Differential expression, during the estrous cycle and pre- and postimplantation conceptus development, of messenger ribonucleic acids encoding components of the pig uterine insulin-like growth factor system.

The temporal patterns of endometrial expression for mRNAs encoding insulin-like growth factor-I (IGF-I), IGF-II, IGF-binding protein-2 (IGFBP-2), and the type I IGF receptor (IGF-IR) were elucidated in cyclic and pregnant pigs. Peak levels of IGF-I mRNAs occurred on day 12 in cyclic and early pregnant gilts, while IGFBP-2 mRNA levels were lowest on day 10. Pregnant gilt endometrium had higher levels of both RNA classes than the corresponding cyclic endometrium. IGF-II and IGF-IR mRNAs remained low during this period. In pregnant pig endometrium and rat uterus, levels of IGF-I mRNA decreased, while those of IGF-II and IGFBP-2 mRNAs increased with stage of pregnancy. Decreased endometrial production of IGF-I mRNA during pregnancy paralleled that in the myometrium. IGF-II mRNA tissue abundance was placenta greater than endometrium greater than myometrium. In contrast, IGFBP-2 mRNA levels were higher in endometrium than in placenta and myometrium. Endometrial expression of IGF-II mRNAs was limited to surface and glandular epithelial cells; epithelial and stromal cells expressed IGFBP-2 mRNAs at comparable levels. Expression of IGF-IR mRNAs was low and did not change with pregnancy. The endometria of two breeds of pigs that exhibit different levels of prolificacy were also examined for IGF mRNAs. On day 12, endometrium from the Large White breed with high conceptus mortality had higher levels of IGF-II and IGFBP-2 mRNAs than did endometrium from the Meishan breed with low conceptus mortality. Expression of IGF-I mRNAs was higher in endometria of Meishan than Large White gilts on day 12. The differential expression of IGF mRNAs with stage of gestation and the correlation of relative ratios of IGF mRNAs with prolificacy during the critical period of maternal recognition of pregnancy suggest an important role(s) for IGFs in conceptus and fetal development.

Growth kinetics of the granulosa cell population in ovarian follicles: an approach by mathematical modelling.

This paper describes, from a mathematical viewpoint, the cellular changes in the granulosa of ovarian follicles during their terminal development. A dynamic model takes into account the processes of (1) cell division, (2) exit from the cell cycle towards differentiation, and (3) apoptotic cell death. Proliferative cells leave the cycle in an irreversible way. The risk of entering apoptosis applies to non-cycling cells. Changes in the cell numbers and in the growth fraction are derived from differential equations. The transitions between the different cell states are ruled by time-dependent rates. Numerical applications of the model concern ovulating and degenerating ovarian follicles in the ewe. The main feature of the ovulating case is the progressive exhaustion of the proliferating compartment for the benefit of the non-cycling cells. From an initial mainly proliferative state the granulosa progressively switches to a highly differentiated state, so that the growth fraction continuously decreases. In the atretic cases, the pattern of changes in the total viable cell number is influenced by the follicular age at the onset of the apoptotic process and by the intensity of the cell death rate. As apoptosis affects the non-cycling cells, the growth fraction is no longer strictly decreasing. The sensitivity of the model to the parameters is studied in a more general framework than the granulosa cell population.

A highly sensitive microtitre plate enzyme immunoassay for oestradiol-17 beta.

A specific and sensitive solid-phase microtitre plate enzyme-linked immunosorbent assay for oestradiol-17 beta (E2) is described. After coating with an IgG anti-E2 fraction, we used E2-6-carboxymethyl-oxime-beta-galactosidase in a competitive binding assay and revealed the bound activity with a fluorogenic substrate. Two methods for the competitive binding assay were tested: (1) a classical one (method A) defined as a 'two-step competition' because the E2 sample was first incubated alone, and then E2-beta-galactosidase conjugate was added; (2) and a new one (method B) also performed in two steps but in which the E2 sample was evaporated to dryness. The detection limit of method A was 100 pg/ml (9 pg/well). Method B was more sensitive since 1 pg/ml (0.3 pg/well) was statistically different from 0 pg/ml. Specificity was equivalent with both methods while precision was better in B. Thus, this new method may be able to measure very low levels of oestradiol-17 beta in, for example, the blood of domestic mammals.

Nuclear testosterone receptors in the ovine testis.

Nuclear [3H] testosterone-receptor complexes were demonstrated in hypophysectomized ram testis after in vitro direct labelling. The nuclear binding was maximal after a 45 min incubation of the tissue. The receptors are extractable by 0.4 M KC1 or NaSCN with a 25-30% efficiency. They migrate towards the anodic region during electrophoresis on agar gel. Nuclear androgen receptors were characterized in intact lamb testis by a testosterone exchange assay. After precipitation by protamine sulphate, the receptors were labelled with [3H]testosterone during a 12 h incubation at 4 degrees C. The exchange activity was linear between 0.1 and 0.9 mg of DNA per ml of incubation buffer. The receptors bind testosterone with a limited capacity (40-180 fmoles per mg DNA) and a dissociation constant Kd of 2 x 10(-9) M. Their relative affinities for steroids are dihydrotestosterone greater than testosterone greater than estradiol greater than progesterone greater than 5 alpha-androstanediol greater than cyproterone acute greater than R5020.

Characterization of a cytoplasmic androgen receptor in the ram testis.

An androgen receptor has been characterized in the cytosol fraction of testes from hypophysectomized adult rams after in vitro labelling with [3H]testosterone. It can be distinguished from the testicular androgen-binding protein (ABP) and from the plasma 5 alpha-dihydrotestosterone-binding protein by electrophoresis on 3.25% acrylamide gels (Rx = 0.5) and on agar gels (anodic migration). It sediments in the 4S region in sucrose gradient containing 0.4 M KCl. Its complex with testosterone dissociates very slowly (t 1/2 = 29 h at 0 degrees C), and is destroyed by heating at 50 degrees C for 30 min and by pronase. Its relative affinities for steroids are 5 alpha-DHT greater than T greater than 5 alpha-androstanediols greater than cyproterone acetate greater than estradiol greater than progesterone. The number of binding sites is limited (about 20 fmoles/mg protein) and the apparent equilibrium dissociation constant (KD) is 5 x 10(-9) M.

Studies of the androgen binding protein in the rete testis fluid of the ram and its relation to sexual season.

An androgen binding protein (ABP) with an electrophoretic mobility (Rf) of 0.56 is present in the rete testis fluid of adult rams. Its steroid specificity was found to be in the following order: 5alpha-DHT, testosterone, oestradiol-17 beta, dehydroepiandrosterone 5beta-DHT, androstenedione, cyproterone, cyproterone acetate, cortisol and progesterone. The characteristics of the ABP are similar to those found for the ABP of the testis and the epididymis of the rat and the rabbit. The concentration of ABP, determined by the dextran-coated charcoal method and sometimes confirmed by the steady-state polyacrylamide gel electrophoresis method, was significantly higher in the breeding season than in the non-breeding season (4.40 +/- 0.98 X 10(-9) M vs. 2.60 +/- 0.62 X 10(-9) M; P less than 0.037). The affinity constant of the ABP was independent of the season (2.45 +/- 0.21 X 10(9) M-1 vs. 2.66 +/- 0.1 X 10(9) M-1; NS). In addition, ABP was positively correlated with 5alpha-DHT (r = 0.506; P less than 0.0009), testosterone (r = 0.445; P less than 0.0003), total protein (r = 0.329; P less than 0.02) and spermatozoa (r = 0.406; P less than 0.006) in the RTF and with blood plasma testosterone (r = 0.584; P less than 0.0001). Furthermore, testosterone and 5alpha-DHT in RTF were positively correlated (r = 0.582; P less than 0.0001). These androgens were also correlated with plasma testosterone (r = 0.262, P less than 0.052 for testosterone in RTF; r = 0.341, P less than 0.018 for 5 alpha-DHT). Total proteins and spermatozoa were found to be positively correlated in the RTF (r = 0.789; P less than 0.0001).

Composition of uterine flushings from Large White and prolific Chinese Meishan gilts.

This study examined differences in selected components of uterine secretions from Large White and prolific Chinese Meishan gilts during the oestrous cycle or early pregnancy. Total recoverable protein, uteroferrin (measured as acid phosphatase activity), acyl aminopeptidase, calcium, sodium, potassium, immunoglobulins A and G, glucose, fructose, oestradiol-17 beta, and prostaglandins F2 alpha (PGF2 alpha) and E2 (PGE2) in uterine flushings were measured. During the oestrous cycle, breed effects were detected only for total protein (P = 0.07), which tended to be higher for Large White gilts. However, for pregnant gilts, total recoverable glucose (P less than 0.05), fructose (P less than 0.05) sodium (P less than 0.05), immunoglobulin A (P less than 0.01), PGF (P less than 0.01), PGE (P less than 0.01), and acyl aminopeptidase (P less than 0.05) were greater in uterine flushings from Meishan gilts. Only uteroferrin was higher (P = 0.06) in uterine flushings from Large White gilts. Concentrations of prolactin were higher (P less than 0.05) in plasma from cyclic or pregnant Meishan gilts, but concentrations of total oestrogens and progesterone were not affected by pregnancy status or breed. These results suggest that Meishan conceptuses, individually or collectively, are more stimulatory to endometrial secretion and/or transport of the components of histotroph into the uterine lumen, or that the endometrium of Meishan gilts is more sensitive to conceptus signals responsible for the accumulation of histotroph in the uterine lumen.

Failure to produce transgenic offspring by intra-tubal insemination of gilts with DNA-treated sperm.

The reproducibility of the use of sperm cells as vectors of foreign DNA in the genome of pigs was verified in the present study and the effectiveness of four different procedures for sperm treatment was assessed. For each gilt, approximately 6 x 10(6) ejaculated boar spermatozoa were incubated for 30 min in 1 mL TALP medium containing 3 micrograms of linearized pSV2CAT plasmid DNA. Before incubation, spermatozoa were treated in four experimental groups: (1) cells were stored at 16 degrees C for 24 h and then washed three times in TALP; (2) cells from the fresh, undiluted sperm-rich fraction of an ejaculate were used immediately after collection, following the same procedure as (1); (3) cells were treated as in (2) with an extra wash; and (4) incubation with DNA was performed in TALP medium supplemented with 0.5 mg mL(-1) poly-L-lysine hydrobromide. As determined by immunolocalization, plasmid DNA molecules were found to be associated with 12-17.1% spermatozoa, depending on sperm treatment. Of 35 inseminated gilts, 20 gave birth to a total of 126 piglets. None of the piglets showed sign of exogenous DNA incorporation in any of the tissues tested, as assessed by the polymerase chain reaction and Southern blot. The potential of modifying the pig genome through "transformed' spermatozoa was not confirmed by these experiments.

Dynamics of ovarian follicular development in cattle during the estrous cycle, early pregnancy and in response to PMSG.

Ovarian follicular dynamics of cattle were examined during the estrous cycle, early pregnancy and in response to PMSG. Number and size of follicles were monitored by ultrasonographic examinations. During the estrous cycle, distinct periods of follicular dominance (measured by the increase in difference in size between the largest and second largest follicle) occurred in both the luteal (Days 6-8) and proestrus (18-22) phases of the estrous cycle (two follicular waves). Associated with the well timed development of the first dominant follicle was a change in distribution of follicle numbers in small (less than 5 mm; increased on Days 2-4), medium (6-8 mm; increased on Days 3-5) and large (greater than or equal to 9 mm; increased on Days 6-9) follicular size classes. Follicular development was greater on the ovary bearing the CL for the period that the CL was present. The dominant follicle formed during the first follicular wave was capable of ovulating (6 of 8 heifers) following an injection of a synthetic analogue of prostaglandin F-2 alpha on Day 9 of the estrous cycle. During early pregnancy (Days 6-34), follicular development (size of largest follicle, number of follicles and total accumulated size of all follicles) on the ovary bearing the CL was suppressed between Days 24 and 34 of pregnancy. This was a local effect in that follicular development was sustained on the contralateral ovary. Therefore, the CL or conceptus may be regulating follicular development in a manner to help prevent luteolysis. Associated with the injection of PMSG was an initial increase in the number of small follicles followed by their recruitment into medium and large size classes leading to ovulation. Number of follicles greater than 5 mm on the Day of estrus was related (r = .97) to the number of subsequent embryos and oocytes collected. Ultrasonography is a valuable technique to monitor ovarian follicular dynamics in cattle, and can thereby be used to infer changes in physiological and endocrine states.

Dynamics of ovarian follicular development in cattle following hysterectomy and during early pregnancy.

Three experiments were conducted to evaluate ovarian follicular dynamics and functional activity during pregnancy in cattle. In 11 pregnant Charolais cows of Experiment I, size of largest follicle, number of follicles and accumulated follicle size were reduced by day 27 of pregnancy on the ovary bearing the corpus luteum (CL) but not on the non-CL bearing ovary. In experiment II, local attenuation of ovarian follicular development on the CL bearing ovary of seven pregnant heifers was evident compared to the contralateral ovary without the CL. However, in four hysterectomized heifers, follicular development was sustained on both the CL- and non-CL bearing ovaries when CL maintenance was achieved without presence of the uterus or conceptus. In Experiment III, steroidogenic characteristics of the largest and second largest follicles at 17 d postestrus were evaluated for seven pregnant and six cyclic cattle. Follicle by physiological status interactions were detected for both aromatase activity of the follicle and follicular fluid concentrations of estradiol and progesterone. In cyclic cows, the largest follicle had appreciably more aromatase activity than did the second largest follicle; whereas, aromatase activity of the largest follicle from pregnant cows was less than that of cyclic cows. However, in pregnant cows the second largest follicle became the estrogen-active follicle, and this follicle occurred with a higher frequency on the ovary contralateral to the CL-bearing ovary. These changes in aromatase activity were reflected by parallel changes in estrogen concentrations of follicular fluid. The higher progesterone concentration in follicular fluid of the largest follicle in pregnant cows provided further confirmation of their atretic status. In conclusion, during early pregnancy the conceptus and/or uterus ipsilateral to the conceptus appear to secrete compounds which alter local follicular steroidogenic activity and attenuate subsequent follicular growth between 17 to 34 d of pregnancy on the CL-bearing ovary. This local mechanism acting within the ovary may contribute to the antiluteolytic effects of early pregnancy in cattle.

Effect of sperm survival and CTC staining pattern on in vitro fertilization of porcine oocytes.

Polyspermy in pig oocytes fertilized in vitro remains unacceptably high. In this study, we evaluated the effects of gamete coincubation time, and determined if the proportion of capacitated spermatozoa would be predictive of the fertilizing ability of frozen-thawed semen in vitro. Cumulus-oocyte complexes were collected from slaughterhouse prepubertal gilt ovaries and matured in vitro for 44 h in TCM199, with EGF, FSH, cysteamine and follicular fluid. Fertilization was induced with 2 x 10(5) frozen-thawed spermatozoa/ml in TBM. Penetration of oocytes as well as polyspermic fertilization occurred 2 h after insemination. A strong correlation between penetration and polyspermic fertilization rates has been demonstrated, but there was no correlation between the proportion of capacitated spermatozoa, as assessed by chlortetracycline staining, at the time of insemination and fertilization rates. We also compared the results of IVF in three IVF media: TBM, m199 and TALP. Penetration and polyspermy were very different in these three media: 71 +/- 19% and 25 +/- 13% in TBM, 37 +/- 11% and 6 +/- 2% in m199, 10 +/- 2% and 0% in TALP, respectively. Nevertheless, survival of spermatozoa or modifications of the capacitation status were not different in these media after 6 h incubation. We concluded that survival and capacitation characteristics of the semen used for IVF could not be predictive of the IVF results. It seems necessary to act at the oocyte level to control both variability between replicates and the incidence of polyspermy. Improving the spermatozoa penetration blocking system of the oocytes and reducing the number of sperm-binding sites on the zona pellucida (ZP) are our further objectives.

Effect of passive immunization against testosterone on reproductive hormone secretion and ovarian function in dairy cows and pubertal beef heifers.

Multiparous dairy cows were divided in 3 groups from Day 5 up to Day 56 post partum: high energy level (Group H, n=10), low energy level (Group L, n=10) and low energy level plus anti-testosterone bovine immunoglobulins (Group LI, n= 10). Undernutrition decreased body weight, body condition score, milk yield and energy balance in Groups L and LI compared to Group H (P<0.05), but had no effect on secretory pattern of LH. Passive immunization against testosterone increased LH secretion in Group LI (P<0.05). Follicular score and the presence of follicles >/= 10mm on the ovary were not affected by underfeeding but were higher in Group LI than in Group L after immunization (P<0.01). The duration of the first luteal phase was shorter in Group H than in Groups L and LI and maximum progesterone levels reached were higher in Group LI than in Group H (P<0.01). Reproductive performance was not depressed by underfeeding and immunization. In the pubertal beef heifers maintained in anestrus by undernutrition had very low LH secretion. After passive immunization against testosterone, the increase of LH pulses number became almost significant (P=0.07). Following injection of exogenous LH, the number of follicles >/= 9mm was higher in immunized (Group I, n=8) than in control heifers (Group C, n=7). Group I developed a dominant follicle sooner and of greater size than Group C. Passive immunization against testosterone increased LH secretion and follicular development.

Effect of growth factors, EGF and IGF-I, and estradiol on in vitro maturation of sheep oocytes.

The objective of these experiments was to determine the effect of exogenous addition of insulin-like growth factor-I (IGF-I, 100 ng/mL), epidermal growth factor (EGF, 10 ng/mL) and estradiol (E2, 100 ng/mL) to the maturation medium of sheep oocytes on their subsequent development in vitro. Addition of IGF-I to the maturation medium did not improve nuclear or cytoplasmic maturation of sheep oocytes at the concentration tested. However, EGF improved significantly the resumption of meiosis (84% oocytes in metaphase II stage after IVM vs. 59% in medium alone). Cleavage rate and blastocyst development rates were improved (P<0.01) after addition of EGF (60% and 29%, respectively), as compared with maturation in TCM 199 alone (39% and 19%, respectively), but remained lower than rates observed after maturation in complete medium containing follicular fluid (FF, 10%) and FSH (81% and 35%, respectively). No additive effect of EGF over FSH was observed during these experiments. Addition of FF to FSH containing maturation medium improved significantly both cleavage (P<0.001) and blastocyst rates (P<0.05). Addition of E2 to the IVM medium is not required when medium already contains FF. However, in defined conditions supplementation of maturation medium with E2 had a positive effect. These results suggest that EGF, FSH and E2 may play an important role in the nuclear and cytoplasmic maturation of sheep oocytes in vitro.

Cloned ovarian cells from sheep: Cholesterol dependency in progesterone synthesis.

The TV4 cell line is derived from sheep ovarian tissues trypsinized for 60 min and developed from a clone after serial dilutions. The TV4 cells had a doubling time of 24 h in B2 medium with 10% fetal calf serum and 10% BSA. TV4 cells synthesized progesterone (P4) in the presence of cholesterol. As the concentration of cholesterol increased (0, 92.5 and 125 mg/l), synthesis of P4 increased (P<0.01) from 1.05 +/- 0.20 to 30.6 +/- 3.03 ng/ml. Kinetics of P4 production were determined; a linear production response (y = 5.816 + 1.05 x, y = ng/ml, x = hour of incubation; R(2) = 0.97) was observed with up to 35 ng/ml of P4 obtained by 30 h of incubation. Follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone, or FSH and testosterone did not have any effect on estradiol-17beta (E2) or P4 production. Aromatase activity measured by RIA and HPLC following incubation with either nonradiolabeled or labeled testosterone was undetectable. In conclusion, this study established a cell line from the sheep ovary which has a high ability of divide and produce progesterone.

Meiotic and developmental competence of prepubertal and adult swine oocytes.

The present study was conducted to compare meiotic and cytoplasmic competence of prepubertal and adult porcine oocytes, and the effects of EGF (0 to 100 ng/mL), FSH (0 to 400 ng/mL) and prepubertal pFF (0 to 10%) on nuclear maturation. Prepubertal oocytes were less responsive to FSH and pFF than were adult oocytes in terms of stimulation of nuclear maturation. The best nuclear maturation rates for prepubertal oocytes were obtained with 10 ng/mL EGF and 400 ng/mL FSH, whereas for adult oocytes no additional effect of EGF was seen in the presence of 400 ng/mL FSH. Supplementation with pFF had no additional effect on MII yield over that obtained with EGF plus FSH. After maturation in the presence of EGF, FSH and cysteamine, fertilization rates were not different between adult and prepubertal oocytes, but polyspermy was more frequent in prepubertal oocytes (31 +/- 17% vs. 17 +/- 7% in prepubertal and adult oocytes, respectively, P < 0.05). The addition of pFF to maturation medium decreased oocyte fertilization of adult oocytes and polyspermic fertilization in prepubertal oocytes. Blastocyst yield and developmental competence were significantly reduced in prepubertal oocytes compared to adult oocytes. The mean cell numbers in blastocysts cultured for 7 days ranged from 61 to 74, and did not differ among groups. Finally, the viability of the 2- to 4-cell embryos and blastocysts produced was assessed by embryo transfer experiments. One offspring was obtained after transfer of 2- to 4-cell embryos, and one after transfer of in vitro-produced blastocysts. In conclusion, although prepubertal gilt oocytes appeared less meiotically and developmentally competent than their adult counterparts, they can be used to produce blastocysts able to develop to term.

Correlations between chemical parameters, mitogenic activity and embryotrophic activity of bovine oviduct-conditioned medium.

To establish parameters predicting the quality of bovine oviduct epithelial cell-conditioned media, we compared media conditioned by oviduct cells from cows at Day 2 (n = 3) and Day 15 (n = 3) of the estrous cycle. In addition, we tested the influence of time of conditioning. Media were evaluated for their embryotrophic activity using a cumulus cell co-culture system as a control. The same media were tested for their mitogenic activity on NIH 3T3 cells and for chemical parameters, including total protein, and de novo synthesized protein as well as for concentrations of glucose, lactate and ammonium. Analysis of variance did not reveal a significant effect by stage of the estrous cycle on the embryotrophic activity of conditioned media. However, there was a significant effect by time of conditioning on the proportion of 5- to 8-cell embryos (P < 0.01) and of blastocysts and hatched blastocysts (P < 0.05). None of the conditioned media (19 to 31% blastocysts) was superior to the cumulus cell co-culture system (32% blastocysts). In the conditioned media, the proportion of 5- to 8-cell embryos correlated positively with mitogenic activity on 3T3 cells (r = 0.64; P < 0.05), whereas the proportion of blastocysts was not significantly correlated with this parameter. In summary, our results provide evidence for an effect of time of conditioning on embryotrophic activity of oviduct epithelial cell-conditioned media. The fact that mitogens for NIH 3T3 cells affect the proportion of 5- to 8-cell embryos but not of blastocysts suggests different culture requirements for early and late preimplantation stage development of bovine embryos.

Follicular growth and maturation in hyperprolific and large white sows.

The mechanisms whereby hyperprolific sows achieve their increased ovulation rate (+5 oocytes on average) compared with normal Large White sows were explored in this study. The following specific questions were asked. Is increased ovulation rate related to 1) increased follicular populations within the ovaries or 2) alterations in terminal follicular growth and maturation? The population of antral follicles in six sows of each genotype was studied using histological techniques on ovaries obtained at the preovulatory stage. No difference between the total number of antral follicles, number of healthy or atretic follicles in specific size classes, and in size of the preovulatory follicles could be detected. The number of granulosa cells contained in preovulatory follicles was also similar between genotypes. Terminal follicular growth and maturation were studied in 15 sows of each genotype killed at d 1, 3, or 5 (n = 5.d-1.genotype-1) after the end of Regumate administration (d 0). Small (< or = 3.5 mm) follicles were counted, and follicles > 3.5 mm were dissected, measured, and incubated in vitro. Steroid concentrations (estradiol and testosterone) in culture medium were then measured. The two genotypes differed in the patterns of growth of their ovulatory follicles, because at d 3 all ovulatory follicles were obvious in Large White sows. In contrast, between d 3 and 5, seven additional ovulatory follicles grew in hyperprolific sows. Differences in follicular maturation between genotypes were also detected. Whereas testosterone concentrations in culture medium were similar in the two genotypes, estradiol concentrations were markedly (P < .01) increased in hyperprolific follicles. This indicates that these follicles may have an increased aromatizing ability. How this generates the altered pattern of follicular growth and the increased ovulation rate of hyperprolific sows remains to be established.

Effects of various hormone treatments on induction of lactation in the ewe.

In Exp. I and II, 52 of 68 ewes were induced into lactation with twice-daily injections of estradiol-17 beta (E2-beta) and progesterone (P4; .5 and 1.25 mg/kg body weight/day) for 7 days. Additional treatments were twice-daily injections (days 18 to 20) of hydrocortisone, growth hormone, thyroxine and thyrotropin releasing hormone alone or in various combinations. In Exp. III, 12 ewes were induced into lactation. In this experiment, all ewes were injected with E2-beta and hydrocortisone, as previously, but four ewes (III-2) had P4 injections extended to day 20, and four ewes (III-3) were not injected with P4. Across experiments, lowest milk yields during lactation and the lowest percentage of ewes induced into lactation (58%) occurred when only E2-beta and P4 were injected. Inclusion of hydrocortisone injections (50 mg/day) induced the highest percentage of ewes into lactation (86%, 38 of 44), the highest peak daily yields of milk and the highest total yields during lactation. Including injections of growth hormone, thyroxine or thyrotropin releasing hormone alone or in combinations did not produce better results than injections of E2-beta and P4 alone. Injections of E2-beta and hydrocortisone without concurrent injections of P4 were less effective. Intramuscular injections of P4 (10 mg/day) from days 8 to 20 did not inhibit lactogenesis or subsequent lactation. Across all experiments, 76% of multiparous (52/68) and 50% of nulliparous (6/12) ewes produced greater than 100 ml milk/day during their lactation (34 to 95 days). However, yields of milk for ewes that lactated were only 25 to 50% of those from postpartum ewes. The importance of including injections of hydrocortisone in the induction procedure was established, but determination of optimum time to inject and potential importance of other hormones requires additional research.

Induction and synchronization of ovulations of nulliparous and multiparous sows with an injection of gonadotropin-releasing hormone agonist (Receptal).

The objective of this study was to determine if administration of a set dose (10 microg) of a gonadotropin-releasing hormone agonist, buserelin (Receptal; Rc), at set times after altrenogest (Regumate; RU) treatment or after weaning was able to induce and synchronize ovulation in female swine (gilts and sows). The pubertal (n=187) gilts were allocated to four groups, all synchronized with RU. Group 1 (RU) was inseminated twice at detected estrus, Group 2 (RU+Rc120) and Group 4 (RU+Rc104) received 10 microg Rc at 120 or 104 h after the end of RU treatment, respectively, and Group 3 (RU+eCG+Rc104) was treated with 800 IU equine chorionic gonadotropin (eCG) at 24h and Rc 104 h after the end of RU treatment, respectively. Gilts were inseminated twice at predetermined times, namely 144 and 168 h (Group 2), 128 and 144 h (Group 3), and 144 and 152 h (Group 4) after the end of RU treatment, respectively. Pregnant gilts were slaughtered at 30 d. Administration of Rc 104 h after the end of RU feeding synchronized ovulation over a 24-h time window in 97.9% and 100% of the gilts of Groups 3 and 4, respectively, whereas Rc administration at 120 h (Group 2) only successfully synchronized 88.9% of the gilts over 24h. Ovulation rates of gilts of Groups 2 and 4 were similar to that of the control group. Pregnancy rates were numerically higher in Groups 2 and 3 (92% and 96%, respectively) compared with those of Groups 1 and 4 (84% and 81%, respectively). Combination of eCG with Rc administration at 104 h (Group 3) increased ovulation rate (+4 CL) but decreased embryo survival to 62% at Day 30. The weaned sow experiment involved 61 sows of a range of parities (2.7+/-0.9), allocated to two control groups (Control 104 group and Control 94 group) and two treated groups (Rc104 group and Rc94 group), which received 10 microg Rc at 104 and 94 h after weaning, respectively. The females were inseminated at detected estrus. All pregnant sows farrowed. After treatment with Rc 94 h after weaning, 100% of sows ovulated over a 24-h time window versus only 68.7% of controls. Farrowing rate and litter size of the sows treated with Rc at that time were unaffected compared with that of control sows. In contrast, Rc administration at 104 h after weaning may have been too late; only 66.7% of the treated sows ovulated during a 24-h period. This proportion was numerically lower but not significantly different than that for control sows. Farrowing rate and litter size of treated sows were not significantly different than that of controls. Administration of Rc at the dose and times selected in this study tightened synchrony of ovulation in gilts and in sows after weaning. It remains to be established if such a synchrony is suitable to obtain good fertility after a single artificial insemination at a predetermined time.