Testosterone antagonist (flutamide) blocks ovulation and preovulatory surges of progesterone, luteinizing hormone and oestradiol in laying hens

The preovulatory release of luteinizing hormone (LH) in the domestic hen occurs after the initiation of a preovulatory surge of testosterone. The objective of this study was to determine whether this testosterone surge has functional significance in the endocrine control of ovulation. Groups of laying hens (n = 10-22) were treated with the androgen receptor antagonist, flutamide, at 8 h intervals for 24 h at doses of 0, 31.25, 62.5, 125 and 250 mg. All doses reduced egg laying (P < 0.001), with the highest dose being the most effective. In a second study, laying hens (n = 9) were treated with 250 mg flutamide at 8 h intervals for 24 h with a control group being given placebo (n = 10). Blood samples were taken for hormone measurements at 2 h intervals for 18 h starting 4 h before the onset of darkness. The percentage of hens laying per day did not differ between groups before treatment (control, 88% vs flutamide, 86%). Ovulation was blocked in all hens treated with flutamide within 2 days while the control hens continued to lay at the pretreatment rate (80%). Preovulatory surges of plasma testosterone, progesterone, oestradiol and LH were observed in control hens but with the exception of testosterone, flutamide treatment blocked the progesterone, oestradiol and LH surges. LH concentrations declined progressively with time in the flutamide-treated hens. It is concluded that inhibition of testosterone action blocks egg laying and the preovulatory surges of progesterone, luteinizing hormone and oestradiol demonstrating a key role for the preovulatory release of testosterone in the endocrine control of ovulation in the domestic hen.     

Effect of elevating luteinizing hormone action using low doses of human chorionic gonadotropin on double ovulation,follicle dynamics,and circulating follicle-stimulating hormone in lactating dairy cows

Double ovulation and twin pregnancy are undesirable traits in dairy cattle. Based on previous physiological observations, we tested the hypothesis that increased LH action [low-dose human chorionic gonadotropin (hCG)] before the expected time of diameter deviation would change circulating FSH concentrations, maximum size of the second largest (F2) and third largest (F3) follicles, and frequency of multiple ovulations in lactating dairy cows with minimal progesterone (P4) concentrations. In replicate 1, multiparous, nonbred lactating Holstein dairy cows (n = 18) had ovulation synchronized. On d 5 after ovulation, all cows had their corpus luteum regressed and were submitted to follicle (≥3 mm) aspiration 24 h later to induce emergence of a new follicular wave. Cows were then randomized to NoP4 (untreated) and NoP4+hCG (100 IU of hCG every 24 h for 4 d after follicle aspiration). Ultrasound evaluations and blood sample collections were performed every 12 h for 7 d after follicle aspiration. All cows were then treated with 200 μg of GnRH to induce ovulation. In replicate 2, cows (n = 16) were resubmitted to similar procedures (i.e., corpus luteum regression, follicle aspiration, randomization, ultrasound evaluations every 12 h, GnRH 7 d after aspiration). However, cows in replicate 2 received an intravaginal P4 device that had been previously used (～18 d). Only cows with single (n = 15) and double (n = 16) ovulations were used in the analysis. No significant differences were detected for frequency of double ovulation, follicle sizes, and FSH concentrations across replicates (NoP4 vs. LowP4 and NoP4+hCG vs. LowP4+hCG), so data were combined. Double ovulation was 40% for control cows with no hCG (CONT) and 62.5% with hCG (hCG). Double ovulation increased as the maximum size of F2 increased: <9.5 mm and 9.5–11.5 mm (7.7%) and ≥11.5 mm (94.1%). The hCG group had more cows with F2 > 11.5 (69%) than with 9.5 ≥ F2 ≤ 11.5 (25%) and F2 < 9.5 (6%). In agreement, F2 and F3 maximum size were larger in the hCG group, but FSH concentrations were lower after F1 > 8.5 mm compared with CONT. In contrast, FSH concentrations were greater before deviation (F1 closest value to 8.5 mm) in cows with double ovulations than in those with single ovulations, regardless of hCG treatment. In addition, time from aspiration to deviation was shorter in cows with double rather than single ovulation and in cows treated with hCG as a result of faster F1, F2, and F3 growth rates before diameter deviation. In conclusion, greater FSH and follicle growth before deviation seems to be a primary driver of greater frequency of double ovulation in lactating cows with low circulating P4. Moreover, the increase in follicle growth before deviation and in the maximum size of F2 during hCG treatment suggests that increased LH may also have a role in stimulating double ovulation.     

Oxytetracycline in bovine plasma, milk, and urine after intrauterine administration.

Six healthy lactating cows with normal but varying stages of estrous cycles were treated with chelated or nonchelated oxytetracycline by intrauterine infusion, 4 mg/kg body weight. The chelated drug was a mixture of special metal halides and oxytetracycline while the nonchelated form was a commercial oxytetracycline solution suitable for intramuscular or intravenous administration. Oxytetracycline concentrations in plasma, milk, and urine were determined at timed intervals following treatment. Both forms of the drug appeared in the plasma, but the chelated form was a small amount. The nonchelated oxytetracycline also appeared in the milk. However, no oxytetracycline was in milk of cows treated with chelated oxytetracycline. Both forms of the drug were excreted in the urine. Milk should be withheld from market 24 and 0 h after administration of nonchelated and chelated forms.     

Response of bovine serum prolactin and growth hormone to duodenal, abomasal, and oral administration of thyrotropin-releasing hormone.

Thyrotropin-releasing hormone was injected into the duodenum of two 500-kg steers, placed into the abomasum of two prepubertal bulls, and fed to four bull calves (1 to 3 wk of age) to test the effect on concentrations of prolactin and growth hormone in blood serum. Before 20 and 200 mg of thyrotropin-releasing hormone were injected into the duodenum, prolactin in serum averaged 7.5 and 9.4 ng/ml and increased to 52.5 and 129.6 ng/ml at 45 and 35 min after treatment. Average growth hormone concentration of serum was increased also, but the response was more variable than prolactin. Peak concentrations of prolactin and growth hormone in blood serum were 5 to 10 times greater after treatment with thyrotropin-releasing hormone (40 mg/100 kg body weight into abomasum) than before treatment. Within 30 min after oral administration of thyrotropin-releasing hormone (0, .5, 1, and 2 mg/kg body weight) growth hormone concentration of serum was 30, 306, 356, and 317% greater than pretreatment. Prolactin concentration of serum, however, was increased in only one calf.     

Effect of feeding calcium soaps to early postpartum dairy cows on plasma prostaglandin F2 alpha, luteinizing hormone, and follicular growth

Multiparous Holstein cows (n = 18) were fed a total mixed ration containing corn silage, corn grain, whole cottonseed, soybean meal, dried distillers grains, and chopped bermudagrass hay (control) or same diet plus Ca salts of long-chain fatty acids (2.2% of diet DM) for the first 60 d postpartum. Predicted energy balance was calculated from DM intake, milk yield and composition, and BW. On d 25 postpartum, cows were injected with 25 mg of prostaglandin F2 alpha and treated for 15 d with an intravaginal device containing 1.9 g progesterone. Profiles of 15-keto-13,14-dihydro-prostaglandin F2 alpha (d 1 to 21) and plasma triglycerides (d 7 to 60) were similar between groups. Average number of follicles, determined by ultrasonography prior to d 25, tended to differ between groups; controls had more 3- to 5-mm and fewer 6- to 9-mm follicles than the group of fat-fed cows. Basal, smoothed mean concentration, and average luteinizing hormone amplitude, determined by 10-min samples for 8 h on d 10, were not significantly different between groups. Increasing predicted energy balance was associated with increased pulse amplitude and diameter of the largest follicle on d 10. During the progesterone treatment period and the postprogesterone treatment estrous cycle, cows fed fat had greater numbers of 3- to 5-mm and greater than 15-mm follicles. In conclusion, feeding fat did not influence 15-keto-13,14-dihydro-prostaglandin F2 alpha or luteinizing hormone dynamics but did alter the average number of follicles within different size classes and the diameter of largest and second largest follicle after progesterone treatment.     

Use of 1 alpha-hydroxyvitamin D3 in prevention of bovine parturient paresis. 8. Maternal and neonatal plasma calcium, parathyroid hormone, and vitamin D metabolites concentrations

Thirteen Israeli Friesian cows (3.71 average calvings) in the second or later lactation, fed a daily diet containing 90 g of Ca and 50 g of P, were injected once intramuscularly with 700 micrograms 1 alpha-hydroxy-vitamin D3 in order to investigate its placental transfer and its subsequent metabolism in the neonate. The injection of the vitamin 96 to 24 h before calving slightly increased plasma Ca at parturition, whereas uninjected controls displayed a prominent hypo-calcemia. On the 10th and 20th d after calving, difference in the plasma Ca concentration of the two groups was not significant. At parturition, plasma parathyroid hormone concentration was significantly higher and plasma 1,25-dihydroxyvitamin D lower in the control than in the treated cows. At parturition the plasma concentrations of Ca, parathyroid hormone, hydroxyproline, and 24,25-hydroxyvitamin D were higher in the calves than in their dams. Plasma concentrations of 25-hydroxyvitamin D were markedly higher and 1,25-hydroxyvitamin D was slightly higher in cows than in their offsprings.     

The role of angiotensin II in the early stages of bovine ovulation

There is evidence that the renin-angiotensin system plays an important role in ovulation in cattle. Using an in vivo model, we investigated the role of angiotensin (Ang) II in bovine ovulation by injecting Ang II receptor antagonists into ovulatory follicles. Animals (n = 102) were pre-synchronized and, when the follicles reached 12 mm, they were given the respective treatment and the cows received GnRH agonist (i.m.) to induce ovulation. The ovulation rate was significantly lower when 100 mu M saralasin (Ang II receptor antagonist) was intrafollicularly injected (14.3%) in comparison with saline solution (83.3%). Based on these results, a second experiment was carried out to determine the timing of Ang II's critical role in ovulation. Saralasin inhibited ovulation only when applied at 0 and 6 h (16.7 and 42.9% ovulation rate in the 0- and 6-h groups respectively), but not at 12 h (100%) following GnRH agonist treatment. To investigate the subtypes of Ang II receptors implicated in the LH-induced ovulation, losartan (LO; AT(1)-Ang II receptor antagonist), PD123 319 (AT(2)-Ang II receptor antagonist), LO+PD123 319, or saline were intrafollicularly injected when the cows were challenged with GnRH agonist. Ovulation was inhibited by PD123 319 and LO+PD123 319 (50.0 and 33.3% on ovulation rate respectively), but not by LO or saline solution (100% ovulation in both groups). From these results, we suggest that Ang II plays a pivotal role in the early mechanism of bovine ovulation via the AT(2) receptor subtype.     

Effects of cyclic steroid hormone replacement on prolactin and luteinizing hormone surges in female rats

The ability of steroid hormones to produce an LH or prolactin (PRL) surge was determined in rats ovariectomized at 6, 9 or 13 weeks of age and subjected to one, three or six cycles of estrogen and progesterone replacement. Sensitivity to steroid replacement was dependent on the age of the animal at the time of ovariectomy. Repeated cyclic steroid hormone replacement significantly increased the magnitude of the PRL response, but not the LH response, in animals ovariectomized at 6 weeks. The LH response was significantly altered by cyclic steroid replacement only in animals ovariectomized at 13 weeks. These results indicate that the mechanisms involved in the regulation of PRL secretion are influenced by steroid hormone replacement and that cyclic steroid hormone exposure increases the magnitude of the PRL secretory response.     

Immunoglobulins,growth factors and growth hormone in bovine colostrum and the effects of processing

In colostrum collected 0–80 h postpartum the contents of immunoglobulins (Igs), transforming growth factor beta-2 (TGF-β2), insulin-like growth factor-1 (IGF-1) and growth hormone (GH) were analysed. Colostrum initially contained 90 mg mL⁻¹ IgG1, 2.8 mg mL⁻¹ IgG2, 1.6 mg mL⁻¹ IgA, 4.5 mg mL⁻¹ IgM, and these concentrations declined by 92%, 87%, 93% and 84%, respectively, in the samples collected later. Of the growth factors, colostrum initially contained 289–310 ng mL⁻¹ TGF-β2 and the concentration diminished to 66 ng mL⁻¹. The content of IGF-1 and GH postpartum decreased from 870 to 150 ng mL⁻¹, and from 0.17 to <0.03 ng mL⁻¹, respectively. Heat treatment and freeze-drying of colostral whey decreased the content of Igs to 75%, while the contents of IGF-1 and TGF-β2 were unaffected. A similar processing, including filtration steps reduced also the IGF-1 and TGF-β2 by 25%. IgM seems to be the most sensitive of the Igs to processing.     

Effect of monopropylene glycol on luteinizing hormone, metabolites, and postpartum anovulatory intervals in primiparous dairy cows

This study examined the effect of monopropylene glycol (MPG) supplementation on LH secretion, postpartum interval to first ovulation, and milk production in heifers calving with poor body condition score (BCS). Forty-seven heifers were allocated to 3 treatments: 1) heifers with high BCS (BCH; n = 13) that calved at a BCS of 3.4 (BCS scale of 1 to 5); 2) heifers with low BCS (BCL; n = 17) that calved at a BCS of 2.8; and 3) heifers with low BCS that calved at a BCS of 2.8 and were assigned to receive MPG supplementation (BCL + MPG; n = 17) and grazed pasture ad libitum. Monopropylene glycol was drenched (250 mL) twice daily for 16 wk after calving. Patterns of change in plasma LH were measured at 2 and 5 wk after calving. Pulsatile release of LH at 2 and 5 wk was greater in BCL + MPG and BCH cows compared with the BCL control cows. The BCL + MPG cows had lower NEFA concentrations than did the BCL cows during wk 1 to 6 after calving. At 12 wk postpartum, the proportion of cows cycling was 77, 82, and 28% for the BCH, BCL + MPG, and BCL treatments, respectively. Mean milk fat yield was greater for the BCH treatment during the first 12 wk postpartum compared with the BCL + MPG or BCL treatments, which did not differ from each other. Results of this study indicate that MPG supplementation reduced the interval from calving to first ovulation in heifers having poor body condition at calving.     

Derivation of an aquatic predicted no-effect concentration for the synthetic hormone, 17 alpha-ethinyl estradiol

17alpha-Ethinyl estradiol (EE2) is a synthetic estrogen widely used in combination with other steroid hormones in oral contraceptives and in the contraceptive patch. EE2 has been detected in sewage treatment plant effluents in the low nanogram -per-liter range and occasionally in surface waters in the U.S., U.K., Canada, Brazil, Germany, and elsewhere. The mode of action is receptor-mediated, and estrogen receptors exist in mammals and other vertebrates. A large number of studies on the effects of EE2 on aquatic organisms exist. One hundred English language studies published between 1994 and 2007, one as yet unpublished study, and findings published in conference proceedings (in German) were compared to published data quality criteria to identify the most relevant studies for deriving a predicted no-effect concentration (PNEC). Reproduction in fish was identified as the most sensitive end point in aquatic species. A species sensitivity distribution was constructed using no observed effect concentrations (NOECs) for reproductive effects from 39 papers in 26 species, resulting in a median hazardous concentration at which 5% of the species tested are affected (HC5,50) of 0.35 ng/L. After comparing this HC5,50 to all of the laboratory and field-derived toxicity information available for EE2, we recommend using 0.35 ng/L as the PNEC for EE2 in surface water. This PNEC is below 95% of the existing NOECs for effects on reproduction and is also below virtually all of the NOECs for vitellogenin induction in the key fish reproduction studies.     

Luteinizing hormone and testosterone concentrations in plasma of bull calves treated with gonadotropin releasing hormone.

Twelve bull calves, 2 to 24 days of age, were assigned to two groups of six calves and administered a single 50 microng intramuscular injection of gonadotropin releasing hormone. Bull calves in group 1 were 2 to 5 days of age (mean 3.0 days), and bull calves in group 2 were 10 to 24 days of age (mean 17.1 days). Blood plasma for radioimmunoassay of luteinizing hormone and testosterone was collected at .5-h intervals for 3-h and at 4- and 6-h following treatment. With increasing age, preinjection luteinizing hormone concentrations in plasma decreased while preinjection testosterone concentrations increased. Mean preinjection luteinizing hormone concentrations for group 1 (1.01 +/- .07 ng/ml; mean +/- standard error) were higher than for group 2 (.61 +/- .12 ng/ml). Preinjection luteinizing hormone and testosterone concentrations were correlated--.63. Following treatment, luteinizing hormone increased with mean peak concentrations of 7.28 +/- .74 and 6.95 +/- 1.79 ng/ml for groups 1 and 2. Peak testosterone response following gonadotropin releasing hormone was higher for group 2 than 1 (1077 +/- 253 versus 436 +/- 116 pg/ml).     

Enhanced precision of estrus and luteinizing hormone after progesterone and prostaglandin in heifers

Objectives were to determine if low progesterone and increased basal luteinizing hormone for 3 d after injecting prostaglandin F2 alpha would affect precision of preovulatory surges of luteinizing hormone and onset of estrus in Holstein heifers. In experiments 1 and 2, heifers received progesterone devices for 10 d and prostaglandin F2 alpha on the 7th d after inserting progesterone device. For controls, progesterone devices were removed when prostaglandin F2 alpha was injected, and control devices were installed for 3 d. In experiment 1, jugular blood was sampled every 2 or 4 h from injection of prostaglandin F2 alpha until 108 h after removing progesterone to determine basal and peak secretion of luteinizing hormone. In treatment heifers, basal luteinizing hormone tended to increase, and intervals to peak secretion of luteinizing hormone were shorter and more precise than for controls. In experiment 2, heifers were observed for estrus for 30 min every 4 h for 96 h after removing progesterone devices. Intervals to onset of estrus for experimental heifers were shorter and more precise than for controls. After exogenous prostaglandin F2 alpha, 1 to 2 ng of progesterone per milliliter of serum for 2 to 3 d allows increased basal luteinizing hormone and is associated with increased precision of preovulatory secretion of luteinizing hormone and onset of estrus. This may be an acceptable method to control ovulation precisely for fixed time insemination of cattle.     

Effects of induced parturition and estradiol on feed intake, liver triglyceride concentration, and plasma metabolites of transition dairy cows.

The effect of induced parturition and estradiol on feed intake, liver triglyceride, plasma metabolites, and milk yield was evaluated in fifty-six Holstein cows and heifers. Cows were assigned to treatments on d 260 of gestation and were on trial until d 10 postpartum for measurement of dry matter intake (DMI), plasma metabolites, and liver triglyceride and until d 31 postpartum to measure milk yield. Fourteen animals per group (9 cows and 5 heifers) received either a placebo, 1 mg of fenprostalene, 50 mg of estradiol-17 beta benzoate, or both on d 276 of gestation. Cows that received fenprostalene consumed more dry matter (DM) for the last 8 d prepartum than did cows that did not receive fenprostalene (9.6 kg/d vs. 8.5 kg/d, respectively) but consumed less DM for the first 10 d postpartum (10.9 kg/d vs. 13.1 kg/d, respectively). Cows injected with estradiol-17 beta benzoate tended to consume less DM postpartum than did cows not injected with estradiol-17 beta benzoate (11.3 kg/d vs. 12.7 kg/d, respectively). There was no effect of treatment on milk yield; however, a fenprostalene by day interaction resulted from lower milk yield on d 3, 4, 5, 7, and 10 relative to calving in cows that received fenprostalene. Administration of fenprostalene resulted in a delay in the peak plasma nonesterified fatty acid (NEFA) concentration until 2 d after calving. Plasma glucose concentrations were greatest 1 d prior to calving for cows that received fenprostalene, whereas plasma glucose concentrations peaked on the day of calving for cows that did not receive fenprostalene. Liver triglyceride increased over time; however, there was no effect of treatment on liver triglyceride. Calving induction improved DMI for the last 8 d prepartum, but a concomitant decrease in liver triglyceride after calving did not result. Estradiol-17 beta benzoate had no effect on plasma metabolites or liver triglyceride, indicating that the physiological rise in estradiol prior to calving does not have a primary role in lipolysis or hepatic fatty acid metabolism in the dairy cow.     

Gonadotropin-releasing hormone and prostaglandin F2 alpha for postpartum dairy cows: estrous, ovulation, and fertility traits

A study of 234 Holstein cows was conducted to determine if early postpartum treatments of gonadotropin-releasing hormone, prostaglandin F2 alpha, or both would alter frequency and occurrence of estrus and ovulation as well as subsequent fertility. Cows in groups 1 and 2 received gonadotropin-releasing hormone (200 micrograms) between 10 and 14 d postpartum, and cows in groups 3 and 4 received saline. Ten days later, cows in groups 2 and 3 received prostaglandin F2 alpha (25 mg), and cows in groups 1 and 4 received saline. Treatment with gonadotropin-releasing hormone reduced intervals to first ovulation and first detected estrus as well as increasing the proportion of cows with three or more ovulations before first service from 57% for saline-treated controls to 83%. Treatment with prostaglandin F2 alpha reduced intervals to second and third ovulation and shortened the first estrous cycle. Treatments for cows in groups 1 and 3 increased the proportion of cows having estrous cycles of normal duration. Interval from calving to conception was reduced by 43 to 48 d for cows with an abnormal puerperium treated with either gonadotropin-releasing hormone or prostaglandin F2 alpha compared with controls (group 4) and by 27 to 29 d overall in all cows in groups 1 and 3. Cows with normal or abnormal puerperium in groups 1 and 3 required 26 to 41% fewer services per conception than controls. We conclude that treatments of gonadotropin releasing hormone or prostaglandin F2 alpha, but not the treatment combination, improved fertility of dairy cows, especially those that experienced puerperal problems.     

The role of neuropeptide Y and interaction with leptin in regulating feed intake and luteinizing hormone and growth hormone secretion in the pig

Two experiments (EXP) were conducted in ovariectomized prepubertal gilts to test the hypothesis that neuropeptide Y (NPY) stimulates appetite and modulates LH and GH secretion, and that leptin modifies such acute effects of NPY on feeding behavior and LH and GH secretion. In EXP I, gilts received intracerebroventricular (ICV) injections of 0.9% saline (saline; n = 6), or 10 microg (n = 7), 50 microg (n = 5) or 100 microg (n = 7) NPY in saline and blood samples were collected. In EXP II, gilts received ICV injections of S (n = 4), or 50 microg leptin (n = 4), or 100 microg NPY (n = 4) or 100 microg NPY +50 microg leptin (n = 4) in saline, and feed intake was measured at 4, 20 and 44 h after feed presentation and blood samples collected. In EXP I, NPY suppressed LH secretion and the 100 microg dose stimulated GH secretion. In EXP II, NPY reversed the inhibitory effect of leptin on feed intake and suppressed LH secretion, but serum GH concentrations were unaffected. These results support the hypothesis that NPY modulates feed intake, and LH and GH secretion and may serve as a neural link between metabolic state and the reproductive and growth axis in the pig.