In vitro Development of Buffalo Oocytes in Media-containing Fluids from Different Size Class Follicles

Studies were conducted to investigate the effect of supplementation of fluid from different sized class [small (SFF, < 3 mm), medium (MFF, 3-8 mm) and large (LFF, > 8 mm)] of normal and cystic (CFF) ovarian follicles in oocyte culture media on oocyte maturation rate and embryo development in vitro and to test the efficacy of follicular fluid (FF) from different size classes as a whole oocyte maturation medium. Results suggested that FF were capable of developing buffalo oocytes to embryonic stage in vitro although its efficacy was lower than that of serum. Regardless of high maturation rates after in vitro maturation (IVM) in media containing FF or IVM in whole FF, low blastocyst rates were obtained after in vitro fertilization (IVF) and culture of embryos. Follicular fluid from small follicles had significantly (p < 0.05) higher potential of developing buffalo oocytes to embryonic stage in vitro than that from medium and large follicles. Cystic FF was not capable of supporting development of buffalo oocytes in vitro.     

Follicular Fluid Prolactin and the Periovulatory Prolactin Surge in the Mare

Prolactin may play multiple roles in equine reproduction. Prolactin appears to be associated with seasonal reproduction, and fluctuating prolactin levels during the estrous cycle suggest that it may play a role in estrous cyclicity as well. The purpose of this research was to investigate the activity of prolactin during the follicular phase of the estrous cycle. In experiment 1, prolactin concentrations were determined from plasma samples collected at least every other day throughout the estrous cycle. Periovulatory (ovulation ± 1 day) prolactin concentrations were compared with concentrations during early diestrus (days 2−10 postovulation). In experiment 2, prolactin concentrations were measured in follicular fluid collected from 74 follicles of various sizes. Follicles were grouped into small (≤20 mm), medium (21−35 mm), and large (>35 mm) size categories. Prolactin concentrations increased during the periovulatory period in cycling mares. This periovulatory surge was superimposed on baseline prolactin concentrations that varied with season. Prolactin was present in significant quantities in the follicular fluid. Follicular fluid prolactin concentrations were lowest in small follicles and increased in medium and large follicles. Concentrations did not differ between medium and large follicles. Follicular fluid prolactin concentrations were lower in autumnal follicles compared with summer follicles of comparable size. It is possible that the short-term surge in circulating prolactin around ovulation could be linked to the significant levels of prolactin in follicular fluid. Ovulation releases a relatively large volume of fluid into the peritoneum. The prolactin in this fluid could be a contributor to the periovulatory prolactin surge.     

Comparison of leptin levels in serum and follicular fluid during the oestrous cycle in cows

Leptin is mainly synthesised in white adipose tissue. Besides its effects on body weight and metabolic homeostasis, leptin also has effects on puberty, sexual maturation and reproduction. In this study the relationship between leptin, IGF-1, oestradiol (E2) and progesterone levels were investigated in serum and follicular fluid from cows. This study included 72 healthy, Brown Swiss cows aged 4-5 years. Samples from the jugular vein and follicular fluids were collected. Phases of the oestrus cycle of cows were classified according to their serum progesterone levels (< 3.18 nmol/l, follicular phase and the others as luteal phase). Follicles were grouped as large (> or = 8 mm) or small (< 8 mm). Leptin, IGF-1, oestradiol and progesterone levels were measured from serum and follicular fluid. Leptin concentrations were found to be significantly higher in luteal-phase follicular fluid of small follicles (P < 0.05). These were classified as atretic follicles. There was a positive correlation between serum and follicular fluid leptin levels in the luteal phase. Serum leptin was found to have a positive correlation with follicular fluid progesterone level (P = 0.01) in the preovulatory follicles. The present study shows that there is a relationship between the concentration of leptin in follicular fluid and atresia in small follicles.     

Dynamics of Follicular Fluid in One‐humped Camel (Camelus dromedarius)

In the present study, ovarian follicular fluid and serum biochemical, hormonal, electrolytes and amino acids profiles in female dromedary camel (Camelus dromedarius), were investigated. Fluid from small (2–6 mm) and large follicles (7–20 mm) and blood samples were collected from 25 clinically healthy adult female camels. The concentrations of glucose, cholesterol, triglycerides, high‐density lipoproteins, urea, total proteins, albumin, globulin, fibrinogen, alanine aminotransferase, aspartate aminotransferase and tri‐iodothyronine were lower (p ≤ 0.05) in large follicles when compared with the small follicles. However, the concentrations of low‐density lipoproteins, uric acid, creatinine, alkaline phosphatase and acid phosphatase in small and large follicles did not differ. The concentrations of oestradiol 17‐β and progesterone were higher (p ≤ 0.05) in large follicles. The serum concentrations of these hormones were many folds lower (p ≤ 0.05) than those of follicular fluid. Among electrolytes, the concentration of phosphorus was higher (p ≤ 0.05) in the large follicles, while that of potassium and chloride were lower (p ≤ 0.05) in the small follicles. Serum concentrations of sodium, chloride, calcium and phosphorous were higher (p ≤ 0.05), while that of potassium lower (p ≤ 0.05) than corresponding concentrations in the follicular fluid. The concentrations of leucine and arginine were higher (p ≤ 0.05) in follicular fluid when compared with serum concentrations, while the reverse was true for other amino acids. In conclusion, this study is indicative of either low or high concentrations of certain biochemical metabolites, hormones, electrolytes and amino acids in small and large follicles for the individual roles that they play in the growth and development of follicles in the one‐humped she‐camel.     

Changes in follicular fluid steroids, insulin-like growth factors (IGF) and IGF-binding protein concentration, and proteolytic activity during equine follicular development

The objective of the present study was to evaluate changes in equine follicular fluid insulin-like growth factor binding protein (IGFBP) proteolytic activity as well as steroid, IGF, and IGFBP concentrations during follicular development in the mare. Mares (n = 14) were classified as either in the follicular phase (n = 8) or luteal phase (n = 6). Follicles (n = 92) were categorized as small (6 to 15 mm; n = 54), medium (16 to 25 mm; n = 23), or large (> 25 mm; n = 15), and follicular fluid was collected. Estradiol and androstenedione levels in follicular fluid were greater (P < 0.05), and IGFBP-3 concentrations tended to be greater (P < 0.10) in large than in small or medium follicles, whereas IGFBP-2, -4, and -5 levels were less (P < 0.05) in large than in small or medium follicles. Estradiol and androstenedione concentrations were negatively correlated (P < 0.01) with IGFBP-2, -4, and -5 but not IGFBP-3 concentrations. To evaluate proteolysis of IGFBP, follicular fluid was incubated with human 125I-labeled IGFBP-2, -3, and -5 and protein separated by 12% SDS-PAGE. Follicular fluid caused little or no proteolysis of 125I-lableled IGFBP-2 or -3, and the small amount of proteolysis of IGFBP-2 and -3 did not differ (P > 0.10) among follicle classes. However, more 125I-labeled IGFBP-5 was cleaved (P < 0.05) by follicular fluid from large follicles collected during the follicular phase than large follicles during the luteal phase, and small or medium follicles from follicular and luteal phase mares indicating that a protease to IGFBP-5 exists in estrogen-dominant equine follicles. This IGFBP-5 protease was inhibited by kallikrein/serine protease and metalloprotease inhibitors. We conclude that the tendency of estrogen-dominant follicles of mares to have greater levels of IGFBP-3 and lesser levels of IGFBP-2 does not appear to be due to differences in proteolysis, whereas changes in IGFBP-5 levels are likely due to changes in activity of a serine protease or metalloprotease. Changes in IGFBP may alter levels of bioavailable IGF that stimulate steroidogenesis and mitogenesis in developing mare follicles.     

Intrafollicular Concentrations of Steroid Hormones and PGF2α in Relation to Follicular Development in the Mares during the Breeding Season

The concentrations of androstenedione, estradiol-17β, progesterone and PGF_2α contained in the follicular fluid produced by the follicles in collected ovaries of mares that have had estrous phase during the breeding season were measured and analyzed the relation between the growth stage of follicles and the hormone levels in the follicular fluid. An ultrasonographic diagnostic instrument was used to measure the diameter of the follicles in order to categorize the follicles into three groups the following: 8 small follicles (from 1.0 to less than1.5 cm), 8 medium follicles (from 1.5 to less than 3.0 cm), and 8 large follicles (from 3.0 to 5.0 cm), respectively. The analysis of the follicular fluid in ovaries of estrous mares showed that the concentrations of androstenedione were significantly higher in the medium or large follicles than in the small follicles and the concentrations of estradiol-17β were significantly higher in larger follicles than in the small or medium follicles (P<0.05). The concentrations of progesterone and PGF_2α, on the other hand, did not significantly vary regardless of follicluar size. In the follicles within the mare ovaries that have had estrous stage, the concentrations of the hormones related the ovulation, namely androstenedione and estradiol-17β, were higher with larger follicles.     

Low Molecular Mass Factors from Follicular Fluid Inhibit Steroidogenesis in Bovine Granulosa Cells

Gonadotropins are required for follicular growth and differentiation, but increasing amounts of evidence indicate that intrafollicular factors modulate their effects at granulosa cell level. In order to study the effect of factors present in bovine follicular fluid, a partial purification of low molecular mass factors from fluids collected from small (< 5 mm), medium (5–8 mm) and large (> 8 mm) follicles was performed and the biological activity of these peptides on steroidogenesis of granulosa cells from small and large follicles was examined. The purification was carried out by filtration through membranes in 25 and 10 kDa molecular weight cutoffs. After filtration, samples were analysed by polyacrylamide gel electrophoresis and protein concentration was measured by spectrophotometric analysis. Granulosa cells from small and large follicles were cultured in serum‐free DMEM/Ham's F12 (1 : 1) plus transferrin (5 mg/l) and selenium (5 µg/l) for 2 days. At the end of the culture period, media were renewed and follicular extracts from two different preparations (< 25 and < 10 kDa) were added at the concentrations of 1–10–100–1000 ng/ml. After 24 h the media were collected and stored until estradiol 17β (E2) and progesterone (P4) determination by validated radio‐immuno‐assays. Basal P4 production was 18.3 ± 1.4 (mean ± SEM)and 9.8 ± 1.8 ng/24 h per 3 × 10⁴ cells from small and large follicles, respectively. Both < 10 and < 25 kDa extracts reduced P4 production by cells from both the types of follicles (p < 0.05). Basal E2 release was 671.8 ± 21.4 and 5500 ± 800 pg/24 h per 3 × 10⁴ cells from small and large follicles, respectively. Both extracts reduced E2 production by either cells from small and large follicles (p < 0.05). No differences were observed in the inhibition of steroidogenesis by purifications obtained from large, medium or small follicles. Results of this study indicate that factors present in bovine follicular fluid can reduce steroidogenesis in granulosa cells in vitro.     

Effect of porcine somatotropin on number of granulosa cell luteinizing hormone/human chorionic gonadotropin receptors, oocyte viability, and concentrations of steroids and insulin-like growth factors I and II in follicular fluid of lean and obese gilts.

Prepubertal gilts of obese (n = 24) or lean (n = 24) genetic lines were injected (s.c.) daily with 0, 2, or 4 mg of porcine somatotropin (pST) for 6 wk starting at 160 d of age to determine whether pST affects follicular function. Blood and ovaries were collected at slaughter 24 h after the last injection. Surface follicles greater than or equal to 1.0 mm in diameter were counted, and pools of follicular fluid (FFL) and granulosa cells were collected from 1.0- to 3.9-mm (small) and 4.0- to 6.9-mm (medium) follicles. Oocytes were collected from small and medium follicles and evaluated for maturational stage and viability. Porcine somatotropin increased (P less than .08) the numbers of small but not the numbers of medium follicles per gilt (P greater than .10). Oocyte maturation and viability were not affected by pST or genetic line. Porcine somatotropin increased (P less than .05) concentrations of insulin-like growth factor I (IGF-I) in serum and FFL of both obese and lean gilts; IGF-I was lower (P less than .01) in lean gilts. Treatment with pST decreased (P less than .05) IGF-II in FFL of lean but not in that of obese gilts. Dose of pST and line had no effect on concentrations of progesterone in FFL of small or medium follicles or on concentrations of estradiol in FFL of small follicles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Impact of dietary lysine intake during lactation on follicular development and oocyte maturation after weaning in primiparous sows

Primiparous sows (n = 36) were used to evaluate the effects of dietary lysine intake in lactation on follicular development and oocyte maturation after weaning. Sows were assigned randomly to one of three diets containing .4% (low lysine, LL), 1.0% (medium lysine, ML), or 1.6% (high lysine, HL) total lysine. All diets contained 2.1 Mcal NE/kg and exceeded NRC (1988) requirements for all other nutrients. Actual lysine intakes over an 18-d lactation were 16, 36, and 56 g/d for sows consuming LL, ML, and HL, respectively. Ovarian data were analyzed for sows determined to have been slaughtered during the first proestrus period after weaning, using previously established criteria. Compared with sows fed ML and HL, sows fed LL tended to have lower uterine weight, follicular fluid volume, and follicular fluid (FF) estradiol (E2) content (P < .15), but similar ovarian weight and follicular fluid IGF-I concentration. Within the largest 15 preovulatory follicles, sows fed LL had a lower percentage of large (> or = 7.0 mm) follicles (33 vs 50 and 58%; P < .01) and a higher percentage of medium (5.5 to 7.0 mm) follicles (62 vs 44 and 39%; P < .01) but a similar percentage of small (< or = 5.5 mm) follicles (4.4 vs 5.9 and 3.7%; P > .15), respectively, compared with sows fed ML or HL. Standardized pools of oocytes aspirated from follicles of prepubertal gilts were incubated for 44 h with pooled FF recovered from the largest 15 follicles of each experimental sow. Fewer oocyte nuclei matured to metaphase II of meiosis when cultured with FF recovered from sows fed LL, than from sows fed ML or HL (47.1 vs 59.8 and 63.8%, respectively; P < .01). Our results suggest that low lysine (protein) intake in primiparous lactating sows impaired follicular development and reduced the ability of follicles to support oocyte maturation. However, high compared with medium lysine (protein) intake had no further positive effects on ovarian function.     

Histological Characteristics and Steroid Concentration of Ovarian Follicles at Different Stages of Development in Pregnant and Non-pregnant Dairy Cows

The aim of the study was to investigate the histological characteristics and steroid concentrations in follicular fluid of different populations of follicles at different stages of development, during pregnancy and the oestrous cycle in cows. Follicles from ovaries collected at a slaughterhouse were allocated into three size categories (small, 2–5.9 mm; medium, 6–13.9 mm; and large, 14–20 mm) in pregnant and non-pregnant cows. Slices were stained with HE and PAS for histological analysis. Follicular fluid was pooled according to size and pregnancy status and estradiol, testosterone and progesterone concentrations in follicular fluid were determined by RIA. Characteristics of healthy follicles did not differ, regardless of follicle size or pregnancy status. Total histological atresia was significantly higher in pregnant cows than in non-pregnant cows (p < 0.05). Estradiol increased and testosterone decreased significantly, while follicles increased in size, in both non-pregnant and pregnant cows (p < 0.05). Nonpregnant cows had the highest estradiol values in follicles of all sizes. Medium and large follicles from pregnant cows showed the lowest testosterone concentration (p < 0.05). Progesterone levels increased with follicle size only in non-pregnant animals. In large follicles, progesterone concentration was significantly higher in non-pregnant cows than in pregnant cows (p < 0.05). Considering steroid concentration and histological findings, most large follicles might be atretic during pregnancy in cattle.     

Insulin-like growth factor binding proteins in granulosa and thecal cells from bovine ovarian follicles at different stages of development

Because IGFBP inhibit IGF-stimulated cellular proliferation and differentiation, it is hypothesized that variations among IGFBP in individual follicles might contribute to the regulation of recruitment, selection, dominance, and turnover of ovarian follicles. Sources of IGFBP in fluid of bovine follicles are not well established; thus, objectives of this study were to determine levels of IGFBP binding activities and messenger RNA (mRNA) in granulosa and theca interna cells at different stages of follicular development (small [< 6 mm], medium [6 to < 8 mm], and large [> or = 8 mm]) and to characterize associations of these levels measured in the cells with levels of IGFBP and steroids in follicular fluid. Thecal and granulosa cells from large healthy follicles contained two- to twentyfold less (P < 0.05) IGFBP-2, -3, and -5 than cells from small, medium, and large atretic follicles. Thecal cells from small, medium, and large atretic follicles contained more (P < 0.05) IGFBP-3 and -4 than granulosa cells from these follicles, whereas granulosa cells from these follicles contained more IGFBP-2 activity than thecal cells. Differences in IGF binding activity were paralleled by differences in levels of mRNA for the respective IGFBP. Developmental differences in IGFBP activity in follicular fluid were positively associated with activity in granulosa and/or thecal cells, with the exception of IGFBP-4, which was low in fluid from large healthy follicles but markedly increased (mRNA and binding activity) in granulosa cells from these follicles. It is concluded that developmental changes in follicular fluid IGFBP-2 and -5 binding activities seem to be controlled in part by alterations in synthesis of these IGFBP by granulosa and thecal cells, whereas diminished IGFBP-4 in fluid from large healthy follicles occurs concomitantly with increased levels of IGFBP-4 mRNA and activity in granulosa cells, implicating posttranslational regulation by specific proteases.     

Bovine oocytes in secondary follicles grow in medium containing bovine plasma after vitrification

There has been no culture system that supports the growth of bovine oocytes for more than 2 weeks. In the present study, bovine secondary follicles were cultured for 4 weeks, and the effects of supplemented protein components and FSH in the culture medium on the growth of the oocytes were examined. The effect of vitrification of secondary follicles on the subsequent oocyte growth was also examined. Secondary follicles (150 to 200 μm in diameter) containing growing oocytes (approximately 60 μm in diameter) were dissected from ovaries and cultured in a medium supplemented with FSH (0, 25 or 50 ng/ml) and one of the following four kinds of protein components: bovine serum albumin (BSA), bovine plasma (BPL), fetal calf serum (FCS) and bovine follicular fluid (BFF). In BSA- and BPL-supplemented media with 0 or 25 ng/ml FSH, more than 50% of follicles showed no degenerative signs during culture, and oocytes significantly increased in size after 4 weeks (P<0.05). Higher percentages of granulosa cell-enclosed oocytes were recovered from the follicles cultured in BPL-supplemented media with 0 and 25 ng/ml FSH, and the oocytes grew to 90 μm or more in diameter. In FCS- and BFF-supplemented media, FSH increased the numbers of degenerating follicles. Next, vitrified-warmed secondary follicles were cultured in BPL-supplemented medium. One third of the follicles showed no degenerative signs, and the oocytes increased in diameter to 88.8 ± 3.1 μm after 4 weeks of culture. These results suggest that a BPL-supplemented medium supports oocyte growth in bovine secondary follicles for 4 weeks, even after vitrification and warming of the follicles.     

Follicular fluid stimulates capacitation and acrosome reaction in alpaca sperm (Vicugna pacos)

The follicular fluid exerts an effect on the sperm capacitation of several species; however, these effects vary according to species, both in the sperm motility and in the subsequent acrosome reaction. In this study, the effect of alpaca follicular fluid (aFF) on the motility and acrosome reaction of alpaca spermatozoa was observed, using follicular fluid of three follicle sizes: small (<3 mm), medium (3‐6 mm) and large (>6 mm), in a concentration of 30%. Sperm motility at the first hour of incubation with aFF of small follicles was 48.0%, with aFF of medium follicles it was 43.33% and with aFF of large follicles, it was 34.53%, while control averaged 26.00%. At the second hour, control achieved an average of 28.13%, treatment with aFF from small follicles showed an average of 46.53%, with aFF from medium follicles it was 40.00% and with aFF from large follicles it was 35.60%. The acrosome reaction after 4 hours of incubation was 30.06% for control, whereas for aFF of small follicles it was 66.3%, with aFF of medium follicles it was 58.86% and for aFF of large follicles, it was 67.63%. In the case of sperm motility, a significant difference is demonstrated for all treatments in relation to the control at the first hour, whereas only the treatments with aFF of small and medium follicles show a significant difference with respect to the control at the second hour. In the case of the acrosome reaction, all treatments with follicular fluid show a significant difference with respect to the control. It was concluded that alpaca follicular fluid favours sperm capacitation and the acrosome reaction in alpaca spermatozoa.     

Biochemical analysis of bovine follicular fluid: albumin, total protein, lysosomal enzymes, ions, steroids and ascorbic acid content in relation to follicular size, rank, atresia classification and day of estrous cycle

To investigate some biochemical changes during bovine follicle development, ovaries were obtained from cyclic heifers (7 to 11 heifers/d on each day of the 21-d estrous cycle; N = 152). Follicular fluid from the two largest follicles from both ovaries and a pool from small follicles (N = 30/cow) were collected from each animal and analyzed for ionic, enzymatic and endocrine changes in relation to day of the estrous cycle, follicle size, rank and atretic or growing status. Follicular fluid alkaline phosphatase activity and ascorbate concentrations were highest in all follicular sizes during the earlier portion of the estrous cycle (d 1 to 12; P less than .05), then decreased to the lowest levels (d 13 to 21). As follicular size (diameter) increased lactate dehydrogenase (LDH), acid and alkaline phosphatase activity was reduced in follicular fluid (P less than .05). Alkaline phosphatase and LDH activity tended to be increased in atretic follicles (P less than .10), and was correlated with increased progesterone and androgen concentrations of follicular fluid (r = .4, P less than .05). Both albumin and total protein concentrations decreased as follicular diameter increased (P less than .05). Sodium concentrations in follicular fluid were greater in growing-antral than atretic follicles, and increased with follicular enlargement (P less than .05). Follicular potassium concentrations increased as the estrous cycle progressed (P less than .05), and tended to be elevated in atretic follicles (nonsignificant). Both Ca and Mg concentrations increased with follicular enlargement (P less than .05). Dehydroepiandrosterone and testosterone were the predominant androgens in follicular fluid (androstenedione, the lowest concentration); their concentration decreased with follicle development (P less than .05), but were quite variable. Estradiol was increased in growing follicles (P less than .01). Estrone and estradiol concentrations increased as ovulation approached, particularly in small follicles (less than or equal to 4 mm diameter). Changes of biochemical components found in follicular fluid that relate to the growth and atresia process may provide a more sensitive and accurate method to classify follicle status, and thus aid in understanding the complexity of events associated with maturation of the bovine follicle and oocyte.     

Effects of intraovarian infusion of insulin-like growth factor-I on ovarian follicular function in cattle

The objective of this study was to determine if increased secretion of intraovarian insulin-like growth factor-I (IGF-I), experimentally induced via minipumps, affects follicular function in cattle. Fourteen cycling Holstein cows were divided equally into two groups: Control, osmotic minipumps (containing vehicle) surgically inserted into each ovary, or IGF-I treated, osmotic minipumps as in Controls but pumping 2.0 microg of recombinant human IGF-I per hr for 7 days. All cows were synchronized with prostaglandin F(2alpha) 0.10) between Control and IGF-I-treated cows during Days 2 to 6 of treatment. IGF-I treatment increased (P<0.05) estradiol concentrations in follicular fluid of small follicles, but had no effect (P<0.10) on estradiol concentrations in follicular fluid of large follicles, or on progesterone, androstenedione, or IGF binding protein concentrations in small or large follicles. We conclude that a 7-day infusion of IGF-I directly into the stroma of the ovary altered follicular growth and follicular fluid estradiol concentrations.     

Ovarian follicular growth, function and turnover in cattle: a review

Studies in cattle assessing changes in number and size of antral follicles, concentrations of estradiol, androgens and progesterone in serum and follicular fluid, and numbers of gonadotropin receptors per follicle during repetitive estrous cycles and postpartum anestrus are reviewed. The rate of growth of small follicles (1 to 3 mm) into larger follicles increases as the estrous cycle progresses from d 1 to 18 (d 0 = estrus). Size of the largest antral follicle present on the ovary also increases with advancement of the estrous cycle. Most large follicles (greater than 10 mm) persist on the ovarian surface for 5 d or more between d 3 and 13 of the bovine estrous cycle. After d 13, most of these large follicles are replaced more frequently by new growing follicles (turnover) with an increased probability for recruitment of the ovulatory follicle after d 18. More research is needed to determine the time required for growth of bovine follicles from small to large antral size and evoke recruitment of the ovulatory follicle. Factors that regulate selection of the ovulatory follicle are unknown but may involve increased frequency of LH pulses in blood, altered blood flow and(or) changes in intrafollicular steroids and proteins. Quantitative evaluation of ovarian follicles indicated occurrence of consistent short-term changes in fluid estradiol and numbers of luteinizing hormone receptors in cells of large follicles only during the pre-ovulatory period. Presumably, low concentrations of follicular estradiol found during most of the estrous cycle are not due to a lack of aromatizable precursor or follicle-stimulating hormone receptors. Follicular fluid concentrations of progesterone increase only near the time of ovulation. Little is known about changes in follicular growth, turnover and function during postpartum anestrus in cattle. However, preliminary data suggest that the steroidogenic capacity of large follicles changes markedly during the postpartum period.     

Ovarian follicular development in cattle selected for twin ovulations and births

Comparisons of numbers of antral ovarian follicles and corpora lutea (CL), of blood hormone concentrations, and of follicular fluid steroid concentrations and IGFBP activity were conducted between cows selected (twinner) and unselected (control) for twin births to elucidate genetic differences in the regulation of ovarian follicular development. Ovarian follicular development was synchronized among cows by a single i.m. injection of PGF2alpha on d 18 of the estrous cycle; six cows per population were slaughtered at 0, 24, 48, and 72 h after PGF2alpha. Jugular vein blood was collected from each animal at PGF2alpha injection and at 24-h intervals until slaughter. Ovaries of twinner cows contained more small (< or = 5 mm in diameter, P < 0.05), medium (5.1 to 9.9 mm, P < 0.05), and large (> or = 10.0 mm, P < 0.01) follicles and more (P < 0.01) CL than ovaries of controls. Follicular fluid concentrations of estradiol, androstenedione, testosterone, and progesterone reflected the stage of follicular development and were similar for twinner and control follicles at the same stage. Earlier initiation of follicular development and/or selection of twin-dominant follicles in some twinner cows resulted in greater concentrations of estradiol in plasma at 0, 24, and 48 h and of estradiol, androstenedione, and testosterone in follicular fluid of large follicles at 0 h after PGF2alpha for twinner vs. control cows (follicular status x time x population, P < 0.01). Binding activities of IGFBP-5 and -4 were absent or reduced (P < 0.01) in follicular fluid of developing medium and large estro-gen-active (estradiol:progesterone ratio > 1) follicles but increased with atresia. Only preovulatory Graafian follicles lacked IGFBP-2 binding, suggesting a possible role for IGFBP-2 in selection of the dominant follicle. Concentrations of IGF-I were twofold greater (P < 0.01), but GH (P = 0.10) and cholesterol (P < 0.05) were less in blood of twinners. Three generations of selection of cattle for twin ovulations and births enhanced ovarian follicular development as manifested by increased numbers of follicles within a follicular wave and subsequent selection of twin dominant follicles. Because gonadotropin secretion and ovarian steroidogenesis were similar for control and twinner cattle, enhanced follicular development in twinners may result from decreased inhibition by the dominant follicle(s), increased ovarian sensitivity to gonadotropins, and/or increased intragonadal stimulation, possibly by increased IGF-I.     

Follicular fluid concentrations of free insulin-like growth factor (IGF)-I during follicular development in mares

The objective of the present study was to evaluate changes in concentrations of free insulin-like growth factor (IGF)-I in follicular fluid (FFL) during follicle development in the mare. Mares (n = 14) were classified as either in the follicular phase (n = 8) or luteal phase (n = 6). Follicles (n = 92) were categorized as small (6–15 mm; n = 54), medium (16–25 mm; n = 23) or large (>25 mm; n = 15) and FFL was collected. Free IGF-I levels in FFL in large follicles of follicular phase mares were greater (P < 0.05) than in large follicles of luteal phase mares and small or medium follicles of luteal and follicular phase mares. Free IGF-I concentrations were greater (P < 0.05) in large follicles of luteal phase mares than small but not medium follicles of luteal phase mares. FFL ratio of estradiol:progesterone paralleled changes in free IGF-I. Free IGF-I concentrations were negatively correlated (P < 0.05) with insulin-like growth factor binding protein (IGFBP)-2, -4 and -5 but not IGFBP-3 levels. In addition, free IGF-I concentrations in FFL were positively correlated (P < 0.01) with FFL estradiol, progesterone, androstenedione, estradiol:progesterone ratio, total IGF-I and total IGF-II. We conclude that increases in intrafollicular levels of bioavailable (free) IGF-I are associated with increased steroidogenesis in developing mare follicles.     

Effects of intermittent injections of LHRH on specific binding of 125I-labeled gonadotropins to granulosa and theca, and concentrations of steroids in serum and ovarian follicles during postpartum anovulation in suckled beef cows

To examine ovarian follicular response to low-dose injections of luteinizing hormone-releasing hormone (LHRH), 32 anovulatory, suckled beef cows were allotted to one of four treatment groups and injected with either saline or 500 ng LHRH every 2 h for 48 or 96 h, starting 21.4 +/- .4 d after parturition. Two hours after the last injection of LHRH, cows were ovariectomized and 10 to 15 ovarian follicles per pair of ovaries were removed and categorized by diameter as small (1.0 to 3.9 mm), medium (4.0 to 7.9 mm) or large (greater than or equal to 8.0 mm). Injections of LHRH did not affect (P greater than .10) steroid levels in small follicles or numbers of gonadotropin receptors in small and medium follicles. Concentrations of progesterone in fluid of medium follicles increased 1.5-fold (P less than .05) after 96 h of LHRH, whereas concentrations of estradiol and androstenedione were unchanged. In fluid of large follicles, concentrations of progesterone were fourfold greater (P less than .05) in LHRH-treated than in control cows at 48 h, but by 96 h progesterone was twofold greater (P less than .05) in control than LHRH-treated cows. In large follicles, concentrations of estradiol were unchanged (P greater than .10) after 48 h of LHRH injections but after 96 h estradiol was twofold greater (P less than .05) in LHRH-treated than control cows. Increased concentrations of estradiol in large follicles coincided with increased numbers of binding sites for human chorionic gonadotropin (hCG) but not follicle stimulating hormone (FSH) in granulosa and theca.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of antral follicle populations during the estrous cycle in pigs selected for ovulation rate

The objectives of this study were to characterize and compare ovarian follicular populations in Gene Pool Control (GPC, randomly selected) and Relax Select line (RS, nine generations of selection for high ovulation rate followed by six generations of random selection) gilts during different stages of the estrous cycle. Thirty-five RS and 23 GPC gilts were allotted randomly within litter for ovary recovery on either d 3, 15 or 19 of the estrous cycle. Surface follicles on the ovaries were classified by size (small, less than 3 mm; medium, 3 to 6.9 mm; large, 7 to 12 mm), and counts were recorded for each ovary. Ovarian weight (OW), number of corpora lutea (CL), follicular fluid volume (FFV) from small, medium and large follicles, residual ovarian weight and follicular fluid weight (FFW) also were recorded. Total numbers of small and medium follicles were greatest on d 15, whereas total number of large follicles and FFW were greatest on d 19. The OW, FFW and follicle numbers of all classes were lowest on d 3. The RS gilts expressed longer interestrous intervals (21.9 vs 20.4 d, P less than .05) and higher ovulation rates (18.5 vs 15.3 CL, P less than .01) than GPC gilts. The left ovary of RS gilts was responsible for most of the ovulation rate advantage (10.3 vs 7.4 CL, P less than .01) Overall, GPC gilts had more total small follicles than RS gilts (P less than .01). The advantage was due primarily to higher numbers of small follicles at d 15.(ABSTRACT TRUNCATED AT 250 WORDS)