Effect of addition of a progesterone intravaginal insert to a timed insemination protocol using estradiol cypionate on ovulation rate, pregnancy rate, and late embryonic loss in lactating dairy cows

The objectives of this study were to determine the effects of incorporating a progesterone intravaginal insert (CIDR) between the day of GnRH and PGF2alpha treatments of a timed AI protocol using estradiol cypionate (ECP) to synchronize ovulation on display of estrus, ovulation rate, pregnancy rate, and late embryonic loss in lactating cows. Holstein cows, 227 from Site 1 and 458 from Site 2, were presynchronized with two injections of PGF2alpha on study d 0 and 14, and subjected to a timed AI protocol (100 mixrog of GnRH on study d 28, 25 mg of PGF2alpha on study d 35, 1 mg of ECP on study d 36, and timed AI on study d 38) with or without a CIDR insert. Blood was collected on study d 14 and 28 for progesterone measurements to determine cyclicity. Ovaries were scanned on d 35, 37, and 42, and pregnancy diagnosed on d 65 and 79, which corresponded to 27 and 41 d after AI. Cows receiving a CIDR had similar rates of detected estrus (77.2 vs. 73.8%), ovulation (85.6 vs. 86.6%), and pregnancy at 27 (35.8 vs. 38.8%) and 41 d (29.3 vs. 32.3%) after AI, and late embryonic loss between 27 and 41 d after AI (18.3 vs. 16.8%) compared with control cows. The CIDR eliminated cows in estrus before the last PGF2alpha injection and decreased (P < 0.001) the proportion of cows bearing a corpus luteum (CL) at the last PGF2alpha injection because of less ovulation in response to the GnRH and greater spontaneous CL regression. Cyclic cows had greater (P = 0.03) pregnancy rates than anovulatory cows at 41 d after AI (33.8 vs. 20.4%) because of decreased (P = 0.06) late embryonic loss (16.0 vs. 30.3%). The ovulatory follicle was larger (P < 0.001) in cows in estrus, and a greater proportion of cows with follicles > or = 15 mm displayed estrus (P < 0.001) and ovulated (P = 0.05) compared with cows with follicles <15 mm. Pregnancy rates were greater (P < 0.001) for cows displaying estrus, which were related to the greater (P < 0.001) ovulation rate and decreased (P = 0.08) late embryonic loss for cows in estrus at AI. Cows that were cyclic and responded to the presynchronization protocol (high progesterone at GnRH and CL at PGF2alpha) had the highest pregnancy rates. Incorporation of a CIDR insert into a presynchronized timed AI protocol using ECP to induce estrus and ovulation did not improve pregnancy rates in lactating dairy cows. Improvements in pregnancy rates in cows treated with ECP to induce ovulation in a timed AI protocol are expected when more cows display estrus, thereby increasing ovulation rate.     

Collection and in Vitro Maturation of Equine Oocytes from Estrus,Diestrus and Pregnant Mares

Two experiments were conducted to evaluate oocyte collection rates and in vitro nuclear maturation rates of equine oocytes obtained during diestrus and pregnancy, and to compare these rates with maturation rates in oocytes derived from preovulatory follicles. In Experiment I, transvaginal ultrasound-guided aspiration of follicle was performed during estrus and diestrus in 14 mares over four consecutive cycles. Follicular aspirations during estrus were performed 24 to 27 hours after injection of 2500 IU of hCG given when the largest follicle reached 35 mm in diameter. Oocyte recovery rate from preovulatory follicles was 51% (33/65) in 49 aspiration sessions. Cumulus-oocyte complexes from preovulatory follicles were cultured for 12-15 hours in TCM199 + 10% (NCS) at 38.5°C in 5% CO₂ in air, and 22/33 (67%) were in metaphase II. During diestrus, mares were treated (Group I) or not treated (Group II) daily with equine pituitary extract (EPE) during alternate cycles from days 1 to 14 after the preovulatory aspiration. Diestrous follicles were aspirated when four or more follicles greater than 12 mm in diameter were present. EPE had no effect on the number of follicles that developed during estrus or diestrus (p>0.05). Oocytes were recovered from 119 of 383 diestrous follicles (31 %)in 75 sessions. There was no difference in recovery rates between Groups I and II (p>0.05). Maturation rates for oocytes collected during diestrus, after 42 hours of culture in TCM 199 + 1 μg/ml of Estradiol and lnl/ml of EPE, were not significantly different (p>0.05) between Groups I and II (49% vs. 53%). In Experiment II, mares between 50 and 85 days of pregnancy were used as oocyte donors. The oocyte recovery rate was 53% (66/125). After in vitro maturation for 40 hours (compact COC) or 15 hours (expanded COC), 22% (7/32) and 22% (7/32), respectively, of the oocytes were in metaphase II. It was concluded that: 1) Preovulatory follicles yield a higher percentage of oocytes with a higher rate of maturation to metaphase II than follicles of diestrus and pregnant mares. 2) Diestrous follicles yielded fewer oocytes than follicles of pregnant mares but with a higher percentage of oocyte maturation. Further studies are necessary to determine if oocytes recovered from diestrous follicles and matured in vitro can be fertilized successfully.     

Relationship between the peripheral concentrations of estradiol-17β (E2) and preovulatory characteristics of cumulus-oocyte complexes (COCs) during superovulation treatment in Japanese Black cows

The relationship between the peripheral concentrations of estradiol-17β (E(2)) and the preovulatory characteristics of cumulus oocyte complexes (COCs) during superovulation treatment was investigated in Japanese Black cows. A superovulation regimen with FSH treatment in a descending manner was commenced on day 7 (n=3) or day 10 (n=2) of the estrous cycle (day 0=estrus). Peripheral blood was collected to measure E(2) concentrations twice a day throughout the treatment. Ovariectomies were performed at 100 h after the initial FSH treatment in five cows. Every follicle more than 8 mm in diameter was isolated from the ovaries, and cumulus-oocyte complexes (COCs) were gently aspirated. The COCs were then separated into three groups based on the characteristics of the cumulus (compact, expanded and denuded) and subgrouped based on the stage of the nucleus in the oocytes (GV, GVBD). Plasma E(2) concentrations tended to increase gradually and reached the peak level at around 84 h (E(2)-84: n=3) or 96 h (E(2)-96: n=2) after the initial FSH treatment. The ratio of COCs with expanded cumulus was significantly higher in E(2)-84 than in E(2)-96 (P<0.01). However, there was no difference in the ratio of oocytes showing GVBD between E(2)-84 and E(2)-96 (P=0.73), and the characteristics of the cumulus did not affect the stage of the nucleus in the oocytes in either groups (compact, expanded and nude; P=0.61, 0.81 and 1.00). It was possible that the time until the peak plasma E(2) concentrations after the FSH treatment could become an indicator for the maturation of follicles and oocytes in preovulatory follicles during superovulation treatment in Japanese Black cows.     

Effect of ovarian phase and follicle quality on morphology and developmental capacity of the bovine cumulus-oocyte complex

In order to study the effect of the follicular environment on the quality and developmental competence of cumulus-oocyte complexes (COC), COC were collected from nonatretic (NA), light atretic (LA), atretic (A), and heavily atretic (HA) follicles. Cumulus-oocyte complexes were also collected from early-luteal phase ovaries (EL), from late-luteal phase ovaries (LL), from follicular phase ovaries (F), and from ovaries from non-cyclic animals (NC). Each COC was assigned to one of three quality groups based on the appearance of the cumulus investment: 1) COC-A: compact and bright, 2) COC-B: less compact and dark, and 3) COC-C: strongly expanded cumulus with dark spots. There was a high correlation between follicle quality and the distribution of the COC over the three COC qualities. The COC-A were mainly but not exclusively derived from NA follicles, COC-B were mainly derived from all classes of atretic follicles, and COC-C nearly exclusively originated from HA follicles. The developmental capacity of COC-A and COC-B, which was measured by in vitro embryo production, was consistent over the follicle qualities, except for COC-B obtained from HA follicles. They showed lower development (10.6%, P < .05) compared with COC-B from the other follicle qualities (18.9, 18.7, and 19.8%, respectively, for COC-B from NA, A, and LA follicles). The COC-B from atretic follicles produced more blastocysts (19.8%) than COC-A (12.7%, P < .05). The overall percentage of produced embryos per follicle class seemed to increase with increasing signs of atresia, except if the COC were derived from HA follicles. This increased percentage of embryos was, however, not due to a better quality of COC, but to a higher percentage of COC-B coming from these follicles. The stage of the cycle had no effect on the distribution of the COC over the three COC qualities or on the developmental capacity of COC-A or COC-B, except for COC-A from EL ovaries, which produced more (P < .05) blastocysts than COC-A from the other luteal phases (12.5% vs approximately 8%). A follow-up study was performed trying to elucidate why COC-B possess a higher developmental potency than COC-A. The answer was sought in the oocyte maturation. At several time points during maturation, oocytes were evaluated for their nuclear stage. At all time points COC-B seemed to be a few hours ahead of COC-A, and after 24 h of maturation more COC-B had reached the metaphase-2 stage. This might mean that COC-A need a longer maturation period than COC-B.     

注射hCG后不同时间猪所排卵泡内COC的形态和卵母细胞减数分裂进程

研究了超排处理注射 h CG后不同时间猪卵丘卵母细胞复合体 (COC)的回收率和形态、卵母细胞减数分裂进程以及 COC形态与生发泡 (GV)染色质构型之间的关系。结果表明 :(1)注射 h CG后 4 h COC的回收率为 5 3.1% ,明显低于注射 h CG后 18、2 2、2 4 h(71.2 %、76 .5 %、70 .0 % ,P<0 .0 5 ) ;(2 )注射 h CG后 18、2 2、2 4 h收集的 COC分别有98.9%、98.0 %、91.4 %的卵丘已经发生扩展 ,而注射 h CG后 4 h收集的 COC则无一发生卵丘扩展 ;(3)注射 h CG后 4h,有少量卵母细胞开始恢复减数分裂 ,至 18h左右开始发生生发泡破裂 (GVBD) ,到 2 2～ 2 4 h有 5 8.4 %～ 6 0 .0 %的卵母细胞发生 GVBD;(4)外层卵丘扩展、放射冠轻微扩展的卵母细胞处于 GV- 1期的比例 (6 9.6 % )明显高于卵丘完全扩展的卵母细胞 (47.8% )和放射冠部分扩展、卵丘已经脱落的卵母细胞 (2 9.3% ) ,而后 2者发生 GVBD的比例(40 %、4 4 % )则略高于前者 (2 7% )。     

Changes in follicular endocrinology during final maturation of porcine oocytes

Thirty-one gilts were ovariectomized between 21 and 34 hr after the onset of estrus to compare changes in follicular endocrinology with stages of oocyte maturation. Oocytes were recovered from 6 to 8 mm follicles and classified by stage of meiosis. Remaining follicular fluid was assayed for steroids and dermatan sulfate. Amounts of prostaglandin F2 alpha (PGF2 alpha) and E2 (PGE2) were measured in intramural tissues. Coincident with germinal vesicle breakdown, the follicular content of all steroids except testosterone decreased (P less than .05). As oocytes approached metaphase II, the amount of progesterone within follicles increased (P less than .05), and estradiol continued to decrease (P less than .05). The pattern of dermatan sulfate content was biphasic and peaked at germinal vesicle breakdown and anaphase stages. Amounts of PGF2 alpha and PGE2 within intramural tissues increased (P less than .05) throughout oocyte maturation. Follicular atresia was evident during estrus; however, more (P less than .05) atretic follicles were recovered at germinal vesicle than metaphase II stages (20 vs 3%, respectively). Follicular development, within a gilt, was skewed (P less than .05) and classification of follicles by hormone content demonstrated that a majority were more mature than a minority of less mature follicles. These data suggest that follicular maturation and oocyte development are highly correlated in swine. Furthermore, partitioning the follicular variability by hour and stage of oocyte maturation allowed for more precise assessment of follicular endocrinology than previously reported.     

Characteristics of the cumulus-oocyte complex during the preovulatory period in superovulated heifers]

The nature of cumulus--oocyte complexes was examined in PMSG (group 1, n = 18) and FSH (group 2, n = 30) stimulated heifers. Laparoscopy was performed 65 h after cloprostenol application. The number of follicles was 13.67 +/- 0.75 and 12.67 +/- 0.81 (P greater than 0.05) in group 1 and 2, respectively (Tab. I). The recovery rate of oocytes was 56% in the first and 67% in the second group (Tab. I). The cumulus oophorus was divided into three groups: compact, expanded and partial (Tab. II). Most oocytes (65 and 75% in the first and second group, respectively) exhibited an expanded cumulus (P greater than 0.05). In the first and second group 11 and 26% (P less than 0.01) of oocytes with the extruded first polar body were aspirated (Tab. III). As judged from the pool of visible follicles, the superovulation response to stimulatory treatment and recovery rate of oocytes in the present experiment were not different from the results published earlier. The degree of the cumulus oophorus expansion is an indicator for the evaluation of cumulus--oocyte complexes. After the preovulatory LH peak the disintegration of cumulus oophorus proceeds from glycosaminoglycan accumulation. In our experiment this effect resulted in a significantly higher number of oocytes with expanded cumulus in both treatments. The enlargement of perivitelline space is related to a subsequent release of the first polar body in the preovulatory period. It can be seen from our results that after FSH treatment it is possible to reach the high number of oocytes with the extruded polar body.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maternal Liver Damage Delays Meiotic Resumption in Bovine Oocytes through Impairment of Signalling Cascades Originated from Low p38MAPK Activity in Cumulus Cells

The main objective of the present study is to investigate the molecular mechanism underlying the delay in progression of nuclear maturation in oocytes derived from cows with damaged livers (DL cows), which was previously reported. In present study, delayed progression of nuclear maturation of oocytes derived from DL cows relative to oocytes derived from cows with healthy livers (HL cows) was accompanied by low maturation promoting factor (MPF) activity (0.43 fold, p < 0.05). When cumulus cells were removed from cumulus‐oocyte complexes and the denuded oocytes were cultured, there was no difference in the progression of nuclear maturation between the two liver conditions. In addition, gap junctional communication (GJC) between the oocyte and cumulus cells was higher in DL cows than in HL cows at 3 and 7 h of in vitro maturation (IVM) (p < 0.05). Supplementation of IVM medium with epidermal growth factor (EGF) increased the ratio of germinal vesicle breakdown (GVBD) of oocytes derived from DL cows to the level seen in oocytes derived from HL cows. Additionally, the level of p38MAPK phosphorylation at 0 h of IVM was significantly lower in cumulus cells derived from DL cows than in cumulus cells derived from HL cows (HL cows, 53.5%; DL cows, 28.9%; p < 0.05). Thus, a low level of p38MAPK phosphorylation in cumulus cells induced slow GJC closure between oocyte and cumulus cells, which resulted in slow meiotic maturation of oocytes derived from DL cows.     

Effect of Initial Cumulus Morphology on Meiotic Dynamic and Status of Mitochondria in Horse Oocytes during IVM

The aim of this investigation was to examine the chromatin configuration of the nucleus, pattern of mitochondrial aggregation and mitochondrial activity in parallel studies in the same horse oocytes. Horse oocytes recovered by ultrasound-guided follicle aspiration in vivo were classified according to two main initial cumulus morphologies as having compact or expanded cumulus. The percentage of oocytes with a diplotene meiotic configuration at the time of recovery from the follicles was highest in compact oocytes. Oocytes with expanded cumulus layers at the time of recovery matured more rapidly in vitro and reached a proportion >50% at the metaphase II stage (M 2) sooner during in vitro maturation (IVM), than did compact oocytes. The mitochondrial aggregation pattern changed from finely distributed (Type 1) through crystalline (Type 2) to an aggregated, granulated appearance (Type 3) during IVM. The pattern of mitochondrial aggregation at the time of recovery was associated with the initial cumulus morphology of the oocyte, in that compact oocytes had a higher proportion of Type 1 aggregation, whereas expanded oocytes had a higher proportion of Type 3. The fluorescence intensity of metabolic active mitochondria, measured by fluorescence intensity (Em 570) per oocyte after MitoTracker CMTM Ros orange labelling, increased in the oocytes during IVM and depended on initial cumulus investment. Oocytes with the granulated type of aggregated mitochondria Type 3 had the highest level of metabolic activity and were in more progressed stages of meiosis (A 1-M 2). Oocytes initially having expanded layers of cumulus reached significantly higher levels of mitochondrial activity after IVM than did oocytes initially having compact cumuli. During resumption of meiosis the mitochondrial activity of oocytes with initially expanded cumulus increased continuously up to M 2, whereas in oocytes from compact cumulus-oocyte complex (COC), the activity declined after A 1/T 1 stages of meiosis.     

Ovarian 3'5'-cyclic adenosine monophosphate and prostaglandins: a sequential comparison of gonadotropin-stimulated events in small and large ovine follicles

Luteinizing hormone (LH) stimulates a cascade of ovarian hormonal events that culminate in ovulation. This study was designed to investigate, in sheep, sequential changes in prostaglandin (PG) E2, PGF2 alpha, 6-keto-PGF1 alpha, and cyclic adenosine monophosphate (cAMP) in the theca, granulosa and follicular fluid of large preovulatory follicles and small nonovulatory follicles in response to LH. On d 15 postestrus, preovulatory or nonovulatory follicles were injected intrafollicularly with saline or LH. Ewes were then ovariectomized at 0, 2, 4, or 8 h postinjection. Injected follicles were excised; theca, granulosa and fluid were separated, weighed and assayed for cAMP and PG. Contents of cAMP in the theca, granulosa and fluid of preovulatory follicles increased (P less than .01) 2 to 4 h after injection of LH. Increases (P less than .05) in contents of PGE2 and PGF2 alpha in the theca and fluid of preovulatory follicles were observed between 4 and 8 h after injection of LH. The time courses of LH-induced synthesis of PGE2 and PGF2 alpha in preovulatory follicles were parallel. Luteinizing hormone had no effect on PGE2, PGF2 alpha or cAMP in any compartment of small follicles. Contents of 6-keto-PGF1 alpha varied with time in both theca and granulosa of large and small, saline- and LH-injected follicles. Although specific increases in cAMP and PG followed an injection of LH only in large follicles, the parallel temporal relationship of PGE2 and PGF2 alpha did not explain the dichotomous functions ascribed to PGE2 and PGF2 alpha during the periovulatory period.     

Serum concentrations of IGF-I in postpartum beef cows

Four experiments assessed changes in serum IGF-I under various physiologic conditions in postpartum cows. In Exp. 1, anestrous suckled cows (n = 25) were infused for 6 d with either saline or glucose at two different infusion rates. In Exp. 2, anestrous cows (n = 29) received either a saline (weaned and suckled controls) or 3 g/d phlorizin (weaned phlorizin) infusion for 3 d. Calves from the weaned groups were removed from 15 h before and throughout infusions. In Exp. 3, cycling suckled cows (n = 20) received prostaglandin F2 alpha (PGF2 alpha) when the 5-d saline or phlorizin infusion began. In Exp. 4, suckled cows (n = 20) had ad libitum access to feed or received 50% of control feed consumption from 30 to 40 d postpartum. Increasing glucose availability (Exp. 1) increased (P less than .05) serum IGF-I by 30 to 35%. IGF-I remained stable after weaning (Exp. 2) in phlorizin-infused cows (128.8 +/- 12.7 ng/ml), but increased (P less than .05) by 3 d after calf removal in weaned control cows (152.2 +/- 7.5 ng/ml). IGF-I also remained stable in phlorizin-infused cows following PGF2 alpha injection (Exp. 3), but increased in control cows by 2 d after PGF2 alpha (156.8 +/- 18.3 on d 2 vs. 133.7 +/- 9.8 ng/ml pre-injection; P less than .05) and remained elevated (P less than .05) during the periovulatory period. In cows receiving restricted feed intake (Exp. 4), IGF-I decreased by approximately 50% within 4 d of feed restriction (71.3 +/- 9.4 vs 137.4 +/- 16.6 ng/ml; P less than .05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Nitrogen metabolism and fertility in cattle: II. Development of oocytes recovered from heifers offered diets differing in their rate of nitrogen release in the rumen

In vitro blastocyst production was determined for oocytes recovered postmortem from 48 beef x dairy heifers offered low (Low NH3) or high (High NH3) plasma ammonia-generating diets during the period of late antral follicle development. Following the establishment of a reference estrus (d 0), the experimental diets were offered for an 18-d period starting on d 3 and during which a second estrus was induced (d 16) 4 d before the animals were slaughtered. Blood samples collected at varying intervals were analyzed for ammonia, urea, progesterone, and LH. Ovarian folliculogenesis was monitored daily by transrectal ultrasonography. Ovaries were collected at slaughter and cumulus-oocyte complexes were aspirated from small (1 to 4 mm) and medium-sized (> 4 to 8 mm) sized follicles. In vitro-matured and -fertilized putative d-1 zygotes were cultured for a further 7 d in vitro and embryo development and metabolism were assessed. Relative to the low-NH3-generating diet, the high-NH3-generating diet increased peak postprandial levels of plasma ammonia (326.1 +/- 43.3 vs 52.1 +/- 7.4 micromol/L; P < .001), mean levels of plasma urea (7.0 vs 5.7 mmol/L; SED = .2; P < .001), peak levels of plasma progesterone prior to induced luteolysis (8.9 +/- .4 vs 6.8 +/- .3 microg/L; P < .001), and follicular fluid levels of ammonia (267 +/- 18 vs 205 +/- 20 nmol/mL; P < .05) and progesterone (351 +/- 69 vs 199 +/- 26 ng/mL; P < .05). The timing and level of the preovulatory LH surge was not affected by dietary treatment. Of oocytes cultured, cleavage (47.4 vs 62.4%; P = .02) and blastocyst production (10.9 vs 20.6%; P = .06) rates were reduced when the oocytes were derived from heifers offered the high- rather than the low-NH3-generating diets. There were interactions between dietary treatment and follicle size class, which indicated that fewer blastocysts were produced from cleaved oocytes derived from medium-sized follicles of heifers offered the high-NH3 treatment but that de novo protein synthesis was increased in such embryos. In conclusion, exposure to high levels of ammonia and(or) urea in vivo can significantly compromise the subsequent capacity of oocytes to develop to blastocysts in vitro, and oocytes recovered from medium-sized follicles are particularly sensitive to this effect.     

The Phosphodiesterase Inhibitor,Isobutyl-1-Methylxanthine Prevents the Sudden Drop in Cyclic Adenosine Monophosphate Concentration and Modulates Glucose Metabolism of Equine Cumulus–Oocyte Complexes Matured in Vitro

Spontaneous nuclear maturation of mammalian oocytes can occur when physically removed from the ovarian follicle during in vitro oocyte maturation (IVM), largely because of a decrease in cyclic adenosine monophosphate (cAMP) concentration. Modulation of oocyte cAMP during IVM by using phosphodiesterase inhibitors has been shown to maintain elevated oocyte cAMP concentrations and control meiotic resumption of bovine and ovine oocytes. This study determined the effect of inclusion of isobutyl-1-methylxanthine (IBMX) during collection and the first 12 hours of incubation of equine oocytes on cAMP concentration and glucose metabolism of cumulus–oocyte complexes (COCs). Abattoir-derived COCs were collected in aspiration medium with (Asp-IBMX) or without (Asp) IBMX. Cumulus–oocyte complexes were then incubated for 12 hours in IVM medium with (Mat-IBMX) or without (Mat) IBMX, followed by additional 24 hours in Mat medium. The cAMP concentration, glucose consumption, lactate production, and metaphase II rates of the COCs were assessed. Cumulus–oocyte complexes aspirated into Asp-IBMX (62.2 ± 2.6 fmol per COC) medium had higher cAMP concentration than Asp (31.8 ± 2.8 fmol per COC) control group (P < .05). Likewise, at 12 hours of IVM, Mat-IBMX group (33.2 ± 2.1 fmol per COC) had higher cAMP concentration than the Mat group (7.68 ± 0.5 fmol per COC; P < .05). Glucose consumption and lactate production were lower during the first 12 hours of incubation in COCs cultured in Mat-IBMX (P < .05). Isobutyl-1-methylxanthine prevented the rapid drop in cAMP concentration and altered metabolism of glucose by the COC. Preventing the sudden drop in cAMP prevents the premature nuclear maturation of in vitro–matured oocytes causing poor developmental competence.     

Influence of co‐culture with denuded oocytes during in vitro maturation on fertilization and developmental competence of cumulus‐enclosed porcine oocytes in a defined system

Co‐culture of cumulus‐oocyte complexes (COCs) with denuded oocytes (DOs) during in vitro maturation (IVM) was reported to improve the developmental competence of oocytes via oocyte‐secreted factors in cattle. The aim of the present study was to investigate if addition of DOs during IVM can improve in vitro fertilization (IVF) and in vitro culture (IVC) results for oocytes in a defined in vitro production system in pigs. The maturation medium was porcine oocyte medium supplemented with gonadotropins, dbcAMP and β‐mercaptoethanol. Cumulus‐oocyte complexes were matured without DOs or with DOs in different ratios (9 COC, 9 COC+16 DO and 9 COC+36 DO). Consequently; oocytes were subjected to IVF as intact COCs or after denudation to examine if DO addition during IVM would affect cumulus or oocyte properties. After fertilization, penetration and normal fertilization rates of zygotes were not different between all tested groups irrespective of denudation before IVF. When zygotes were cultured for 6 days, no difference could be observed between all treatment groups in cleavage rate, blastocyst rate and cell number per blastocyst. In conclusion, irrespective of the ratio, co‐culture with DOs during IVM did not improve fertilization parameters and embryo development of cumulus‐enclosed porcine oocytes in a defined system.     

Effects of low-dose follicle-stimulating hormone administration on follicular dynamics and preovulatory follicle characteristics in dairy cows during the summer

The well-documented phenomenon of reduced conception rate in dairy cows during the hot season involves impaired functioning of the ovarian follicles and their enclosed oocytes. Three experiments were performed to examine the administration of low doses of follicle-stimulating hormone (FSH) to induce turnover of follicles that are damaged upon summer thermal stress and to examine whether this FSH administration has beneficial effects on preovulatory follicles. In experiment 1, synchronized heifers were treated with 100 mg of Folltropin-V (n = 7) or 4.4 mg of Ovagen (n = 6) on day 3 of the estrous cycle. Treatment with both FSH sources resulted in greater (P < 0.05) numbers of follicles than in control animals (n = 12) on day 6 of the estrous cycle, indicating that low doses of FSH can increase the number of emerging follicles in a follicular wave. In experiment 2, milking cows were assigned to a control group (n = 4) or treated with 2.2 mg (FSH-2.2; n = 6) or 4.4 mg (FSH-4.4; n = 5) Ovagen. Follicle-stimulating hormone was administrated on day 3 or 4 and day 10 or 11 of the estrous cycle, coinciding with emergence of the first and second follicular waves, respectively. The number of follicles emerging during the first wave tended to be higher (P < 0.1) in FSH-4.4-treated cows than in controls. The second-wave dominant follicles emerged 2 d later in the treated cows and were smaller in diameter (P < 0.05) than controls, 2 d before aspiration. Despite being younger, the preovulatory follicles of FSH-4.4 cows expressed a steroidogenic capacity that was similar to controls with a tendency toward greater insulin concentrations (P < 0.09). In experiment 3, milking cows were assigned to a control group (n = 6) or treated with 4.4 mg Ovagen (FSH-4.4; n = 6). Follicle-stimulating hormone was administrated on day 3 and day 12 or 13 of the estrous cycle. The number of emerging follicles was higher (P < 0.05) in the treated vs control cows. However, the features of the preovulatory follicle developed in the subsequent cycle did not differ between groups. In summary, low doses of FSH can efficiently induce follicular turnover accompanied by a modest effect on the preovulatory follicle of the treated cycle. It appears that the administration of low doses of FSH, precisely timed to synchronize with the emergence of follicular waves, might have a beneficial effect on the preovulatory follicle and its enclosed oocyte.     

Light and transmission electron microscopy of immature camelus dromedarius oocyte

In order to provide a consistent system for laboratory production of embryos, the characteristics of immature camel oocyte must first be described. The objective of this study was to define ultrastructural features of immature camel oocyte. Ovaries were obtained from camels at a local abattoir, and then transported to the laboratory within 2 h. Camelus cumulus oocyte complexes (COCs) were aspirated from 2-6 mm follicles using a 22-gauge needle. Excellent and good quality COCs were selected and prepared for transmission electron microscopy study using a cavity slide. The fine structure of camel oocyte is morphologically similar to that of other mammalian oocytes. However, some minor differences exist between COC of camel and other mammalian species. Different size and shape of membrane-bound vesicles, lipid droplet, mitochondria and cortical granules were distributed throughout the ooplasm. Discrete or in association with endoplasmic reticulum, Golgi complexes were observed in the periphery of the oocytes. The majority of the oocytes were in the germinal vesicle stage.     

Comparison of follicular and oocyte development and reproductive hormone secretion during the ovulatory period in Hungarian native breed, Mangalica, and Landrace gilts

Only a very small amount of physiological data is available about the low fertility (mean litter size is 5.7+/-0.8) of Hungarian native breed, Mangalica (M), sows. The aim of the present paper is to reveal the differences in preovulatory follicle development and intrafollicular oocyte maturation between M and Landrace (L) gilts, with special reference to the peri- and postovulatory secretion and peripheral concentrations of estradiol-17beta (E2), progesterone (P4), and luteinizing hormone (LH). The number of preovulatory follicles was 6.8+/-1.4 and 19.6+/-6.6 in M and L gilts, respectively. A lower degree of cumulus expansion and a lower percentage of mature oocytes (TI/M II) was noted in M. Higher LH and E2 peak levels, a longer E2 to LH peak interval, and lower embryo survival was confirmed. Interestingly, despite the lower number of corpora lutea, a higher peripheral blood level of P4 was shown in M than in L gilts. Both diminished follicular development and protracted oocyte maturation may be involved in low fecundity in M, and the present findings may explain these reproductive phenomena.     

Effect of prostaglandin F2 alpha-induced luteolysis on in vivo and in vitro progesterone production by individual placentomes of cows

This study investigated placental progesterone production by bovine placentomes. Catheters were placed in the femoral artery (FA) and in the caruncular artery (CA), caruncular vein (CV) and lymphatic vessel of a prominent placentome of 13 cows at 200 d of gestation. Four of the 13 cows were given prostaglandin F2 alpha (PGF2 alpha) after surgery, and blood and lymph were collected for progesterone determination. After 24 h, progesterone was higher (P less than .01) in FA and CA plasma from control cows that FA and CA plasma from PGF2 alpha-treated cows (5.11 +/- .29 and 5.17 +/- .64 vs 1.41 +/- .08 and 1.15 +/- .08 ng/ml, respectively), but CV concentrations were similar (3.38 +/- .30 vs 2.56 +/- .24, respectively). There was a net uptake of progesterone by placentomes from control cows (P less than .01) but a net secretion in PGF2 alpha-treated cows (P less than .05). Lymph contained low progesterone concentrations regardless of treatment. Cows were slaughtered at 240 d of gestation. Placentomes were removed and perfused with pregnenolone through the maternal and fetal arteries. Fetal venous effluent contained more progesterone than maternal venous effluent (P less than .001) in both groups, and fetal venous effluent of placentomes from PGF2 alpha-treated cows contained more progesterone than that from control cows (P less than .05). Maternal and fetal components of other placentomes were cultured alone or in co-culture along with pregnenolone and (or) epostane. Fetal tissue produced more progesterone (P less than .001) than maternal tissue when each was cultured alone, but fetal tissue production declined when co-cultured with maternal tissue.(ABSTRACT TRUNCATED AT 250 WORDS)     

次黄嘌呤对猪卵母细胞体外自发成熟抑制作用的研究

利用猪卵母细胞体外无血清培养技术,研究了次黄嘌呤（ＨＸ）对猪卵母细胞体外自发成熟的抑制作用。猪卵丘－卵组细胞复合体（ＣＯＣ）和裸卵母细胞（ＤＯ）取自初情期猪卵巢,培养在Ｍ－１９９培养液中,并施以各种自理培养不同时间后,观察卵母细胞核成熟（ＧＶＢＤ）情况。实验结果表明：⑴ＨＸ（１－４mmol/Ｌ）对猪ＣＯＣ的自发成熟具有抑制 作用,且具有剂量领事关系。４mmol/Ｌ的ＨＸ对ＣＯＣ和ＤＯ自发成熟的抑制作用,且具