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Veterinarians and scientists involved in applied and basic research in cattle require a lexicon of terms that is used uniformly so that diagnoses and inference of results between and among studies can be correctly interpreted and substantiated or negated and therapy and hypotheses can be formulated without unnecessary confusion and redundancy in treatments and experiments. This review provides a compilation of many of the classical and contemporary terms used in association with ovarian dynamics primarily during the estrous cycle in cattle, which can also apply to other reproductive states. While many classical terms used to describe healthy and diseased conditions associated with follicles and corpora lutea are still applicable today, there are some that have become antiquated (e.g., cystic corpus luteum, cystic ovarian degeneration, luteolysis, and granulosa cell tumor), due, in part, to advanced technology (e.g., ultrasonography) and a more thorough understanding of ovarian function. In this regard, older terms have been revised (e.g., corpus luteum with a cavity, follicular and luteinized-follicular cysts, structural and functional luteal regression, and granulosa-theca cell tumor) and newer terms have been coined (e.g., follicle deviation) and advocated herein. Defining and adopting terminology used in bovine reproduction that is clear, precise and understandable and available in a single source, is expected to make the exchange of clinical and research information and outcomes more effective, safe, and economical.

Relaxin-like factor/insulin-like peptide (INSL)-3 is highly expressed in the bovine corpus luteum throughout the estrous cycle and pregnancy. Demonstration of translation of the relaxin-like factor message was previously shown for the follicle but not the corpus luteum. In this study, relaxin-like factor mRNA was highly expressed in the corpus luteum on days 7 and 14 of pregnancy. Tissues were fixed in 10% neutral buffered formalin, and utilizing two different antibodies to relaxin-like factor, W3 rabbit anti-bovine and 2-8F mouse anti-bovine, relaxin-like factor was localized in fibroblast-like cells. Staining was also observed in the Leydig cells of bovine testicular tissue. No staining was observed in small and large steroidogenic luteal cells, indicating a nonsteroidogenic source of luteal relaxin-like factor. Definitive cell type identification is currently being determined via electron microscopy.

The purpose of this study was to evaluate associations of lactation, somatic cell count score (SCCS) at breeding, milk yield, lactation number, interval from calving to breeding (days open), number of times inseminated, and season of breeding on fetal loss for lactating Holstein females (both first-parity and multiparous cows) and nonlactating Holstein heifers in a hot climate. Females were palpated between d 40 and 50 of gestation and again at d 70 to 80 to determine pregnancy status. Early fetal loss was defined as a loss that occurred after d 40 to 50 but before d 70 to 80. Mid-to-late fetal loss represented losses after d 70 to 80 but before expected calving. Lactating females had higher early (P = 0.055) and mid-to-late fetal loss (P < 0.05) than nonlactating heifers. Those lactating females with increased days open experienced greater early (P < 0.05) and mid-to-late fetal loss (P = 0.055), whereas lactating females with an elevated SCCS encountered greater mid-to-late fetal loss (P < 0.01). Milk yield, lactation number, number of times inseminated, and season were not associated with early or mid-to-late fetal loss. For nonlactating heifers, there were no associations between number of times inseminated, season, or age at breeding on early or mid-to-late fetal loss. In conclusion, lactating females were more likely to suffer early and mid-to-late fetal loss than nonlactating heifers. Also, days open and SCCS at breeding were related to ability of lactating females to maintain pregnancy, but there were no relationships between fetal loss and milk yield, lactation number, number of times inseminated, or season.

Spatial and temporal heterogeneity of superficial muscle strain during in situ fixed-end contractions.Drost MR, Maenhout M, Willems PJ, Oomens CW, Baaijens FP, Hesselink MK.Department of Movement Sciences, Nutrition and Toxicology Research Institute Maastricht, Maastricht University, P.O. Box 616, 6200, Maastricht, The Netherlands. maarten.drost@bw.unimaas.nlNumerical models of contracting muscle offer a powerful tool to study local mechanical load. For validation of these models, the spatial and temporal distribution of strain was quantified in fixed-end contracting rat tibialis anterior muscle in situ at optimal muscle length (L(o)) and at 120 degrees plantar flexion as well as at 125 and 33Hz stimulation frequency. We studied the hypothesis that after termination of stimulation in situ muscle segments near the motor endplates elongate while segments away from the endplates shorten. We show that both spatial and temporal inhomogeneities in muscle deformation occurred during contraction. Muscle plateau shortening strain equalled 4.1%. Maximal plateau shortening of a muscle segment was much larger (9.6%) and occurred distally (at 0.26 of the scaled length of the muscle). Manipulating torque levels by decreasing the stimulation frequency at the same muscle length induced a decrease in torque ( approximately 20%) with a smaller effect on the level and no effect on the pattern of muscle deformation. During relaxation, distal segments actively shortened at the expense of proximal muscle segments, which elongated. The segments undergoing lengthening were nearer to motor endplates than segments undergoing shortening.In conclusion, the present study provides experimental data on magnitude of contraction-induced deformation needed for validation of numerical models. Local muscle deformation is heterogeneous both temporally and spatially and may be related to proximity to the motor endplates

Timed embryo transfer (TET) using in vitro produced (IVP) embryos without estrus detection can be used to reduce adverse effects of heat stress on fertility. One limitation is the poor survival of IVP embryos after cryopreservation. Objectives of this study were to confirm beneficial effects of TET on pregnancy rate during heat stress as compared to timed artificial insemination (TAI), and to determine if cryopreservation by vitrification could improve survival of IVP embryos transferred to dairy cattle under heat stress conditions. For vitrified embryos (TET-V), a three-step pre-equilibration procedure was used to vitrify excellent and good quality Day 7 IVP Holstein blastocysts. For fresh IVP embryos (TET-F), Holstein oocytes were matured and fertilized; resultant embryos were cultured in modified KSOM for 7 days using the same method as for production of vitrified embryos. Excellent and good quality blastocysts on Day 7 were transported to the cooperating dairy in a portable incubator. Nonpregnant, lactating Holsteins (n = 155) were treated with GnRH (100 microg, i.m., Day 0), followed 7 days later by prostaglandin F2alpha (PGF2alpha, 25 mg, i.m.) and GnRH (100 microg) on Day 9. Cows in the TAI treatment (n = 68) were inseminated the next day (Day 10) with semen from a single bull that also was used to produce embryos. Cows in the other treatments (n = 33 for TET-F; n = 54 for TET-V) received an embryo on Day 17 (i.e. Day 7 after anticipated ovulation and Day 8 after second GnRH treatment). The proportion of cows that responded to synchronization based on plasma progesterone concentrations on Day 10 and Day 17 was 67.7%. Pregnancy rate for all cows on Day 45 was higher (P0.05) in the TET-F treatment than for the TAI and TET-V treatments (19.0 +/- 5.0,6.2 +/- 3.6, and 6.5 +/- 4.1%). For cows responding to synchronization, pregnancy rate was also higher (P0.05) for TET-F than for other treatments (26.7 +/- 6.4, 5.0 +/- 4.3, and 7.4 +/- 4.7%). In the TET-F treatment group, cows producing more milk had lower (P0.05) pregnancy rates than cows producing less milk. In conclusion, ET of fresh IVP embryos can improve pregnancy rate under heat stress conditions, but pregnancy rate following transfer of vitrified embryos was no better than that following TAI.

J Biomech 2001 Oct;34(10):1365-8 Related Articles, Books, LinkOut Passive transverse mechanical properties of skeletal muscle under in vivo compression.Bosboom EM, Hesselink MK, Oomens CW, Bouten CV, Drost MR, Baaijens FP.Department of Materials Technology, Eindhoven University of Technology, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands. e.m.h.bosboom@tue.nlThe objective of the present study is to determine the passive transverse mechanical properties of skeletal muscle. Compression experiments were performed on four rat tibialis anterior muscles. To assess the stress- and strain-distributions in the muscle during the experiment, a plane stress model of the cross section was developed for each muscle. The incompressible viscoelastic Ogden model was used to describe the passive muscle behaviour. The four material parameters were determined by fitting calculated indentation forces on measured indentation forces. The elastic parameters, mu and alpha, were 15.6+/-5.4 kPa and 21.4+/-5.7, respectively. The viscoelastic parameters, delta and tau, were 0.549+/-0.056 and 6.01+/-0.42 s. When applying the estimated material parameters in a three-dimensional finite element model, the measured behaviour can be accurately simulated.

Three experiments were conducted to evaluate the effect of an induced first wave persistent dominant follicle on folliculogenesis and ovulatory responses induced by FSH. On d 6 of a synchronized estrous cycle (d 0 = estrus), cows were treated with a Syncromate-B implant and two injections of PGF2, (25 mg, 0700 h; 15 mg, 1900 h, i.m.). Cows in the control group retained a first-wave persistent dominant follicle, but in the aspirated group, the first-wave dominant follicle was removed via transvaginal aspiration on d 10 (d 0 = estrus). Beginning on d 12, cows received 32 mg of FSH-P i.m. in decreasing doses at 12-h intervals over a 4-d period. On d 15, the Syncromate-B implant was removed, and cows were ovariectomized (experiment 1, n = 8) or inseminated (experiment 2, n = 11) at 10 and 22 h after the onset of estrus. Cows in experiment 3 received a used controlled intravaginal drug releasing (CIDR) device and two injections of PGF2alpha (25 mg, 0700 h; 15 mg, 1900 h; i.m.) on d 6. On d 8, the first-wave dominant follicle was aspirated (n = 6) or left intact (n = 5), and FSH treatment was initiated (20 mg of Folltropin in decreasing doses at 12-h intervals over a 4-d period), and on d 10 the used CIDR device was removed from all cows. Ovarian follicle size and number were examined daily by ultrasonography from d 5 of the estrous cycle. The persistent dominant follicle increased in size from 10.7 mm on d 5 to 15.4 mm on d 10 (experiments 1 and 2), and from 9 mm on d 5 to 20.4 mm on d 11 (experiment 3). From d 11 to 14, the number of class 1 (2 to 5 mm) follicles was lower in the aspirated group than in the control group; the number of class 2 (6 to 9 mm) follicles was higher on d 12 and 13 for the aspirated group (experiments 1 and 2). The number of class 3 (or =10 mm) follicles was higher in the aspirated group on d 14 to 16, but the same on d 17. Ovarian and embryo responses to superovulation did not differ between groups. In experiment 3, the numbers of class 1, 2, and 3 follicles, as well as ovarian and embryo responses following ovulation did not differ between groups. Initiation of exogenous FSH treatment appears to override any systemic inhibitory effect that a persistent dominant follicle may be exerting at the pituitary and possibly the ovary.

Body condition may influence pregnancy rates to a timed insemination (Ovsynch/TAI) protocol and affect the economical performance of dairy farms. The objectives were to compare pregnancy rates using the Ovsynch/TAI protocol for the first service of lactating dairy cows with body condition scores2.5 (scale: 1 to 5, low BCS group) versusor = 2.5 (control group) and to estimate the economic impact of the effect of body condition on reproductive performance. At 63 +/- 3 d post partum, cows were assigned to 2 experimental groups (low BCS = 81; control = 126), and were treated with GnRH at d 0 and with PGF2alpha 7 d later. At 48 h after PGF2alpha, cows received an injection of GnRH and were inseminated 16 h later. Pregnancy rates to the Ovsynch/TAI protocol were lower for the low BCS group than for the control group at 27 d (18.1 +/- 6.1%33.8 +/- 4.5%; P0.02) and at 45 d (11.1 +/- 5.4%25.6 +/- 4.1%; P0.02) after insemination. Economic analysis indicated that reducing the percentage of the herd in low body condition increases net revenues per cow per year. Body condition influenced pregnancy rates to the Ovsynch/TAI protocol.

A reservoir for sperm has been found in the oviductal isthmus in several species. Sperm are apparently trapped in the reservoir by binding to the oviductal epithelium, although other factors may be involved. We hypothesized that binding sites for bovine sperm are limited to the isthmus and are regulated by the hormonal state of the cow. Ipsilateral oviducts were obtained from heifers that were preovulatory (in estrus), had ovulated recently (within 12 h), or were in diestrus (Day 10). The isthmic and the ampullar epithelium were milked out and incubated separately in serum-free (SFRE-199-2) medium, at 39 degrees C in 5% CO2. Frozen-thawed sperm from bulls were added to the epithelium and coincubated for 15 min. The number of spermatozoa that bound to explants was not affected by stage of cycle or by anatomic origin of the explants (p > 0.05). In an additional experiment, oviducts were infused with sperm in vivo and then prepared for scanning electron microscopy, which revealed that sperm were associated with ciliated epithelium in both the isthmus and ampulla. Thus, bovine sperm may form a reservoir in the isthmic end of the oviduct because it is the first oviductal region that they encounter.

Two experiments tested the effect of recombinant ovine and bovine interferon-tau on corpus luteum lifespan, interestrous interval, and oxytocin-induced uterine secretion of prostaglandin F2 alpha. Cows received intrauterine injections of 100 micrograms of recombinant ovine interferon-tau plus 1.4 mg of BSA or of 1.5 mg of BSA alone in Experiment 1 and 200 micrograms of recombinant bovine interferon-tau plus 1.3 mg of BSA or 1.5 mg of BSA alone in Experiment 2. Twice daily injections (0700 and 1900 h) were split evenly between the uterine horns from d 14 to 24 of the experimental estrous cycle via an AI pipette in Experiment 1 and via intrauterine catheters in Experiment 2. On d 17, cows were injected with 100 IU of oxytocin, and plasma was collected for analysis of 13,14-dihydro-15-keto-prostaglandinF2 alpha. Recombinant ovine interferon-tau extended the lifespan of the corpus luteum (27.5 vs. 19.2 d) and interestrous interval (30.5 vs. 20.6 d) and abolished the oxytocin-induced increase in 13,14-dihydro-15-keto-prostaglandinF2 alpha, which peaked at 30 min for the BSA control group (210.8 pg/ml). Recombinant bovine interferon-tau also extended the lifespan of the corpus luteum (29.0 vs. 21.4 d) and interestrous interval (31.5 vs. 22.6 d) and abolished the oxytocin-induced increase in 13,14-dihydro-15-keto-prostaglandin F2 alpha, which peaked at 30 min for the BSA control group (205.6 pg/ml). In conclusion, recombinant ovine interferon-tau and recombinant bovine interferon-tau were effective antiluteolytic agents in cattle.

Gonadotropin-releasing hormone (GnRH) and its analogues cause an acute secretion of luteinizing hormone (LH) and follicle stimulating hormone (FSH) such that concentrations in peripheral blood are elevated for a 3–5 h period. GnRH-induced alterations in the function of the corpus luteum (CL) or follicle appear to be indirect through alterations in LH and FSH secretion. Repeated injections of GnRH during diestrus or single injections late in diestrus will cause acute increases in plasma progesterone and a delay in CL regression. Injections or continual administration of GnRH during early phases of CL development appear to augment CL differentiation and alter subsequent CL function. These effects are attributable to induced increases in LH. Injections of GnRH during the estrous cycle will re-synchronize follicle development owing to ovulation or luteinization of the dominant follicle leading to subsequent recruitment and selection of a new dominant follicle during a 7 day period. Injection of GnRH followed by injection of prostaglandin (PGF2α) at 6 or 7 days is a system of estrous synchronization in which follicle development and CL regression are both synchronized and fertility at the induced estrus is good. Injection of GnRH during the luteal phase post-insemination (e.g. Days 12–14) or post-embryo transfer, to alter CL and/or follicular function, has not resulted in a consistent increase in pregnancy rates. An overall assessment of studies that injected GnRH at the time of insemination in first service postpartum cows or in repeat breeders is rather disappointing. Considerable variation existed among studies within both types of cow populations relative to significant differences, directions of pregnancy rate change ( + vs. −), and magnitude of pregnancy rate increases. Recent findings indicate that timing of GnRH injections closer to the onset of estrus may be beneficial in increasing the conception rate. Utilization of GnRH in combination with progesterone and PGF results in an acute treatment sequence to program follicular development, ovulation and a subsequent cycle in cows with follicular cysts. The use of GnRH, with or without PGF, as a reproductive management program in the early postpartum period has not shown a clear improvement in subsequent reproductive efficiency. Development of precise systems to control ovarian function and reproductive efficiency with GnRH and other pharmaceutical agents is possible. However, such advancements must be founded on a clear understanding of GnRH-induced physiological effects and ability to capture any advantage by good management of the farm unit.

The objective of this study was to determine whether bovine embryos become more resistant to deleterious effects of maternal heat stress as early embryonic development progresses. Superovulated, lactating Holstein cows were bred by AI and assigned to be heat stressed on d 1, 3, 5, or 7 of pregnancy (d 0 = day of estrus) or not heat stressed (control). Embryos were retrieved from the uterus on d 8 and evaluated for viability and stage of development. Compared with embryos of control cows, embryos of cows receiving heat stress on d 1 had decreased viability and development. Maternal heat stress on other days had no detrimental effect on embryonic viability or stage of development. Bovine embryos become more resistant to adverse effects of maternal heat stress as pregnancy progresses; substantial resistance develops by d 3. This information may be useful in design of environmental modification systems that provide cooling at critical periods of gestation to enhance pregnancy rates during summer in hot climates.

Therapeutic application of gonadotrophin releasing hormone (GnRH) and its analogs was limited initially to the treatment of cystic follicular degeneration in cows. The role of GnRH was subsequently expanded to control of normal ovarian follicles for fertility enhancement in repeat breeder cows, and regulation of the dominant follicle in estrus synchronization programs as well as for protection of some slowly developing bovine conceptuses. Therapeutic use of GnRH is rare in small ruminants where its application is focused primarily on out-of-season breeding. In pigs GnRH is used in reproductive management for induction of estrus in prepubertal gilts and postpartum sows, as well as for increasing litter size. In horses GnRH is also used for breeding management specifically to schedule ovulation. GnRH seeks to replace human chorionic gonadotrophin because of the latter's propensity to cause antibody formation after repeated use. However, GnRH must be given repeatedly, at frequent intervals, by continuous infusion or by implant to be effective in the mare. Stimulation of follicular development and ovulation are dependent on the amount of GnRH given. While there are no current, practical clinical applications in dogs and cats, there is considerable interest in the chronic administration of GnRH as a contraceptive agent. Immunization against GnRH can also have a contraceptive effect in dogs.

Bovine interferon-alpha I1 (bIFN-alpha) may be useful for enhancing fertility in sheep and cattle because it has extensive sequence homology with ovine and bovine trophoblast protein-1 and, like those proteins, extends corpus luteum lifespan. To test the effectiveness of bIFN-alpha to enhance fertility, several experiments were performed in which inseminated heifers were given i.m. injections of bIFN-alpha approximately at the time of embryo-mediated signals that result in maintenance of the corpus luteum. In Exp. 1, heifers given 20 mg of bIFN-alpha daily from d 14 to 17 tended (P less than .07) to have lower pregnancy rates at d 110 to 112 of gestation (36/75; 48% vs 43/72; 60%). Similar results were obtained in Exp. 2 when heifers received a single injection of 40 mg of bIFN-alpha or placebo at d 13 after estrus; pregnancy rates at d 42 were 39/104 (38%) for bIFN-alpha and 47/98 (48%) for placebo. In Exp. 3, heifers were given gradually increasing doses of bIFN-alpha or placebo from d 11 to 19, because such a regimen had been shown to reduce the number of heifers experiencing hyperthermia after bIFN-alpha injection. Pregnancy rates were 42/95 (44%) for bIFN-alpha and 62/111 (56%) for placebo. Across all three experiments, pregnancy rates were lower (P less than .01) for heifers treated with bIFN-alpha (117/274; 43%) than for heifers treated with placebo (152/281; 54%). In conclusion, these results demonstrate that, under the administration systems used, bIFN-alpha does not increase pregnancy rate, but rather tends to reduce it.

The objective of Experiment I, replicated twice, was to evaluate whether fertility of estrus-synchronized dairy heifers could be improved by postinsemination progesterone supplementation. Estrous cycles were synchronized using two injections of prostaglandin (PG) F(2alpha) adiministered 11 days apart. Heifers displaying estrus were inseminated and assigned to control (n = 155) and treated (n = 159) groups. Treatment consisted of intravaginal insertion of controlled internal drug release (CIDR) devices for Days 7 to 13 (Day 0 = day of estrus). The conception rate for CIDR-treated heifers (57.9%) did not differ significantly from that of the controls (53.6%). The return-to-estrus rate and pattern of return estruses were not affected by treatment, but indicated that early embryonic mortality may have occurred in some of the heifers diagnosed nonpregnant. The objective of Experiment II was to evaluate if used CIDR devices were effective in resynchronizing returns to estrus in previously synchronized inseminated but nonpregnant and noninseminated heifers. Estrous cycles of dairy heifers of breeding age were synchronized with PGF(2alpha). Heifers displaying estrus were assigned to be inseminated (n = 117) or not inseminated (n = 35). All heifers were treated with 9-day used CIDR devices for Days 17 to 22 after synchronized estrus in order to resynchronize returns to estrus. Of the inseminated but nonpregnant heifers (n = 41), 78.1% were detected in estrus after CIDR removal (versus 94.3% of noninseminated heifers [n = 35]; P0.05) and 61.0% of the estruses occurred within 4 days of CIDR removal (versus 91.4% of noninseminated; P0.05). Estruses of synchronized inseminated nonpregnant heifers occurred over a longer period compared with those of noninseminated heifers (P0.025). The results indicate that response to the resynchronization protocol was altered by the outcome (early embryo death or failed fertilization) of the previous unsuccessful insemination, and support the hypothesis that delayed returns to estrus can be attributable to a pregnancy which was initiated but failed to establish itself. Such factors should be considered when evaluating responses of cattle to treatments designed to enhance fertility.

The effectiveness of various delivery vehicles and routes of administration in providing a sustained concentration of circulating relaxin in the cow was evaluated. Porcine relaxin (1 mg) was administered to cows during estrus as follows: 1) intramuscularly in 5% beeswax in sesame oil, 2) subcutaneously in beeswax/oil, 3) intracervically in K-Y gel, 4) intramuscularly in saline, and 5) intravenously in saline. Blood samples were taken over a 48-h period after administration, and plasma concentrations of relaxin were determined by radioimmunoassay. The half-life of relaxin from intravenous administration was determined to be 16.6 min. The beeswax/oil carrier diminished the release of relaxin, with plasma concentrations consistently lower than those seen with relaxin in a saline carrier. Relaxin, administered intramuscularly in saline, reached a peak concentration of 12.6 ± 1.2 ng/ml (Mean ± SEM) at 1.1 ± 0.2 h compared with only 2.8 ± 1.3 ng/ml at 1.8 ± 0.5 h when administered intramuscularly in oil. In addition, use of the beeswax/oil carrier unexpectedly acted like an adjuvant, resulting in the development of antibodies against porcine relaxin. After relaxin was deposited in the cervix, systemic concentrations of the hormone were marginally detectable (0.4 ± 0.2 ng/ml).

Conditioned culture media of ovine placental tissues or fetal skeletal muscle collected at Days 60, 100 and 140 of pregnancy were tested for their ability to inhibit in vitro lymphocyte proliferation. All conditioned cultures contained low-molecular-weight factors which suppressed [ 3 H]Tdr uptake by PHA-stimulated sheep lymphocytes. Non-dialyzable, inhibitory molecules were released only by placental tissues, with the greatest activity occurring for fetal cotyledonary placenta at Days 100 and 140 of gestation. Dialyzed conditioned culture medium of Day 100 cotyledonary placenta suppressed lymphocyte proliferation stimulated by PHA, PWM and MLR, but had no effect on ConA-stimulated cultures. Inhibitory actions were effective only during the first 24 h of lymphocyte activation and were abrogated with exogenous IL-2. Size-fractionation by gel filtration chromatography of the conditioned medium resolved the inhibitory activity into a peak of molecular weight > 4 × 10 6 Da and another peak with estimated molecular weight range of 46–162 kDa. Inhibitory action of the high molecular weight fraction was destroyed by sodium periodate oxidation whereas the lower molecular weight peak was sensitive to pronase. Presence of these immunosuppressive factors at the maternal-conceptus interface may contribute to down-regulation of local immune functions in the uterus and survival of the allogeneic conceptus.

Studies were performed to determine 1) cyclic changes in uterine immune function that could be responsible for the increased susceptibility of the diestrous uterus toinfection and 2) whether these changes are mediated by progesterone or estradiol-17#. The number and functional activity (cytochrome c reduction) of polymorphonuclear leukocytes (PMNL) collected from the uterus after oyster glycogen infusion was not statistically affected by stage of the cycle or steroid treatment. While not significant, the number of PMNL tended to be higher at diestrus than at estrus. The number of PMNL also tended to be higher for ovariectomized cows treated with progesterone than with estradiol-17/3 or vehicle. For cows given oyster glycogen, uterine IgG was not sitmificantly affected by treatment and uterine IgA was undetectable. For ovariectomized cows not given oyster glycogen, IgA was detectable in uterine flushings and was higher after 30 d of progesterone treatment than after 12 d of progesterone or vehicle. Additionally, the ability of uterine fluid from ovariectomized cows to inhibit mitogen-induced lymphocyte proliferation was greater (P < 0.01) for uterine fluid collected from cows treated with progesterone for 12 and 30 d than for cows treated with vehicle. It is concluded that alterations in migration of PMNL are unlikely to be the cause of decreased resistance of the uterus to infection during diestrus. Lowered resistance might, in part, be due to the secretion of uterine factors that inhibit lymphocyte function.

Uterine torsion and vaginal prolapse are accidents of gestation; this paper first reviews the predisposition, symptoms, diagnosis, and treatment of uterine torsion, and the predisposition, pathogenesis, symptoms, treatment, and prognosis of vaginal prolapse during late gestation in the cow. Other noninfectious complications of gestation are either fetal or placental in origin. Two relatively unusual fetal problems, mummification and maceration, are also discussed, followed by a comparison of two hydropic conditions (hydrallantois and hydramnios). Hydrops allantois, primarily a placental problem, occurs about nine times more frequently than hydrops amnii, which is a result of fetal anomalies. Abnormal offspring syndrome is a more recent phenomenon associated with the transfer of IVF or SCNT embryos. Finally, reference is made to teratology in general and a website of illustrations is cited.

Embryo transfer techniques in water buffalo were derived from those in cattle. However, the success rate is much lower in buffaloes, due to their inherent lower fertility and poor superovulatory response. The buffalo ovary has a smaller population of recruitable follicles at any given time than the ovary of the cow (89% fewer at birth). In addition, estrus detection is problematic. Progress in the field of embryo transfer in water buffalo has been slow, and is primarily due to a poor response to superovulation. The average yield of transferable embryos is less than one per superovulated donor. In vitro embryo production could considerably improve the efficacy and logistics of embryo production. The technique of Ovum Pick Up is superior to superovulation; it can yield more transferable embryos per donor on a monthly basis (2.0 versus 0.6). The feasibility of intergeneric embryo transfer between buffalo and cattle has been investigated. No pregnancy resulted after transfer of 13 buffalo embryos to synchronized Holstein heifers. Preliminary successes with nucleus transfer of Bubalus bubalis fetal and adult somatic nuclei into enucleated bovine oocytes and subsequent development to the blastocyst stage have been reported.

Fertility in water buffalo (Bubalus bubalis) is considerably lower than that in cattle (Bos taurus and Bos indicus). Poor breeding efficiency is attributed to late onset of puberty, seasonality, poor estrus expression, and long calving intervals. Accurate estrus detection is a prerequisite for efficient reproductive management. Established reproductive management techniques in cattle can be successfully applied to water buffalo because of the similarities in the anatomy, physiology, and endocrinology of reproduction between the two genera.

On December 19, 2005, 14 in vitro-fertilized water buffalo (Bubalus bubalis) embryos, which had been cryopreserved by vitrification, were thawed and transferred into B. bubalis recipients in California. The embryos had been produced in Italy, following transvaginal oocyte pickup (TVOPU), with subsequent in vitro maturation, insemination, and culture. This case study relates our experience in meeting the regulatory criteria, established by the Animal Import/Export Office of the USDA-Animal and Plant Health Inspection Service (USDA-APHIS), in order to successfully import these embryos into the USA.

J. Block, MS, R. M. Rivera, PhD, F. D. Jousan, MS, F. T. Silvestre, BS, F. F. Paula-Lopes, PhD, O. M. Ocon, BS, H. Rosson, MAg, T. R. Bilby, MS, P. J. Hansen, PhD, Department of Animal Sciences, M. Drost, DVM, Department of Large Animal Clinical Sciences, University of Florida, Gainesville, FL 32611, USA R. L. Monson, MS, J. J. Rutledge, PhD, Department of Animal Sciences, University of Wisconsin-Madison, Madison, WI 53706, USA C. R. Looney, PhD, Ovagenix, Bryan, TX 77868, USA

Glycogen storage disease II is an inherited progressive muscular disease in which the lack of functional acid 1-4 alpha-glucosidase results in the accumulation of lysosomal glycogen. In the present study, we examine the effect of these non-contractile inclusions on the mechanical performance of skeletal muscle. To this end, force developed in an isometrically contracting slice of a muscle was calculated with a finite element model. Force was calculated at several inclusion densities and distributions and compared to muscle lacking inclusions. Furthermore, ankle dorsal flexor torque was measured in situ of alpha-glucosidase null mice of 6 months of age and unaffected litter mates as was inclusion density in the dorsal flexor muscles. The calculated force loss was shown to be almost exclusively dependent on the inclusion density and less on the type of inclusion distribution. The force loss predicted by the model (6%) on the basis of measured inclusion density (3.3%) corresponded to the loss in mass-normalized strength in these mice measured in situ (7%). Therefore, we conclude that the mechanical interaction between the non-contractile inclusions and the nearby myofibrils is a key factor in the loss of force per unit muscle mass during early stages of GSD II in mice. As glycogen accumulation reaches higher levels in humans, it is highly probable that the impact of this mechanical interaction is even more severe in human skeletal muscle.

Lysosomal dysfunction in muscle with special reference to glycogen storage disease type II.Hesselink RP, Wagenmakers AJ, Drost MR, Van der Vusse GJ.Department of Movement Sciences, Cardiovascular Research Institute Maastricht, Maastricht University, P.O. Box 616, 6200 MD Maastricht, The Netherlands. Reinout.Hesselink@BW.Unimass.nlThe importance of proper lysosomal activity in cell and tissue homeostasis is underlined by "experiments of nature", i.e. genetic defects in one of the at least 40 lysosomal enzymes/proteins present in the human cell. The complete lack of 1-4 alpha-glucosidase (glycogen storage disease type II (GSD II) or Pompe disease) is life-threatening. Patients suffering from GSD II commonly die before the age of 2 years because of cardiorespiratory insufficiency. Striated muscle cells appear to be particularly vulnerable in GSD II. The high cytoplasmic glycogen content in muscle cells most likely gives rise to a high rate of glycogen engulfment by the lysosomes. The polysaccharides become subsequently trapped in these organelles when 1-4 alpha-glucosidase activity is absent. During the course of the disease, muscle wasting occurs. It is hypothesised that the gradual loss of muscle mass is caused by a combination of disuse atrophy and lipofuscine-mediated apoptosis of myocytes. Moreover, we hypothesise that in the remaining skeletal muscle cells, longitudinal transmission of force is hampered by swollen lysosomes, clustering of non-contractile material and focal regions with degraded contractile proteins, which results in muscle weakness

The production of embryos by superovulation is often reduced in periods of heat stress. The associated reduction in the number of transferable embryos is due to reduced superovulatory response, lower fertilization rate, and reduced embryo quality. There are also reports that success of in vitro fertilization procedures is reduced during warm periods of the year. Heat stress can compromise the reproductive events required for embryo production by decreasing expression of estrus behavior, altering follicular development, compromising oocyte competence, and inhibiting embryonic development. While preventing effects of heat stress can be difficult, several strategies exist to improve embryo production during heat stress. Among these strategies are changing animal housing to reduce the magnitude of heat stress, utilization of cows with increased resistance to heat stress (i.e., cows with lower milk yield or from thermally-adapted breeds), and manipulation of physiological and cellular function to overcome deleterious consequences of heat stress. Effects of heat stress on estrus behavior can be mitigated by use of estrus detection aids or utilization of ovulation synchronization treatments to allow timed embryo transfer. There is some evidence that embryonic survival can be improved by antioxidant administration and that pharmacological treatments can be developed that reduce the degree of hyperthermia experienced by cows exposed to heat stress.

Objectives were to examine how the conceptus and recombinant bovine interferon-tau (rbIFN-tau) regulate intracellular components of the PGF(2a) synthetic pathway and to determine if arachidonic acid (AA) is limiting in endometrial tissue of pregnant cows. In Experiment 1, uteri were collected from either cyclic or pregnant dairy cows on Day 17 post-estrus. Intercaruncular explants were dissected and incubated for 60 min to quantify PGF(2a) production in response to oxytocin (10(-6) M), A23187 (10(-5) M), melittin (10(-5) M), and phorbol 12, 13 dibutyrate (PDBu, 10(-6) M). Additional explants from the same cows were incubated for 24 h with and without AA. Oxytocin and A23187 did not stimulate PGF(2a) in explants from either cyclic or pregnant cows. Both PDBu, melittin, and A23187 + melittin stimulated PGF(2a) production in explants of cyclic cows, but not in explants of pregnant cows. The addition of AA to explant cultures for 24 hr did not increase PGF(2a) production during a subsequent 60-min incubation. In Experiment 2, explants were collected from cows that received intrauterine infusions of either BSA (1.9 mg/1.2 ml) or rbIFN-tau (0.2 mg rbIFN-tau + 1.7 mg BSA/1.2 ml) twice a day from Days 14 to 17 of the estrous cycle. Treatments of rbIFN-tau attenuated PGF(2a) secretion induced by in vitro PDBu and A23187 treatments. However, rbIFN-tau treatment in vivo had no effect on the in vitro induction of PGF(2a) secretion by melittin. IFN-tau may regulate the PGF(2a) synthetic pathway by reducing activity of PKC or PKC mediated events.

The objective was to compare conception rates to embryo transfer relative to AI, during summer heat stress, in lactating dairy cows. Holstein cows (n = 180; 50 to 120 d postpartum) were allocated randomly to 1 of 3 groups: artificial insemination (AI, n = 84), embryo transfer using either embryos collected from superovulated donors (ET-DON, n = 48), or embryos produced in vitro (ET-IVF, n = 48). Embryos from superovulated donors were frozen in 10% glycerol and were rehydrated in a 3-step procedure, in decreasing concentrations of glycerol in a sucrose medium before transfer. Embryos produced in vitro were frozen in 1.5 M ethylene glycol, thawed and transferred without rehydration. Blood samples were collected from AI and ET recipients on Days 0, 7 and 22 for measurement of progesterone in plasma. Conception rate was estimated for the three groups at Day 22 (progesterone1 ng/mL) and confirmed at Day 42 by palpation per rectum. Conception rate estimates at Day 22 did not differ among groups (AI, 60.7%; ET-DON, 60.4%; ET-IVF, 54.2%), but conception rates at Day 42 differed (AI, 21.4%; ET-DON, 35.4%; ET-IVF, 18.8%; AI versus ET: P0.10 and ET-DON versus ET-IVF: P0.05). In cows considered pregnant at 22 d but diagnosed open at 42 d, the interestrous intervals were 28.8 +/- 2.2, 35.2 +/- 3.5 and 31.6 +/- 2.9 d, respectively, for AI, ET-DON and ET-IVF groups. Transfer of embryos collected from nonheat-stressed superovulated donors significantly increased conception rates in heat stressed dairy cattle. However, transfer of IVF-derived embryos had no advantage over AI. Where appropriate mechanisms are in place to attenuate the effects of heat stress, embryo transfer using frozen-thawed donor embryos increases conception rates.

Our objective was to determine whether pregnancy rates in heat-stressed dairy cattle could be enhanced by timed embryo transfer of fresh (nonfrozen) or frozen-thawed in vitro-derived embryos compared to timed insemination. Ovulation in Holstein cows was synchronized by a GnRH injection followed 7 d later by PGF2 alpha and a second treatment with GnRH 48 h later. Control cows (n = 129) were inseminated 16 h (d 0) after the second GnRH injection. On d 7, a fresh (n = 133) or frozen-thawed (n = 142) in vitro-derived embryo was transferred to cows assigned for timed embryo transfer after categorizing the corpus luteum by palpation per rectum as 3 (excellent), 2 (good or fair), 1 (poor), and 0 (nonpalpable). Response to the synchronization treatment, determined by plasma progesterone concentration (ng/ml)or = 1.5 on d 0 andor = 2.0 on d 7, was 76.2%. Mean plasma progesterone concentration on d 7 increased as the quality of corpus luteum improved from category 0 to 3. Concentrations of progesterone in plasma were elevated (or = 2.0 ng/ml) at 21 d in 64.7 (fresh embryo), 40.3 (frozen embryo), and 41.4 +/- 0.1% (timed insemination) of cows, respectively. Cows that received a fresh embryo had a greater pregnancy rate at 45 to 52 d than did cows that received a frozen-thawed embryo or timed insemination (14.34.8, 4.9 +/- 2.3%). Body condition (d 0) of cows influenced the pregnancy rate and plasma progesterone concentrations. In summary, timed embryo transfer with fresh in vitro-produced embryos in heat-stressed dairy cattle improved pregnancy rate relative to timed insemination.

Endometrial oxytocin receptors and total production of PGF by endometrial epithelial cells were measured in 10 cyclic cows after intrauterine injections of recombinant bovine interferon-tau plus BSA or BSA alone. Cows received twice daily injections (via intrauterine catheters) of 200 micrograms of recombinant bovine interferon-tau plus 1.3 mg of BSA (n = 5) or 1.5 mg of BSA (n = 5) from d 14 to 17 after estrus. On d 17, the reproductive tracts of each cow was removed at slaughter, and endometrial epithelial cells were cultured with 0, 2, or 50 ng/ml of recombinant bovine interferon-tau. After 24 h, oxytocin (2 x 10(-7) M) was added to one-half of the culture wells, and the medium was sampled at 0, 30, and 90 min for analysis of total PGF (PGF plus 13, 14-dihydro-15-keto-PGF2 alpha). In vivo treatment with recombinant bovine interferon-tau + BSA reduced total secretion of PGF in culture (1.49 +/- 0.06 vs. 2.80 +/- 0.07 ng/micrograms of DNA), but did not block the oxytocin-induced stimulation in total secretion of PGF. In vitro treatment of cells with recombinant-bovine interferon-tau did not decrease basal secretion of total PGF. Oxytocin receptor binding at d 17 was low in both treatments but slightly attenuated in the group treated with recombinant bovine interferon-tau.

Three experiments were conducted to evaluate a synchronization protocol with AI at a predetermined time. In Exp. 1, 169 dairy heifers were assigned randomly to two groups: 1) timed AI (TAI), consisting of GnRH agonist injection (d 0, 1700), PGF2 alpha injection (d 7, 1700), GnRH agonist injection (d 8, 1700), and AI (d 9, 0800); and 2) AI at estrus (AIE), consisting of GnRH agonist injection (d 0, 1700), PGF2 alpha injection (d 7, 1700), and AI at detected estrus. Pregnancy rate was 25.8% for TAI (n = 89) compared with 48.7% for AIE (n = 80; P < .001). Experiment 2 was comparable to Exp. 1, but the second GnRH agonist injection in TAI was given 48 h after injection of PGF2 alpha. Heifers in TAI (n = 187) were inseminated at detected estrus if estrus occurred within 39 h after administration of PGF2 alpha (n = 47). Pregnancy rates were 45.5% for TAI and 48.0% for AIE (n = 177). Conception rate was reduced for TAI (45.5 [85/187] < 61.2% [85/139]; P < .005). In Exp. 3, the second injection of GnRH agonist, given at 48 h after injection of PGF2 alpha, was replaced with hCG (3,000 IU, i.m.). No differences in pregnancy rate were detected for TAI (52.9% [54/102]) vs AIE (56.1% [55/98]). Conception rate was reduced for TAI (52.9 [54/102] < 72.3% [55/76]; P < .005). Delaying the second GnRH agonist injection by 24 h improved pregnancy rate, but replacing the second injection of GnRH agonist with an injection of hCG did not prevent a reduction in conception rate.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2014-02-26T17:07:20Z No. of bitstreams: 0 Made available in DSpace on 2014-02-26T17:07:20Z (GMT). No. of bitstreams: 0 Previous issue date: 1996-05-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T15:29:26Z No. of bitstreams: 0 Made available in DSpace on 2014-05-20T15:29:26Z (GMT). No. of bitstreams: 0 Previous issue date: 1996-05-01 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). UNIV FLORIDA,DEPT DAIRY & POULTRY SCI,GAINESVILLE,FL 32611 UNIV FLORIDA,DEPT LARGE ANIM CLIN SCI,GAINESVILLE,FL 32611 UNESP,DEPT PHARMACOL,BR-18600 SAO PAULO,BRAZIL S FLORIDA CTR FERTIL,MIAMI,FL 33143 UNESP,DEPT PHARMACOL,BR-18600 SAO PAULO,BRAZIL

An experiment was conducted to evaluate the role of the dominant follicle (DF) of the first wave in regulating follicular and ovulatory responses and embryonic yield to a superovulation regime with FSH-P. Twenty normally cycling Holstein-Freisian heifers (n = 20) were synchronized with GnRH and pgf(2alpha) and randomly assigned to a control or a treated group (n = 10 each). Treated heifers had the first wave dominant follicle removed via transvaginal, ultrasound-guided aspiration on Day 6 after a synchronized estrus. All heifers received a total of 32 mg FSH-P given in decreasing doses at 12 h intervals from Day 8 to Day 11 plus two injections of pgf(2alpha) (35 mg and 20 mg, respectively) on Day 10. Heifers were inseminated at 6 h and 16 h after onset of estrus. Follicular dynamics were examined daily by transrectal ultrasonography from Day 4 to estrus, once following ovulation, and at the time of embryo collection on Day 7. Blood samples were collected daily during the superovulatory treatment and at embryo collection. Follicles were classified as: small,/= 5 mm; medium, 6-9 mm; or large,/= 10 mm. Aspiration of the dominant follicle was associated with an immediate decrease in large follicles, and a linear rate increase in small follicles from Day 4 to Day 8 just prior to the FSH-P injections, (treatmentcontrol: +0.33 vs. -0.22, number of small follicles per day; P0.10). During FSH-P injections, the increase in number of medium follicles was greater (P0.01) for treatment on Day 9-11 (treatmentcontrol: Day 9, 3.21.8; Day 10, 9.24.7; Day 11, 13.18.3; +/- 0.56). Number of large follicles was greater in treatment at Day 11 (5.121.4 +/-0.21; P0.01). Mean number of induced ovulatory follicles (difference between number of follicles at estrus and Day 2 after estrus) was greater in treatment (13.46.3 +/- 1.82; P0.01). Plasma estradiol at Day 11 during FSH-P treatment was greater in treatment (32.515.8 +/- 2.6; P0.01). Plasma progesterone at embryo flushing (Day 7 after ovulation) was greater in treatment (7.44.9; P0.02); technical difficulties at embryo recovery reduced sensitivity of embryonic measurements. No changes in the distribution of unfertilized oocytes and embryo developmental stages were detected between control and treatment groups. Presence of dominant follicle of the first wave inhibited intraovarian follicular responses to exogenous FSH.

In 3 separate trials at 2 locations, dairy heifers (n = 396) were treated with a Controlled Internal Drug Release (CIDR) progesterone device for 9 d. On Day 7 of CIDR treatment, all heifers were injected with PGF(2alpha). Synchronized estruses were detected using a tailpaint and chalk (TPC) scoring system. An animal's tailhead was painted at device insertion, and this strip was covered with a contrasting color of chalk at device removal. Over all trials, 85.1% of the heifers were detected in estrus and were inseminated at 48 or 72 hours after CIDR removal. These synchronized and inseminated heifers were divided into the following treatment groups: 1) untreated controls, receiving no further treatment (n = 138); 2) post-insemination progesterone supplementation with a new (n = 59) or used (n = 29) CIDR device for Days 1 to 8 or 2 to 9, respectively, following insemination; or 3) resynchronization of return to service with a used CIDR device for Days 17 to 22 after insemination (n = 112). The pregnancy rate to first insemination in the control and resynchronized groups (Groups 1 and 3) was 46.4%, but decreased to 18.2% with the post-insemination progesterone supplementation. Resynchronization of returns to service (estrus detected 1 to 4 d following removal of second CIDR) occurred in 58.9% of all nonpregnant heifers in Group 3. In summary, CIDR devices used in conjunction with PGF(2alpha) effectively synchronize estrus in dairy heifers. Progesterone supplementation within 2 d of first insemination for 7 d suppressed fertility. Used CIDR devices inserted for Days 17 to 22 after first insemination resynchronized heifers not pregnant to first insemination.

We examined the effect of intramuscular injections of recombinant bovine interferon-tau on basal body temperature and plasma concentrations of progesterone in cows. Six mature, nonlactating Holstein cows were synchronized for estrus and assigned randomly to a replicated 3 x 3 Latin square design to receive 0, 1, or 5 mg of recombinant bovine interferon-tau intramuscularly on d 9, 12, or 15 of the estrous cycle. Rectal temperatures were measured, and blood was collected just prior to injection, at hourly intervals for 8 h, and at 10 and 12 h after injection. Treatment with 1 and 5 mg of recombinant bovine interferon-tau caused a rise in body temperature that peaked at 6 h (39.5 +/- .13 degrees C) and 4 h (40.3 +/- .13 degrees C), respectively. Analysis of homogeneity of regression for response curves indicated that 5 mg of recombinant bovine interferon-tau decreased plasma progesterone concentrations. This effect was not detected for the 1-mg dose of recombinant bovine interferon-tau. Intramuscular administration of 5 mg of recombinant bovine interferon-tau caused a hyperthermic response that was temporally associated with the decrease in plasma progesterone. Intramuscular administration of 1 mg of recombinant bovine interferon-tau caused a slight rise in temperature occurring 4 h later than that caused by 5 mg of recombinant bovine interferon-tau, and no decrease in plasma progesterone was detected. Recombinant bovine interferon-tau, administered at a 1-mg dose, warrants further study to determine whether it will alter survival rate of cattle embryos.

Nonlactating Holstein cows (n = 12) were assigned randomly to one of two diets (approximately 30% concentrate) in a crossover design. Urea and soybean meal were used to increase CP content of diet from 12.3 to 27.4%. Mean concentrations of plasma urea were 9.8 and 21.3 mg/dl for respective diets. After 35 d on diets, cows were brought to a synchronized estrus (progestin implant plus PGF2 alpha injection). From d 10.5 to 14.5 following estrus, cows received i.m. injections of decreasing doses of FSH to initiate superovulation. On d 3 of FSH treatment, PGF2 alpha was injected with FSH. Cows were bred twice by AI when detected in estrus. Embryos were collected nonsurgically 6.5 d after breeding, and quality was assessed using visual, microscopic, and staining techniques. Development of follicles throughout this period was monitored using ultrasonography. Using ultrasonography, no differences were detected in the number or percentages of preovulatory, anovulatory, and ovulatory follicles induced during superovulation. Number of recovered embryos averaged eight per cow. Numbers and percentages of normal embryos, abnormal or retarded embryos, and unfertilized ova were similar between diets, as were the numbers and percentages of transferable and nontransferable embryos. Excess intake of CP failed to affect the growth or number of ovarian follicles and health and number of embryos of energy adequate, nonlactating dairy cows.

Bovine embryos, recovered from the uterus in vivo or derived from in vitro matured and in vitro fertilized oocytes, were investigated for the presence of the developmentally regulated mouse antigen TEC-3 by indirect immunofluorescence. During preimplantation embryo development TEC-3 is expressed on bovine morulae and blastocysts. It is absent from unfertilized and fertilized oocytes, and from all stages before the 32-cell stage. The finding that TEC-3 is not expressed before the onset of embryonic transcription, which occurs at the eight-cell stage in the bovine, but only when the embryonic genome is active, makes it a potential marker for studying nuclear reprogramming after nuclear transfer. Nuclear transfer embryos were made by electrical fusion of blastomeres from morulae derived in vivo with enucleated metaphase II oocytes. Indirect immunofluorescence with the TEC-03 antibody showed that the TEC-3 antigen, present on blastomeres of the morula stage embryo, disappeared after fusion and was expressed again when the nuclear transfer embryos developed to the morula and blastocyst stage. These data suggest that the bovine embryonic nucleus may be able to revert to the equivalent of an earlier developmental stage when transferred to ooplasm, and is then capable of following the normal developmental program. © 1994 Wiley-Liss, Inc.

The effects of concentration of progesterone in plasma on development and fertility of the first wave dominant follicle were studied in cattle. To identify a source of exogenous progesterone that would permit extension of the first wave dominant follicle, nonlactating Holstein cows (n = 6) received on day 8 of two successive oestrous cycles an injection of PGF2 alpha (25 mg) and a new (1.9 g of progesterone (Period 1)) or used (approximately 1.2 g of progesterone (Period 2)) CIDR-B device that was removed on day 17. Control cows (n = 6) received a new CIDR-B device on day 8 that was removed on day 17 and a PGF2 alpha injection (25 mg) on day 17. Ultrasonography and collection of blood samples were performed on alternate days throughout the experiment. Plasma concentrations of progesterone and oestradiol were different between treatments (P < 0.0001 and P < 0.05, respectively). The dominant follicle was maintained until day 17 and ovulated upon removal of the intravaginal device in 1 of 6, 6 of 6 and 0 of 6 in new CIDR-B, used CIDR-B and control groups, respectively (P < 0.01). The preovulatory dominant follicles were 14.2 +/- 1.6 mm, 20 +/- 1.3 mm and 10 +/- 1.3 mm, respectively (P < 0.001) on day 17. There were fewer 5-9 mm follicles in cows having a persistent dominant follicle (P < 0.01). The interval to onset of oestrus was negatively correlated with size of the dominant follicle on day 17 (P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Postpartum ovarian activity, uterine involution and plasma concentrations of calcium and 15-keto-13, 14 dihydro-prostaglandin F2alpha (PGFM) were assessed in dairy cows with retained fetal membranes (n=10) and milk fever (n=10) at parturition. In addition, calcium and PGFM were evaluated in dairy cows affected with uterine prolapse (n=10) and pyometra (n=14). Cows with retained fetal membrane averaged 24.2+/-3.7 d until their first postpartum ovulation, while controls averaged 29.0+/-3.7 d (P0.10). In cows with retained fetal membranes, the difference in follicular activity between the contralateral and ipsilateral ovaries in relation to the previously gravid uterine horn was appreciably greater post partum when compared with that of the controls. Cows with milk fever had an average of 30.8+/-3.1 d until their first postpartum ovulation, while control cows had an average of 20.4+/-3.3 d (P0.05). The mean diameter of the uterine horns in cows with milk fever was greater (P0.05) compared with that of the controls between Days 15-32 post partum. Concentrations of plasma calcium were lower in cows with retained fetal membranes within 24 h after parturition and during the first week post partum than in the controls (6.27+/-0.18 vs 7.40+/-0.18 mg/100ml, P0.05). Concentration of calcium was lower (P0.05) in cows with milk fever on Day 1 prior to treatment (4.68+/-0.405.8+/-0.45 mg/100ml) than in control cows; however, the calcium (Ca) level was not different during the subsequent 7 d post partum after treatment. Cows with uterine prolapse had lower concentrations of Ca during the first 7 d post partum than the controls (6.10+/-0.15 vs 7.33+/-0.12mg/100ml; P0.01). Cows with pyometra had higher (P0.05) concentrations of plasma PGFM than the controls (208.+/-13.2138.1+/-15.2).

To determine if deleterious effects of heat shock on embryos could be reduced in vitro by glutathione or taurine, morulae from superovulated cows were placed in modified Hams-F10 medium supplemented with 50 nM glutathione (GSH), 50 mM taurine or neither. Morulae were incubated for 2 hours at 38.5 degrees C, then at 42.0 degrees C (heat shock) or 38.5 degrees C for 2 hours and followed by incubation at 38.5 degrees C for 20 hours. Neither GSH nor taurine enhanced viability or blastocyst development at 38.5 degrees C. At 42.0 degrees C, however, GSH and taurine increased (P less than 0.02) viability (73%, 41% and 26% live for GSH, taurine and control); GSH increased (P less than 0.05) blastocyst development (55% for GSH vs. 30% for control). In conclusion, partial thermoprotection of bovine embryos from heat shock can be achieved in vitro by administration of GSH. Taurine is only slightly effective.

Culture medium conditioned with concanavalin A-stimulated mononuclear leukocytes was tested for its ability to stimulate in vitro proliferation of bovine placental cells. The crude preparation of cytokines caused a dose-dependent increase in [3H]thymidine uptake into cells obtained by trypsinization of fresh bovine placentae and placental cell lines established from cellular outgrowths of long-term bovine placental cultures, but had no effect on growth of 3T3 fibroblasts. Growth of trypsinized placental cells was not enhanced by culture in the presence of interleukin-2, interferon-beta 2, interferon-gamma, or tumor necrosis factor-alpha. These results corroborate those of murine studies, suggesting a growth-promoting role for cytokines released into the maternal-fetal interface.

The effect of follicular and/or endocrine environments on superovulatory response was tested. Eighteen nonlactating Holstein cows were superovulated with 32 mg FSH-P given in decreasing doses at 12-h intervals plus two injections of prostaglandin F2-alpha (25 mg each) on the third day of treatment. Cows were assigned randomly to treatments: T1, superovulatory treatment initiated on estrous cycle Day 10.5; T2, CIDR (intravaginal device containing 1.9 g of progesterone) inserted from Days 3 to 9 and superovulation initiated on Day 6.5; T3, identical to T2 but Buserelin (GnRH agonist) was injected (8 microg, i.m.) on Day 3 at the time of CIDR insertion. Embryos were recovered on Day 7 after the superovulatory estrus. Cows were examined daily by ultrasonography and blood was collected for progesterone and estradiol determinations. Mean diameter of the dominant follicle (frequency of first-wave dominant follicle) at the beginning of FSH injections was 13.7 mm (4/6), 11.2 mm (6/6) and 8.7 mm (6/6) (P0.01) for T1, T2 and T3, respectively. Following initiation of superovulation, follicles moved into larger follicle classes (Class I,3 mm; Class II, 3 to 4 mm; Class III, 5 to 9 mm; Class IV9 mm) earliest in T1 (P0.01). Cumulative follicular diameter and plasma concentrations of estradiol at Day 4 of superovulation were higher (P0.01) in T1 (200 mm, 82 pg/ml) compared with T2 (123 mm, 24 pg/ml) and T3 (130 mm, 18 pg/ml). Proportion of cows in estrus prior to 12 h vs 12 to 24 h differed (P0.05) between groups (T1: 5 vs 1; T2: 2 vs 4; T3: 1 vs 5). Mean number of follicles on the last day of superovulation treatment, number of CL and number of embryos plus unfertilized ova recovered were 17.5, 12.2 and 13.3; 13.8, 10 and 8.2 (P0.1) and 8.7, 4.5 and 2.3 (P0.05) for T1, T2 and T3, respectively. The developmental stage of the dominant follicle was associated with not only the number of ovulations, but also the size and periestrous concentrations of plasma estradiol associated with the recruited follicles.

Immunohistochemistry was utilized to determine expression of the major histocompatibility complex (MHC) antigens on Day 8-9 hatched blastocysts and fetal membranes of mid- to late gestation cows and to examine the pattern of leucocytic infiltration into the gravid uterus. Hatched blastocysts were weakly positive for MHC class I antigens. In the mature placenta, chorioallantoic membranes in the interplacentomal area showed positive immunostaining for class I antigens on the chorionic epithelium but had no staining for class II antigens. There was an accumulation of lymphoid cells expressing class II antigens directly beneath the luminal epithelium of the endometrium. In addition, cells staining for leucocyte common antigen were present both within and beneath the luminal epithelium. Some cells positive for class II and leucocyte common antigen (CD45) were also associated with uterine glands. In the placentomes, class I antigens were expressed only on maternal caruncular septa. Fetal cotyledonary villi had no detectable immunostaining for class I and II antigens. No distinct pattern of leucocyte infiltration in the maternal caruncular tissue was observed; the caruncular septa contained some cells that were labelled for CD45 and a few class II-positive cells around blood vessels. The results indicate that the fetal placenta of the cow expresses MHC class I antigens in a regionally defined manner and there is a differential accumulation of lymphoid cells in the uterus.

An experiment was conducted to determine whether pregnancy rates following the transfer of in vitro-produced embryos to heat-stressed cows could be improved by 1) culturing embryos in the presence of IGF-I and 2) treating recipients with GnRH. Lactating Holstein cows (n = 260) were synchronized using a timed ovulation protocol. Embryos were produced in vitro and cultured with or without 100 ng/mL of IGF-I. On d 7 after anticipated ovulation (d 0), a single embryo was transferred to all recipients with a palpable corpus luteum (n = 210). A subset of recipients (n = 164) was injected with either GnRH or placebo on d 11. Plasma progesterone concentrations on d 0 and 7 were used to determine the synchrony of recipients. Pregnancy was diagnosed at d 53 and 81 by rectal palpation. Among all recipients, transfer of IGF-I-treated embryos increased pregnancy rate at d 53 (P0.05) and tended to increase pregnancy rate at d 81 (P0.06). Calving rate also tended to be higher for recipients that received IGF-I-treated embryos (P0.07). Among the subset of synchronized recipients (n = 190), pregnancy rate at d 53 and d 81 and calving rate were higher (P0.05) for IGF-I-treated embryos. The GnRH tended to increase pregnancy rate at d 53 for all recipients (P0.08) and the subset of synchronized recipients (P0.10). There were no effects of GnRH (P0.10) for pregnancy rate at d 81 and calving rate. The overall proportion of male calves was 64.3%. There was no effect (P0.10) of embryo treatment or GnRH on the birth weight or sex ratio of calves. Results of this experiment indicate that treatment of embryos with IGF-I can improve pregnancy and calving rates following transfer of in vitro-produced embryos. Further research is necessary to determine whether the treatment of recipients with GnRH is a practical approach to increase pregnancy rates following in vitro embryo transfer.