Estrogen and progesterone receptors in the ovine cervix during the postpartum period

Cervical estrogen (E) and progesterone (P) receptors were characterized and quantified during the postpartum period in Corriedale ewes lambing in the late breeding season. Cervices and uteri were collected after ovariohysterectomy at 1 d (n = 2), 5 d (n = 4), 17 d (n = 2) or 30 d (n = 2) post partum. The estrogen and progesterone receptors were measured using binding assays with tritiated hormones, dextran charcoal separation and inverse Scatchard analysis. Similar kinetic parameters in cytosolic binding sites for both hormones were found in all cervical and uterine samples, indicating that the binding protein in both tissues is of the same nature. Receptor concentrations (fmol/mg cytosolic protein) in the cervix of early (1 to 5 d, n = 6) and late (17 to 30 d, n = 4) postpartum ewes were 348 +/- 66 vs 994 +/- 145 (P < 0.05) for E and 618 +/- 126 vs 1170 +/- 201 (P < 0.05) for P, respectively. These data suggest an increased synthesis of receptors, probably due to the presence of ovarian estrogen-active follicles. Cervical E and P receptor concentrations were similar or higher than those in the uterus (1.40 +/- 0.15, n = 10 and 1.51 +/- 0.19, n = 10; for E and P respectively), and these receptor ratios did not differ between the early and late postpartum period. The high ratio between cervical/uterine receptors suggests that the ovine cervix may be a very sensitive to steroid action. In conclusion, it was shown that restoration of steroid receptors during the postpartum period in the ovine cervix is similar to receptor dynamics in the uterus, and is probably associated with the recovery of ovarian cyclicity.     

Effect of intravaginal implants of melatonin on the onset of ovarian activity in adult and prepubertal ewes

Melatonin was administered intravaginally in Silastic tubing to adult and prepubertal ewes. In Exp. 1, ewe lambs (born early March) were given intravaginal melatonin implants at a mean age (+/- s.e.m.) of 7.5 +/- 0.1 weeks (Group E, N = 10) or 19.4 +/- 0.2 weeks (Group L, N = 10). The third group (Group C, N = 10) received empty implants. In Exp. 2 mature ewes were given implants on 13 May (Group E, N = 10) or 18 July (Group L, N = 10) or received empty implants (Group C, N = 10) on one of these two dates. Blood samples were taken twice weekly for progesterone assay. In Exp. 1 the mean age (+/- s.e.m.) at puberty (progesterone greater than 2 nmol/l for two consecutive samples) was 35.4 +/- 0.8 weeks. Puberty was advanced by 5.2 weeks in Group L lambs, occurring at a mean age of 30.2 +/- 0.7 weeks (P less than 0.001). In Group E lambs the timing of puberty was unaltered, occurring at a mean age of 34.8 +/- 0.6 weeks. Mature ewes in Group L (Exp. 2) showed increased incidence of ovarian activity (9/10 ewes cycling by 26 September) compared with the control ewes (1/10) (P less than 0.001), but there was no effect in Group E ewes (3/10). The results demonstrate that continuous melatonin administration to adult and prepubertal ewes can mimic the effect of short days in terms of the reproductive response, and that the present and previous exposure to melatonin is critical in determining the response.     

Uterine steroid hormone receptors during the estrous cycle and during hibernation in the Turkish hamster (Mesocricetus brandti)

The Turkish hamster is a long-day breeder that hibernates for 4-5 mo if exposed to a short-day, cold environment. The objective of this study was to assess the uterine responsiveness of the hibernating animal to ovarian steroids. Our approach was 1) to characterize and determine uterine estrogen (E) and progesterone (P) receptors (R) during hibernation as compared to the levels observed in cycling females that had terminated hibernation, and 2) to assess the responsiveness of the uterus to E during hibernation by its ability to induce uterine P receptor. Females were exposed to short days (10L:14D) for 2 mo and then were placed in a cold-room (10L: 14D, 6 +/- 1 degrees C). After 2 or 4 mo in the cold, hibernating animals were killed and uterine steroid receptors were determined by 3H-steroid binding assay. Uterine receptors were also determined in cycling Turkish hamsters on each morning of the estrous cycle. Values for uterine receptors (pmol/g tissue, n = 4-6) during the estrous cycle (estrus, diestrus I, diestrus II, proestrus) were: 4.3 +/- 0.78, 3.9 +/- 0.19, 4.1 +/- 0.25, 3.7 +/- 0.5 for cytosolic ER; 36.6 +/- 5.8, 32.2 +/- 6.8, 36.3 +/- 1.5, 54.4 +/- 1.9 for cytosolic PR; 0.59 +/- 0.11, 0.54 +/- 0.07, 1.06 +/- 0.05, 1.42 +/- 0.17 for nuclear ER. Hibernating (torpid) animals sampled after 2 mo in the cold showed a significant (p less than 0.05) depression of cytosolic ER (2.6 +/- 0.12, n = 5) and cytosolic PR (19.0 +/- 2.6, n = 8) as compared to any day of the estrous cycle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of the antiprogestin RU-486 on progesterone inhibition of occupied nuclear estrogen receptor in the uterus

The ability of the antiprogestin, RU-486, to reverse progesterone (P) antagonism of occupied nuclear E receptor retention was studied in the rat and hamster uterus. RU-486 was shown to effectively displace [3H]P binding from rat uterine cytosolic P receptor in in vitro competition assay. In contrast, no competition by RU-486 for [3H]P binding was observed for uterine cytosolic P receptor from the hamster uterus. In the presence of sustained serum levels (silastic implants) of P and estradiol (E), occupied nuclear E receptor was significantly inhibited in the rat uterus. At 6, 12 and 24h after RU-486 treatment (5 mg/animal, s.c.) uterine receptors for E and P were determined. No significant differences in cytosolic E and P receptors were observed between treated (E + P, + RU-486) and control (E + P alone) animals. However, by 6 h following RU-486 treatment, occupied nuclear E receptor retention increased significantly (0.30 +/- 0.05 vs 0.60 +/- 0.09, pmol/uterus) and reached a peak between 12 h (1.32 +/- 0.09) and 24 h (0.83 +/- 0.09). The increase in nuclear E receptor approached the level observed in animals with an E implant alone (1.55 +/- 0.15). Measurement of uterine fluid accumulation following RU-486 treatment showed an increase which paralleled that observed for occupied nuclear E receptor retention. A similar in vivo experiment in the hamster showed no reversal of P inhibition of occupied nuclear E receptor. These results show that: 1. RU-486 is an effective competitor for rat uterine P receptor but not hamster P receptor; 2. RU-486 can rapidly reverse P inhibition of uterine occupied nuclear E receptor in the presence of sustained serum levels of E and P; 3. The recovery of occupied nuclear E receptor is coincident with a resumption of E action (uterine fluid accumulation). The studies also provide a novel means by which antiprogestin activity can be assessed in vivo in the presence of sustained E and P serum levels, e.g. the reversal of P inhibition of uterine nuclear E receptor retention.     

The effect of oxytocin and PGF2alpha on the uterine involution and pregnancy rates in postpartum Arabian mares

In this study, the effects of oxytocin and an analog of prostaglandin (cloprostenol) on the uterine involution and pregnancy rates were investigated. Mares received 3 ml of 0.9% NaCl in Group C (n=10), 30 IU/mare of oxytocin in Group O (n=10) and 250 microg/mare of cloprostenol in Group P (n=10) within 12h after parturition. The gravid uterine horn's cross-sectional diameter was measured by ultrasonography. The mean uterine diameters did not differ significantly between the treatment (O and P) and the control (C) groups (p>0.05). The difference between the postpartum ovulation periods (Group C: 12.6+/-0.72 days, Group O: 15+/-1.33 days, Group P: 14.6+/-1.11 days), the pregnancy rates at foal heat (Group C: 60%, Group O: 60%, Group P: 80%) and the embryonic death rates at foal heat (Group C: 33.3%, Group O: 16%, Group P: 25%) were not found to be statistically significant between the treatment and the control groups. The mean progesterone concentrations were similar in all groups and decreased continuously from parturition to until foal heat (Group C: from 2.43+/-0.24 to 0.66 ng/ml, Group O: from 3.07+/-0.6 to 0.27+/-0.27 ng/ml and Group P: from 2.8+/-0.44 to 0 ng/ml) (p>0.05). In conclusion, it was decided that the oxytocin and PGF2alpha treatments performed on the mares with the purpose of stimulating involution had no effect on the duration of parturition-first ovulation, the shrinkage of the uterus diameter, the pregnancy and embryonic death rates.     

The influence of estradiol and progesterone on the concentrations of uterine oxytocin receptors and plasma PGFM in response to oxytocin in ovariectomized gilts

Peripubertal gilts (n = 25) were treated with corn oil (CO) or ovarian steroids, one month following an ovariectomy. The first day of treatment was assigned as the first day of the experiment. The gilts received: Group (Gr) I (n = 4)--CO (2 mL x day(-1) from 1st to 12th day), Gr II (n = 4) and Gr III (n = 4)--progesterone (P4; 10 to 100 mg x day(-1) from 1st to 12th day), Gr IV (n = 5)--estradiol benzoate (EB; 400 microg x day(-1) from 1st to 3rd day), Gr V (n = 4) and Gr VI (n = 4)--EB + P4 (EB 400 microg x day(-1) from 1st to 3rd day, 20 microg x day(-1) at 6th and 9th day, 50 microg at 12th day plus P4 10 to 100 mg from 4th to 15th day). All gilts were injected with oxytocin (OT; 20 IU; i.v.) on the following days of the experiment: 13th (Gr I and Gr II), 15th (Gr III and Gr IV), 16th (Gr V) and 18th (Gr VI). Concentrations of the PGF2alpha metabolite--PGFM were determined in blood samples, collected from 30 min before to 120 min after OT injection. Baseline PGFM concentrations (30 min before OT) differed among treatment groups and were the highest in Gr V and Gr VI (P < 0.01 vs. other groups). The magnitude of the PGFM response to OT increased only in four of the five gilts of Gr IV and in three of the four gilts of Gr VI, and it was higher (P = 0.009) in Gr VI than in Gr IV. In the remaining groups, PGFM concentrations did not increase above the baseline in response to OT. The day after OT injection, oxytocin receptors (OTR) were found in the uterine tissues of all animals studied. The lowest OTR concentrations were in Gr I--75.5 +/- 11.2 fmol x mg protein(-1) and the highest in Gr IV--712.9 +/- 86.7 fmol x mg protein(-1); (P < 0.05 vs. other groups). The values of K of OTR differed among groups (P < 0.001) and ranged from 1.62 +/- 0.44 nM in Gr I to 12. 08 +/- 1.9 nM in Gr VI. A positive correlation (r = 0.54; P < 0.01) between plasma E2 and uterine OTR concentrations was observed. In conclusion, E2 and P4 are involved in both PGF2 synthesis/secretion and OTR formation, however, full PGF response to OT does not develop before puberty. Estrogens are evident stimulators of uterine OTR synthesis ingilts.     

Endothelial vasodilator production by uterine and systemic arteries. VII. Estrogen and progesterone effects on eNOS

Uterine blood flow (UBF) and uterine artery endothelial nitric oxide synthase (eNOS) expression are greatest during the follicular vs. luteal phase. 17 beta-Estradiol (E(2)beta) increases UBF and elevates eNOS in ovine uterine but not systemic arteries; progesterone (P(4)) effects on E(2)beta changes of eNOS remain unclear. Nonpregnant ovariectomized sheep received either vehicle (n = 10), P(4) (0.9 g Controlled Internal Drug Release vaginal implants; n = 13), E(2)beta (5 microg/kg bolus + 6 microg x kg(-1) x day(-1); n = 10), or P(4) + E(2)beta (n = 12). Reproductive (uterine/mammary) and nonreproductive (omental/renal) artery endothelial proteins were procured on day 10, and eNOS was measured by Western analysis. P(4) and E(2)beta alone and in combination increased (P < 0.05) eNOS expression in uterine artery endothelium (vehicle = 100 +/- 16%, P(4) = 251 +/- 59%, E(2)beta = 566 +/- 147%, P(4) + E(2)beta = 772 +/- 211% of vehicle). Neither omental, renal, nor mammary artery eNOS was altered, demonstrating the local nature of steroid-induced maintenance of uterine arterial eNOS. In the myometrial microvasculature, eNOS was increased slightly (P = 0.06) with E(2)beta and significantly with P(4) + E(2)beta. Systemic NO(x) was increased with P(4) and P(4) + E(2)beta, but not E(2)beta, suggesting differential regulation of eNOS expression and activity, since P(4) increased eNOS in uterine artery endothelium while E(2)beta and the combination further increased eNOS protein.     

Neonatal exposure to progesterone and estradiol alters uterine morphology and luminal protein content in adult beef heifers

Exposure of the developing urogenital tract to steroids can affect structure and function of adult tissues and compromise reproductive performance. This study was conducted to determine 1) if exposure of neonatal heifer calves to progesterone (P) and estradiol benzoate (E), delivered from a commercial growth-promoting implant, would affect adult uterine morphology or uterine luminal protein content; and 2) whether such effects would be related to neonatal age at the first exposure. At birth (Day 0), 20 crossbred beef heifers were assigned to 1 of 4 treatment groups (n = 5 per group), defined by age at implant placement. Heifers either received an implant on Days 0, 21 or 45, or served as untreated controls. The heifers were maintained together and slaughtered at 15 mo of age, during the luteal phase of an induced estrous cycle, when reproductive tracts and blood samples were obtained. Peripheral plasma P concentrations were determined by RIA. Uterocervical wet weights were recorded, and uterine luminal flushings (ULF) were assayed for total protein. Cross-sections of uterine tissues were evaluated histomorphometrically to determine myometrial and endometrial areas and relative endometrial gland density. Treatment did not affect plasma P concentrations (3.2 +/- 0.5 ng/ml). Regardless of age at treatment, neonatal PE exposure reduced uterocervical wet weight by 35% (112.8 < 173.9 +/- 13.9 g; P < 0.01), myometrial area by 23% (125.3 < 162.8 +/- 8.5 mm2; P < 0.02), and endometrial area by 27% (33.3 < 45.4 +/- 2.7 mm2; P < 0.09) compared with the untreated controls. Endometrial gland density was reduced (P < 0.01) by 40% in treated heifers. This effect was related to age at implant placement. Uterine gland density was reduced (P < 0.01) by 65% in heifers treated at birth, while reductions of 22 and 33% were observed for heifers treated on neonatal Day 21 or 45, respectively. Consistently, ULF protein content was lower (P < 0.01) in the treated heifers (2.67 < 4.98 +/-. 72 mg/ULF). Thus, exposure of newborn calves to PE can have profound effects on adult uterine morphology and environment, the extent of which may depend upon the developmental period when exposure occurs. The potential of such alterations to affect reproductive performance in adult beef heifers remains to be investigated.     

Gonadotrophin secretion and ovarian responses in prepubertal heifers actively immunized against androstenedione and oestradiol-17 beta

Groups of heifer calves received a primary immunization against androstenedione (Group A; N = 11) or oestradiol-17 beta (Group E; N = 10) at 3 months of age and booster injections on 5 occasions at 2- to 3-month intervals. Controls (Group C, N = 11) were immunized against human serum albumin alone using the same protocol. Immunity was achieved against both steroids as judged by the secondary antisteroid antibody titres in Group A (1126 +/- 261; reciprocal of titre) and Group E (10,357 +/- 4067) heifers. In Groups A and E there was a general decline in the respective peak antibody titres after successive booster injections. From 3 to 9 months of age mean plasma concentrations of LH were higher (P less than 0.05) in Group E heifers (0.89 +/- 0.08 ng/ml) than in Group C (0.46 +/- 0.03 ng/ml) and Group A (0.59 +/- 0.05 ng/ml) heifers which did not differ from one another. There were no differences between groups in plasma FSH concentrations. At 10 months of age the LH response to exogenous LHRH was of higher (P less than 0.05) amplitude for heifers in Group E (2.59 +/- 0.56 ng/ml) than for those in Groups C (0.61 +/- 0.07 ng/ml) and A (1.04 +/- 0.22 ng/ml). Elevated plasma progesterone concentrations at 5 months of age were shown by 2 heifers in Group C, 10 in Group A, and 6 in Group E. From 8 to 14 months of age a consistently higher proportion of Group A heifers exhibited elevated progesterone compared with Group C and Group E heifers. After ovarian synchronization and booster injection at 15 months of age a corpus luteum was present in 2 heifers in Group C, 7 in Group A and none in Group E. The ovaries of Group A heifers were different from those of Groups C and E and were characterized by greater numbers of 2-4 mm follicles. It is concluded that active immunization against gonadal steroids influences both LH secretion and ovarian function in prepubertal heifers. Early increases in ovarian activity in androstenedione-immunized heifers are maintained after puberty and may therefore confer some lifetime reproductive advantages.     

Effect of estradiol-17beta on PGF and total protein content in bovine uterine flushings and peripheral plasma concentration of 13, 14-dihydro-15-keto-PGF(2alpha)

Two experiments were conducted to examine the effect of estradiol-17beta (E(2)-17beta) on content of immunoreactive prostagladin F(2)alpha (PGF, ng) and total protein (TUP, mg) in uterine flushings, as well as concentrations of 13, 14-dihydro-15-keto-PGF(2)alpha (PGFM) in plasma (Pg/ml). In experiment 1, Holstein heifers were utilized in a single reversal trial in which either E(2)-17beta (3 mg in 2 ml saline/ethanol 50:50; n=5) or vehicle alone (n=6) were given intravenously on day 14 or 15 of the estrous cycle (Period 1) following an induced estrus (day of estrus = day 0). Treatment (Trt) groups were reversed in Period 2 (Day 14 or 15 of the second estrous cycle). Jugular venous plasma was obtained before treatment (Oh), and at 5, 6, and 9h posttreatment (PT). Uterine flushings were collected nonsurgically in vivo , per cervix, via Foley catheter at 6h PT (20 ml of .9% saline per uterine horn). E(2)-17beta did not significantly alter (E(2)-17beta vs vehicle; x(-) +/- S.E.M.) PGF (1674 +/- .11 +/- 338.39 vs 1889.91 +/- 400.24 ng; P> .10) or TUP (33.25 +/- 2.57 vs 39.16 +/- 3.04 mg; P > .10). However, E(2)-17beta increased (P < .05) plasma PGFM (E(2)-17beta vs vehicle) after treatment (0h, 113.2 vs 163.8; 5h, 312.5 vs 203.9; 6h, 324.5 vs 198.0; 9h, 323.2 vs 246.8, pg/ml). In experiment 2, crossbred beef cattle received comparable treatments of either E(2)-17beta (n=5) or vehicle (n=5) on day 14 or 15 postestrus. Jugular venous plasma was obtained at 0h PT, and at 6h PT. Uterine flushings (1.9% saline, 20 ml per uterine horn) and peripheral plasma were collected at slaughter. Estradiol-17beta increased PGF (30.07 +/- 5.94 vs 8.46 +/- 2.01 ng; P> <.05) in uterine flushings as well as PGFM in plasma (E(2)-17beta : 55.82 +/- 19.13 pg/ml, at 0h and 89.31 +/- 14.02 pg/ml, at 6h, vs saline: 103.46 +/- 50.73 pg/ml, at 0h and 17.78 +/- 14.22, at 6h). Estradiol-17beta stimulated uterine production and release of PGF and protein as measured in flushings (experiment 2) as well as plasma PGFM responses (experiments 1 and 2). Uterine and/or cervical stimulation of experiment 1 may have masked uterine response to E(2)-17beta.     

Effect of insulin on feed intake and reproductive performance of well-nourished nulliparous ewes

Eighty-four nulliparous ewes were used to examine the effect of short-term insulin treatment on feed intake and reproductive performance. Following estrus synchronization, ewes were observed for estrus (= Day 0) and were penned individually beginning on Day 7. Ewes were fed twice daily and feed intake was recorded. On Days 9 through 13, ewes were treated s.c. with 1 IU/kg BW insulin (n = 44) or an equivalent volume of saline (n = 40). On Day 14, ewes were placed with fertile rams and number of ewes in estrus (bred) was recorded. Thirty days post-breeding, ewes were checked for pregnancy via ultrasonography. Feed intake and percentage of ewes in estrus did not differ between saline- and insulin-treated ewes. Similarly, neither pregnancy rate (69 +/- 8.7% vs. 80 +/- 8.1%, respectively) nor lambing rate (61 +/- 8.9% vs. 78 +/- 8.4%, respectively) differed between treatments. The number of lambs born per ewe was, however influenced by a breed-group effect (P < 0.0002). Romanov ewes had more (P < 0.001) lambs than the other breed groups in the study. Therefore, treating well-nourished, nulliparous ewe lambs with insulin did not increase reproductive efficiency, possibly because the ewes were already at a maximal nutritional and/or reproductive state.     

The ratio of plasma bioactive to immunoreactive ACTH-like activity increases with gestational age in the fetal lamb.

The fetal ovine pituitary-adrenal axis plays an important role in the timing of parturition, in fetal lung maturation, and in fetal and neonatal responses to stress. While the ovine pituitary during the last third of gestation (term = 145 days) is capable of secreting immunoreactive ACTH (iACTH) in response to various stimuli, plasma cortisol levels frequently do not reflect the rise in plasma ACTH. Therefore, we examined the relationship between plasma iACTH and steroidogenic ACTH-like activity (bACTH) in a group of immature fetal lambs (Group I: gestational age = 97 +/- 2 days, mean +/- SEM, n = 16) and a group of near-term fetuses (Group II: gestational age = 136 +/- 1 days, n = 13) following acute exteriorization. Plasma iACTH was determined by RIA. Plasma bACTH was determined by the ability of glass-extracted material to stimulate corticosterone (B) production in an acutely dispersed rat adrenal bioassay. Plasma iACTH and bACTH levels varied among animals within age groups, with iACTH tending to be higher in immature fetal lambs (Group I) than near-term lambs (Group II) and bACTH being higher (P < 0.05) near term than earlier (Group I: iACTH = 807 +/- 273 pg/ml, bACTH = 173 +/- 44 pg/ml; Group II: iACTH = 405 +/- 85 pg/ml, bACTH = 371 +/- 96 pg/ml). The proportion of iACTH that had biologic activity (e.g. B/I ratio) was significantly greater in the older than in the younger fetuses (Group II: B/I = 0.862 +/- 0.109; Group I: B/I = 0.462 +/- 0.105 P < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Ultrasonographic evaluation of uterine involution and postpartum follicular dynamics in French jennies (Equus asinus)

Uterine involution and follicular dynamics during postpartum period were studied ultrasonographically in French jennies. For the study of uterine involution in postpartum jennies (n = 6, Group S), sonographic measurements of different parts of the uterus and endometrium were made at three-day interval, starting from the day of foaling and continued up to 33 days postpartum. Uterine dimensions were also recorded in non-pregnant jennies (n = 3, Group C) throughout a cycle and compared with the dimensions of Group S jennies observed on the day of complete involution. Follicular dynamics of first and second postpartum ovulatory cycles were studied and compared with that of the single estrous cycle of Group C jennies. Jugular venous blood samples of Group S jennies were collected at weekly intervals for 49 days, commencing at the appearance of first preovulatory follicle, to support the sonographic findings. The average involution period was 22.5 +/- 1.7 days. However, it was significantly delayed (P < 0.05) in jennies which came into first postpartum ovulatory heat within Day 9 than those who came later (25.0 +/- 1.0 versus 20.0 +/- 1.0). The endometrial layer was not discernible beyond Day 15 postpartum and thus was found to be unreliable index of uterine involution. The follicular growth rate (mm per day) and diameter (mm) of preovulatory follicle in postpartum jennies were similar to that in normal cycling jennies (P > 0.05). The first and second ovulations occurred at 14.6 +/- 0.8 and 39.0 +/- 0.8 days postpartum in Group S jennies. All the corpora lutea, either echogenic or centrally non-echogenic were functionally similar and had similar life span (P > 0.05). In conclusion, the postpartum reproductive events related to uterine involution and ovarian cyclicity apparently resemble that of mares.     

Urinary progesterone in free-ranging red howler monkeys (Alouatta seniculus): preliminary observations of the estrous cycle and gestation

The goals of this study were to develop and validate a radioimmunoassay (RIA) for measurement of unconjugated progesterone (P) concentrations in the urine of red howler monkeys (Alouatta seniculus) and to use urinary P profiles to characterize the reproductive cycle of this species. Analysis of P profiles from two females provided a preliminary estimate of the length of the estrous cycle (mean days +/- S.E.M. = 29.5 +/- 1.5; n = 2), and indicated that one female red howler copulated throughout two apparent estrous cycles. Urinary P concentrations during two confirmed pregnancies (211.8 +/- 29.7 ng P/ml) were higher (P < 0.05) than during the luteal phase (77.4 +/- 10.6 ng P/ml; n = 4) of the cycle.     

Superovulatory and endocrine responses in heifers treated with FSH-P at different stages of the estrous cycle

Forty-two Holstein heifers were superovulated with FSH-P (total dose, 30 mg) and cloprostenol. Treatment was initiated on Day 3 (Group D3, n = 11), Day 6 (Group D6, n = 11), Day 9 (Group D9, n = 10) or Day 12 (Group D12, n = 10) of the estrous cycle. Heifers were bled daily for serum progesterone and estradiol-17beta determinations and every 6 h for a 48-h duration at the expected time of estrus for luteinizing hormone (LH) assay. Ova and embryos were flushed from the reproductive tracts and the number of corpora lutea (CL) were recorded after slaughter on Day 7 post-estrus. Mean (+/- SEM) numbers of observed CL were higher (P < 0.05) in Group D9 (33.3 +/- 4.8) than in Group D3 (15.3 +/- 3.8), with Group D6 (17.0 +/- 2.9) and Group D12 (23.9 +/- 7.3) being intermediate. Similarly, mean (+/- SEM) numbers of fertilized embryos were highest (P < 0.05) in Group D9 (13.3 +/- 2.2). There was also a nonsignificant trend for the number of transferable embryos to be greatest in Group D9. Neither serum progesterone concentrations 3 d after the LH peak nor peak serum estradiol 17beta concentrations differed among groups, but both were significantly correlated with numbers of observed CL and total ova and embryos.     

Reproductive hormone secretion and testicular growth in bull calves actively immunized against testosterone and oestradiol-17 beta

Groups of bull calves received a primary immunization against testosterone (Group T; N = 7) or oestradiol-17 beta (Group E; N = 9) at 3 months of age and booster injections on four occasions at approximately 2 month intervals. Controls (Group C, N = 7) were immunized against human serum albumin alone using the same protocol. Immunity was achieved against both steroids as judged by the secondary antisteroid antibody titres in Group T (730 +/- 231; reciprocal of titre) and Group E (12,205 +/- 4366) bulls; however, peak antibody titres generally declined with successive booster injections. Mean plasma concentrations of LH, FSH and testosterone during the period from 3 to 10 months of age were higher (P less than 0.05) in Group T bulls than in Groups C and E. Group T bulls had larger testes compared with controls from 6 months of age onwards. At castration at 14 months of age, testes of Group T bulls were heavier (P less than 0.05) than those of Groups C and E (179 +/- 13, 145 +/- 8 and 147 +/- 6 g, respectively). At 10 months of age, there were no differences among treatment groups in LH responses to LHRH, but the testosterone responses were greater (P less than 0.05) in bulls in Group T (26.2 +/- 4.9 ng/ml) and Group E (16.6 +/- 1.8 ng/ml) compared with those in Group C (6.9 +/- 0.6 ng/ml). Testosterone responses to hCG determined at 13 months of age were also greater (P less than 0.05) in Groups T and E relative to controls. At 14 months of age daily sperm production rates per bull (X 10(-9)) were higher (P less than 0.10) in Group T bulls (2.2 +/- 0.1) than those in Groups C (1.6 +/- 0.2) and E (1.6 +/- 0.1). These results indicate that early immunity against testosterone is associated with increased gonadotrophin secretion and accelerated growth of the testes in prepubertal bulls. Also, chronic immunity against testosterone or oestradiol-17 beta enhances the steroidogenic response of bull testes to gonadotrophic stimulation. If the above responses observed in young bulls are shown to be sustained, then immunity against gonadal steroids early in life may confer some reproductive advantage in mature animals.     

Effect of bromocriptine on prostacyclin release and cyclic nucleotides on rat aortic and uterine tissues

The effect of bromocriptine mesylate on cyclic nucleotides and PGI2 release by rat aortic and uterine tissues was investigated. Treatment of rats with bromocriptine (10 mg kg-1 I.P. daily for 14 days) increased PGI2 release by the thoracic aorta from 0.67 +/- 0.02 to 1.4 +/- 0.03 ng/mg wet tissue (P less than 0.001; n = 6). This increase was antagonized by treatment with sulpiride (15 mg kg-l). Incubation of the arterial tissue with bromocriptine (50 micrograms ml-1) in vitro also stimulated PGI2 release. Mepacrine (160 micrograms ml-1) significantly decreased both basal and stimulated PGI2 release. Incubation of myometrial tissue from pregnant rats with bromocriptine (50 micrograms ml-1) in vitro significantly decreased PGI2 release from 1.25 +/- 0.07 to 0.60 +/- 0.08 ng/mg wet tissue (P less than 0.05, n = 6). It also elevated uterine cAMP from 40 +/- 2 to 64 +/- 3 pmoles/100 mg wet tissue. Both effects were antagonized by sulpiride. Bromocriptine did not affect uterine cGMP or the cyclic nucleotides in the aorta. It is concluded that the increase in aortic PGI2 was mediated via activation of dopamine D-2 receptors that stimulate phospholipase A2 enzyme. The decrease in myometrial PGI2 release may be related to the increase in uterine cAMP resulting from activation of dopamine D-1 receptors. Previous studies suggested a role for PGI2 in implantation in the rat. The results suggest that the inhibitory effect on uterine PGI2 may underlie the reported inhibition of bromocriptine on implantation. On broad basis, the decrease in uterine PGI2 together with the reported luteolytic effect of bromocriptine point to a potential role for the compound in postcoital contraception.     

Estrous cycle variation of TRPV1-mediated cross-organ sensitization between uterus and NMDA-dependent pelvic-urethra reflex activity

Cross-organ sensitization between the uterus and the lower urinary tract (LUT) underlies the high concurrence of pelvic pain syndrome and LUT dysfunctions, and yet the role of gonadal steroids is still unknown. We tested the hypothesis that cross-organ sensitization on pelvic-urethra reflex activity caused by uterine capsaicin instillation is estrous cycle dependent. When compared with the baseline reflex activity (1.00 +/- 0.00 spikes/stimulation), uterine capsaicin instillation significantly increased reflex activity (45.42 +/- 9.13 spikes/stimulation, P < 0.01, n = 7) that was corroborated by an increase in phosphorylated NMDA NR2B (P < 0.05, n = 4) but not NR2A subunit (P > 0.05, n = 4) expression. Both intrauterine pretreatment with capsazepine (5.02 +/- 2.11 spikes/stimulation, P < 0.01, n = 7) and an intrathecal injection of AP5 (3.21 +/- 0.83 spikes/stimulation, P < 0.01, n = 7) abolished the capsaicin-induced cross-organ sensitization and the increment in the phosphorylated NR2B level (P < 0.05, n = 4). The degrees of the cross-organ sensitization increased in a dose-dependent manner with the concentration of instilled capsaicin from 100 to 300 microM in both the proestrus and metestrus stages, whereas they weakened when the concentrations were higher than 1,000 microM. Moreover, the cross-organ sensitization caused by the uterine capsaicin instillation increased significantly in the rats during the proestrus stage when compared with the metestrus stage (P < 0.01, n = 7). These results suggest that estrogen levels might modulate the cross-organ sensitization between the uterus and the urethra and underlie the high concurrence of pelvic pain syndrome and LUT dysfunctions.     

Ability of ram introduction to induce LH secretion, estrus and ovulation in fall-born ewe lambs during anestrus.

The ability of ram introduction (RI) and progesterone pre-treatment to induce increases in LH secretion and ovulation, and the ability of progesterone pre-treatment with or without estrogen to induce estrus and ovulation in fall-born ewe lambs during seasonal anestrus was investigated. In early July, lambs of mixed breeds (41.8+/-0.6 kg and 250.7+/-1.3 days of age) were assigned to receive no treatment (C, n=7), to be introduced to rams (7:1 ewe:ram ratio; R, n=7), to be treated with progesterone (a used CIDR device) for 5 days (P, n=5), to be treated with progesterone and introduced to rams at CIDR removal (PR, n=11), or to receive the latter treatment plus an injection of estradiol benzoate (25 microg, E2beta i.m.) 24 h after CIDR withdrawal/RI (PER, n=11). Blood samples were collected from all lambs every 4h for 60 h beginning at RI/CIDR withdrawal (0 h), to characterize the LH surge profile and in groups R and C every 15 min for 8 h between 12 and 20 h for determination of LH pulse frequencies. Ultrasonographic examinations of the ovaries were conducted at 0, 36 and 60 h. In ram-exposed groups lambs were also observed for raddle marks every 4h from 0 to 60 h. The LH pulse frequency (pulses/8 h) was higher in group R (P<0.01; 7.7+/- 0.5) than group C lambs (2.7+/- 0.8). More lambs in groups exposed to rams than in the C or P groups showed an LH surge (P<0.05; 0, 100, 0, 72.7 and 100%, for C, R, P, PR and PER groups, respectively). Time from RI/CIDR removal to initiation of the LH surge was greater in lambs in the PR (43.5+/- 3.8h) than in the R (32.6+/- 4.6h; P=0.08) or PER (33+/- 1.2h; P<0.01). Diameter of the largest follicle at 0 h (3.2+/- 0.2mm) was not different among groups. Growth rate of the largest follicle between 0 and 36 h was greater (P<0.05) in RI than in C or P groups. Diameter of the largest follicle at 36 h was larger (P<0.05) in lambs in R (5.6+/- 0.2mm) and PR (5.1+/- 0.5mm) than C (4.0+/- 0.6mm) or P (3.8+/- 0.4mm) groups, and in R than PER (4.3+/- 0.4mm) treatment groups. Only lambs in the RI groups ovulated. Among RI groups the percentage of lambs ovulating was greater in the R (P<0.05; 85.7%) than PR (33.3%) groups with an intermediate response observed in lambs in treatment group PER (71.4%). The estrous response in progesterone pre-treated groups was greater (P<0.05) in lambs also treated with estrogen (PER; 81.8%), than in lambs introduced to rams alone (PR; 45.5%). In conclusion, ram introduction by itself, but not progesterone treatment alone, induces increases in LH pulse frequency, follicular development, and ovulation in fall-born ewe lambs during seasonal anestrus, further, P4 pre-treatment and RI when combined with E2 results in a high estrous response.