with an initial weight of 256.35 kg were assigned to two treatments according to weight and follicular population as evaluated by ultrasonography: in T1 (1.0 × M), animals received 100% of the energy requirements for maintenance; in T2 (1.7 × M), animals received 170% of the energy requirements for maintenance, achieved by the addition of 200 g of Megalac®. After a period of adaptation to the diet, the treatments, blood collection and follicular aspirations were started using a randomized design. The dry matter intake and weight gain were lower in the 1.0 × M group than in the 1.7 × M group. No differences were found in the plasma progesterone concentrations, albumin, glucose, urea or gonadotropin (FSH and LH) levels between the groups. The mean concentrations of cholesterol were higher in the 1.7 × M group. The total number of small (<4 mm) and medium follicles (4-8 mm) was not altered by the treatments, but the number of small follicles increased on days 1 and 2 of the estrous cycle, with higher values found in the 1.7 × M group. The average of the oocytes also increased (9.50±2.1 and 12.5±4.4 for the 1.0 × M and 1.7 × M groups, respectively). The rapid increase in the amount of energy offered in the diet changes the amount of follicles and oocytes available for follicular aspiration (OPU) in young heifers without changing their metabolic profile.

Dynamics of follicle populations and gonadotropin concentrations in fillies age two to ten months

Follicle populations and concentrations of circulating gonadotropins were studied during age 2-10 months in 10 spring-born pony fillies. Blood sampling and ultrasound scanning were done every 4 days and daily for four 30 day periods. During 5-12 weeks, FSH concentrations were lower in 6 fillies with follicles greater than or equal to 6 mm (mean +/- s.e. 1.4 +/- 0.1 ng/ml) than in 4 fillies with follicles <6 mm (2.8 + 0.3 ng/ml). The diameters and numbers of follicles and gonadotropin concentrations increased progressively during age 2-4 months. A plateau in follicle activity and reduced levels of gonadotropins occurred during 5-7 months. During 8-10 months, follicles grew to >10 mm and gonadotropin concentrations increased. Waves of follicular growth were identified during the 30 day periods by significant increases in the diameter of the 10 largest follicles. The waves did not partition into dominant and subordinate follicles. Results indicated an initial postnatal period of negative ovarian feedback, temporally related changes in gonadotropins and follicles for months 3-10, and development of follicles in waves.

GROWTH OF OOCYTE AND NURSE CHAMBER DURING OOGENESIS IN QUEENS OF THE LEAF-CUTTING ANT ATTA SEXDENS RUBROPILOSA FOREL, 1908 (HYM, FORMICIDAE)

Volume changes of the vitellarium components of the leaf-cutting ant Atta sexdens rubropilosa are reported. The oocyte grew to approximately 409 times of its initial volume and reached a maximum value of 1.2 x 10(7) mu m(3). The follicle increase in size at a more or less constant rate up to the 12(th), showing an elevated growth rate thereafter. The mean number of follicles per ovariole was 16.42 +/- 3.58.

Reduction in size of the ovulatory follicle reduces subsequent luteal size and pregnancy rate

We hypothesized that reducing the size of the ovulatory follicle using aspiration and GnRH would reduce the size of the resulting CL, reduce circulating progesterone concentrations, and alter conception rates. Lactating dairy cows (n=52) had synchronized ovulation and AI by treating with GnRH and PGF(2 alpha) as follows: Day -9, GnRH (100 mug); Day -2, PGF(2 alpha) (25 mg); Day 0, GnRH (100 mug); Day 1, AI. Treated cows (aspirated group; n=29) had all follicles > 4 mm in diameter aspirated on Days -5 or -6 in order to start a new follicular wave. Control cows (nonaspirated group; n=23) had no follicle aspiration. The size of follicles and CL were monitored by ultrasonography. The synchronized ovulation rate (ovulation rate to second GnRH injection; 42/52=80.8%) and double ovulation rate of synchronized cows (6/42=14.3%) did not differ (P > 0.05) between groups. Aspiration reduced the size of the ovulatory follicle (P < 0.0001; 11.5 +/- 0.2 vs 14.5 +/- 0.4 mm), and serum estradiol concentrations at second GnRH treatment (P < 0.0002; 2.5 +/- 0.4 vs 5.7 +/- 0.6 pg/mL). The volume of CL was less (P < 0.05) for aspirated than nonaspirated cows on Day 7 (2,862 +/- 228 vs 5,363 +/- 342 mm(3)) or Day 14 (4,652 +/- 283 vs 6,526 +/- 373 mm(3)). Similarly, serum progesterone concentrations were less on Day 7 (P < 0.05) and Day 14 (P < 0.10) for aspirated cows. Pregnancy rate per AI for synchronized cows was lower (P < 0.05) for aspirated (3/21=14.3%) than nonaspirated (10/21=47.6%) cows. In conclusion, ovulation of smaller follicles produced lowered fertility possibly because development of smaller CL decreased circulating progesterone concentrations. (C) 2001 by Elsevier B.V.

Ovarian follicular dynamics in Nelore breed (Bos indicus) cattle

The most common beef cattle raised in Brazil is the Nelore breed (Bos indicus). Information obtained by ultrasonography on follicular growth in Bos taurus cattle has been accumulating rapidly. However, there are few publications to date on follicular development in Bos indicus breeds. The follicular dynamics in Nelore heifers and cows during natural or prostaglandin (PG)-induced estrous cycle were studied. From the detection of estrus onward, all animals were examined daily by ultrasonography for one (n=35) or two (n=10) consecutive estrous cycles. The follicular dynamic in Nelore cattle was characterized by the predominance of 2 follicular waves in the cows (83.3%, n=18, P<0.05) and 3 waves in the heifers (64.7%, n=16, P<0.05). Most of the cattle observed over 2 consecutive estrous cycles presented the same pattern of follicular waves in the first and second cycle, and only 30% showed variation in the number of waves from one cycle to the other. Most of the follicular parameters analyzed were not affected by PG treatment or age but were altered by follicular waves. Consequently, data on cows and heifers were combined according to the number of follicular waves. The ovulatory follicle was larger than the other dominant follicles (P<0.05), and the ovulatory wave was shorter than the preceding waves (P<0.05). The interovulatory interval was longer in animals showing 3 waves than those exhibiting 2 waves (P<0.05). Maximum diameter of the dominant follicle (around 11 mm) and of the corpus luteum (CL, approximately 17 mm) were smatter than those reported for European breeds. In conclusion, the results demonstrate that although the dominant follicle and corpus luteum are smaller than in European breeds, the follicular dynamics in Nelore cattle were similar to those observed in European breeds and were characterized by 2 or 3 follicular waves for cows and heifers, respectively, during the natural or prostaglandin-induced estrous cycle. (C) 1997 by Elsevier B.V.

Synchronization rate, size of the ovulatory follicle, and pregnancy rate after synchronization of ovulation beginning on different days of the estrous cycle in lactating dairy cows

Recently a protocol was developed that precisely synchronizes the time of ovulation in lactating dairy cows (Ovsynch; GnRH-7d-PGF(2 alpha)-2d-GnRH). We evaluated whether initiation of Ovsynch on different days of the estrous cycle altered the effectiveness of this protocol. The percentage of cows (n=156) ovulating to the first GnRH was 64% and varied (P<0.01) by stage of estrous cycle. Treatment with PGF(2 alpha) was effective, with 93% of cows having low progesterone at second GnRH. The overall percentage of cows that ovulated after second GnRH (synchronization rate) was 87% and varied by response to first GnRH (92% if ovulation to first GnRH vs 79% if no ovulation; P<0.05). There were 6% of cows that ovulated before the second injection of GnRH and 7% with no detectable ovulation by 48 h after second GnRH. Maximal diameter of the ovulatory follicle varied by stage of estrous cycle, with cows in which Ovsynch was initiated at midcycle having the smallest follicles. In addition, milk production and serum progesterone concentration on the day of PGF(2 alpha) affected (P<0.05) size of the ovulatory follicle. Using these results we analyzed pregnancy rate at Days 28 and 98 after Al for cows (n=404) in which Ovsynch was initiated on known days of the estrous cycle. Pregnancy rate was lower for cows expected to ovulate larger follicles than those expected to ovulate smaller follicles (P<0.05; 32 vs 42%). Thus, although overall synchronization rate with Ovsynch was above, 85%, there were clear differences in response according to day of protocol initiation. Cows in which Ovsynch was initiated near midcycle had smaller ovulatory follicles and greater pregnancy rates. (C) 1999 by Elsevier B.V.

Ovulation synchronization following commercial application of ultrasound-guided follicle ablation during the estrous cycle in mares

A regimen of progesterone plus estradiol (P&E) was used as a standard for ovarian synchronization to test the efficacy and evaluate the commercial application of ultrasound-guided follicle ablation as a non-steroidal alternative for ovulation synchronization in mares. Recipient mares at a private embryo transfer facility were at unknown stages of the estrous cycle at the start of the experiment on Day 1 when they were randomly assigned to an ablation group (n = 18-21 mares) or to a ME group (n = 20-21 mares). In the ablation group, mares were lightly sedated and all follicles > 10 mm were removed by transvaginal ultrasound-guided follicle aspiration. In the ME group, a combination of progesterone (150 mg) plus estradiol (10 mg) prepared in safflower oil was given daily (im) for 10 d. Two doses of prostaglandin FZ, (PGF, 10 mg/dose, im) were given 12 h apart on Day 5 in the ablation group, or a single dose on Day 10 in the ME group. Human chorionic gonadotropin (hCG, 2500 IU/mare, im) was given at a fixed time, 6 and 10 d after PGF treatment in the ablation and ME groups, respectively, with the expectation of a follicle > 30 mm at the time of treatment. In both the ablation and P&E groups, transrectal ultrasonography was done at the start of the study (Day 1) and again on the day of hCG treatment and daily thereafter to determine the presence of a CL, measure diameter of the largest follicle and detect ovulation. The mean interval from the start of the study and from PGF treatment to ovulation was shorter (P < 0.0001) in the ablation group (13.7 and 9.7 d, respectively) compared to the P&E group (22.3 and 13.2 d, respectively). Following fixed-day treatment with hCG after PGF treatment, the degree of ovulation synchronization was not different (P > 0.05) between the ablation and P&E groups within a 2-d (56 and 70%) or 4-d (83% and 90%) period. Although ultrasound-guided follicle ablation may not be practical in all circumstances, it excluded the conventional 10-d regimen of progesterone and estradiol and was considered an efficacious and feasible, non-steroidal alternative for ovulation synchronization in mares during the estrous cycle. (C 2007 Elsevier B.V. All rights reserved.

Morphological characterization of follicle deviation in Nelore (Bos indicus) heifers and cows

Follicle diameter deviation is defined as the beginning of the differential change in growth rates between the largest and next largest follicles subsequent to wave emergence and is considered a key component of follicle selection. Follicle selection has been extensively studied in European breeds of cattle (Bos taurus) but has not been critically studied in Zebu breeds (Bos indicus). The objectives of the present study were to determine and compare the morphological characteristics of deviation associated with the first post-ovulatory wave (Wave 1) of the estrous cycle in Nelore heifers (n = 8) and nonlactating cows (n = 11). Beginning on the day of ovulation (day 0), the three largest follicles (F1-F3, respectively) were individually tracked every 12 h for 6 d using transrectal ultrasonography. In individual animals, deviation was determined graphically using visual inspection of the diameter profiles of F1, F2 and sometimes F3 (observed deviation) and mathematically using segmented regression analysis of the diameter differences between F1 and F2 or sometimes F3 (calculated deviation). Mean day of emergence of Wave 1 when F1 reached >3 rum (approximately 1 d after ovulation) and growth rate of F1 during deviation (approximately 1.4 mm/d) were not significantly different between heifers and cows. The results of determining the beginning of deviation within heifers and cows using the observed and calculated methods were not significantly different. Averaged over both methods, diameter deviation occurred 2.8 d after ovulation when F1 reached 5.7 mm in heifers, and 2.4 d after ovulation when F1 reached 6.1 mm in cows. In conclusion, the emergence of Wave 1 and growth rates and diameters of the future dominant follicles at the beginning of deviation were similar in Nelore heifers and nonlactating cows, regardless of the methods used to determine deviation. Relative to Holstein cattle, emergence of Wave 1 appeared to occur about 1 d later and diameter of the future dominant follicle at the beginning of deviation was about 2 turn smaller in Nelore. (C) 2004 Elsevier B.V. All rights reserved.

Differential response of the luteal phase and fertility in cattle following ovulation of the first-wave follicle with human chorionic gonadotropin or an agonist of gonadotropin-releasing hormone

A series of experiments with Holstein heifers was conducted to develop the capability of inducing accessory corpus luteum (CL) with a GnRH agonist (Buserelin, 8 mu g; GnRHa) or hCG; (3,000 IU) to increase plasma progesterone concentrations (Exp. 1, 2, and 3) and to test whether induction of accessory CL with hCG will increase conception rates in heifers (Exp. 4) and lactating cows (Exp. 5). In Exp. 1, heifers were treated on d 5 after estrus with GnRHa (n = 8) or saline (n = 7); heifers in Exp. 2 received hCG (n = 5) or saline (n = 4) on d 5. Experiment 3 allowed a contemporary evaluation of heifers treated on d 5 with GnRHa (n = 6), hCG (n = 6), saline (n = 6), or GnRHa at d 5 and hCG at the time of the induced ovulation (n = 5). The GnRHa and hCG were equally effective in inducing an accessory CL (93% induction rate), but the subsequent increase in progesterone concentrations was greater in hCG-treated heifers. A greater half life of hCG may provide longer LH-like stimulation of the first-wave follicle and subsequent developing accessory CL or a greater luteotropic effect on the original CL. Induction of an accessory CL with hCG on d 5 or 6 after insemination did not increase pregnancy rates in fertile heifers (Exp. 4: hCG = 64.8% vs control = 62.9%; n = 243) or lactating dairy cows during summer heat stress (Exp. 5: hCG = 24.2% vs control = 23.5%; n = 201).

Histochemical and ultrastructural cytochemistry of glycogen on ovarioles of Neoponera villosa ants (Hymenoptera: Ponerinae)

In Neoponera villosa ants, we found ovaries of the polytrophic meroistic type which is characterized by the presence of nurse cells forming together with the oocyte, the so-called follicles. The nurse cells have the primary function of supplying the oocyte with RNA, but they contribute to the supply of other elements such as glycogen. With the objetive of detecting the presence of this substance in the ovarioles of workers and queens of N.villosa ante the ovaries were removed and processed according to electron microscopy technic for glycogen detection. Glycogen is a common element in insect oocytes and is abundantly distributed in the cytoplasm of N.villosa workers and queens. However, in ovarian follicles it can only be detected at stages ET and lit of development. Glycogen synthesis probably occurs predominantly in nurse cells which transfer it into the oocyte through the nourish pore. This process requires high energy expenditure that justify the large numbers of mitochondria associated with glycogen in the nurse cell cytoplasm. The amount of glycogen in the nurse cells of queens is slightly greater than workers.

Expression of LH receptor mRNA splice variants in bovine granulosa cells: Changes with follicle size and regulation by FSH in vitro

In cattle, most evidence suggests that granulosa cells express LH receptors (LHR) after (or as) the follicle becomes dominant, however there is some suggestion that granulosa cells from smaller pre-dominant follicles may express several LHR mRNA splice variants. The objective of this study was to measure LHR expression in bovine follicles of defined size and steroiclogenic ability, and in granulosa cells from small follicles (< 6 mm diameter) undergoing differentiation in vitro. Serniquantitative RT-PCR demonstrated that LHR mRNA was undetectable in granulosa cells of follicles < 7 mm diameter (nondominant follicles), and increased with follicle diameter in follicles > 7 mm diameter. Splice variants with deletions of exon 10 and part of exon 11 were detected as previously described, and we detected a novel splice variant with a deletion of exon 3. Cultured granulosa cells contained LHR mRNA, but with significantly greater amounts of variants with deletions of exon 10 and/or exon 11 compared with cells from dominant follicles. FSH increased the abundance of some but not all LHR mRNA splice variants in cultured granulosa cells. The addition of LH to cultured cells did not increase progesterone secretion, despite the presence of LHR mRNA. Collectively, these data suggest that granulosa cells do not acquire functional LHR until follicle dominance occurs.

Effects of dominant follicle aspiration and treatment with recombinant bovine somatotropin (BST) on ovarian follicular development in Nelore (Bos indicus) heifers

Follicle ablation has been recognized as an efficient method of follicular wave synchronization. Treatment with recombinant bovine somatotropin (BST) has been shown to enhance follicular development in <(Bos taurus)under bar>. This experiment assessed the effects of these treatments in Nelore (<(B. indicus)under bar>) heifers. Eight cycling Nelore heifers were randomly assigned to 3 different treatments. on Day 2 of a synchronized cycle (Day 0 = day of ovulation), heifers assigned to Treatments 1 and 2 received 2 mL of saline, whereas heifers assigned to Treatment 3 received 320 mg of BST. on Day 5, the first-wave dominant follicle was ablated by ultrasoundguided transvaginal aspiration in heifers in Treatments 2 and 3, and all heifers received an injection of prostaglandin on Day 11. Aspiration of the dominant follicle advanced and synchronized (P < 0.05) the day of second-wave emergence (6.9 +/- 0.1 vs. 8.4 +/- 0.4) and the day of the pre-wave FSH peak (6.0 +/- 0.0 vs. 6.9 +/- 0.4), and increased FSH peak concentrations (381 +/- 21 vs. 292 +/- 30; pg/mL; P < 0.01). Recombinant bovine somatotropin treatment caused a two-fold increase in plasma insulin-like growth factor-I (IGF-I) concentrations (P < 0.001) and resulted in a 36% increase in the number of small follicles (<5 mm; P < 0.001) compared with saline-treated heifers. In summary, in agreement with previous reports on <(B. taurus)under bar>, dominant follicle aspiration synchronized ovarian follicular development, and BST treatment increased peripheral concentrations of IGF-I in Nelore heifers. Recombinant bovine somatotropin also increased the number of small follicles, but this response appeared to be inferior to that reported for <(B. taurus)under bar>. (C) 2000 by Elsevier B.V.

Ovarian follicle diameter at timed insemination and estrous response influence likelihood of ovulation and pregnancy after estrous synchronization with progesterone or progestin-based protocols in suckled Bos indicus cows

The objectives of the present study were to evaluate factors associated with estrous synchronization responses and pregnancy per insemination (P/AI) in Bos indicus beef cows submitted to progesterone-based fixed-time artificial insemination (FTAI) protocols. A total of 2388 cows (1869 Nellore and 519 crossbred Nellore x Angus) from 10 commercial farms were evaluated to determine the relationships among breed, body condition score (BCS) on the first day of the FTAI protocol, the occurrence of estrus between progesterone device removal and FTAI and diameter of largest ovarian follicle (LF) at FTAI on estrous synchronization responses and P/AI. Cows (n=412 primiparous; 1976 multiparous) received an intravaginal device containing progesterone or an ear implant containing norgestomet (a progestin), and an injection of estradiol at the beginning of the estrous synchronization protocol. Body condition was scored using a 1-5 scale on the first day of the FTAI protocol and at 30-60 days postpartum. Females received 300IU of equine chorionic gonadotropin (eCG) and PGF(2 alpha) on the day the progesterone device/implant was removed and were inseminated 48-60h later. At insemination, cows (n=2388) were submitted to an ultrasonographic exam to determine the diameter of the LF. Follicles were classified into four categories based on mean and standard deviation (SD) of the LF (LF1 = two SD below the mean; LF2 = mean minus one SD; LF3 = mean plus one SD; LF4 = two SD above the mean). Ovulation rate was determined in a subset of cows (n=813) by three consecutive ultrasonographic exams: (1) at time of progesterone device/implant removal, (2) at time of FTAI and (3) 48 h after FTAI. Ovulation was defined as the disappearance of a large follicle (>= 8.0 mm) that was previously recorded. Estrus was determined in a subset of the cows (n = 445) by the activation of a detection of estrous patch placed on the tail head on the day of progesterone device/implant removal. Pregnancy was diagnosed 30 days after FTAI. Pregnancy was influenced (P = 0.001) by follicle diameter [LF1 = 27.5% (81/295), LF2 = 46.6% (328/705), LF3 = 57.9% (647/1118), LF4 = 63.3% (171/270)] and the occurrence of estrus [estrus = 67.7% (174/257) and no estrus = 36.2% (68/188)]. Follicle diameter at FTAI influenced ovulation rate [LF1 = 42.5% (34/80), LF2 = 73.9% (161/218), LF3 = 95.8% (407/425), LF4 = 97.8% (88/90)], the occurrence of estrus [LF1 = 54.8% (51/93), LF2 = 33.6% (43/128), LF3 = 68.9% (126/183), LF4 = 90.2% (37/41)] and P/AI among cows that had ovulations [LF1 =32.4% (11/34), LF2 = 50.3% (81/161), LF3 = 60.0% (244/407), LF4 = 68.2% (60/88)]. Improving estrous responses between progesterone device withdrawal and FTAI and increasing the diameter of the LF at FTAI may be important aspects to achieve improved estrous synchronization responses and P/AI following progesterone/progestin and estradiol based FTAI protocols in suckled Bos indicus cows. (C) 2010 Elsevier B.V. All rights reserved.

Ultrastructure of the ovariole sheath in Diatraea saccharalis (Lepidoptera : Pyralidae)

The ultrastructure of the ovariole sheath along the Diatraea saccharalis ovariole was studied by scanning and transmission electron microscopy. Each ovariole is surrounded by an epithelial sheath, a tunica propria and scattered lumen cells. These three components of the ovariole sheath show different ultrastructural features along the ovariole, in the germarium or in the vitellarium; these differences are more evident in the epithelial sheath cells. The epithelial sheath is composed by two layers of cells, the external one running longitudinally and the internal one running circularly in the ovariole. These cells, in vitellarium, present cytoplasmic bundles of myofilaments that are arranged parallel to the long axis of the cells; these myofilaments are apparently related to the contraction movements of the follicles within the ovariole. The acellular tunica propria, composed of finely filamentous material, is attached to the adjacent follicle cells by adhesive dense plates. Between the epithelial sheath and the tunica propria there is a population of lumen cells, with morphological features of secretory activity.

LH and FSH concentration and follicular development in Nellore heifers submitted to fixed-time artificial insemination protocols with different progesterone concentrations

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