Effects of enclomiphene on estradiol-induced changes in LH secretion in ewes

Experiments were conducted to characterize the ability of the antiestrogen enclomiphene (ENC) to block the effects of estradiol on secretion of LH in ovariectomized ewes. To determine whether ENC could block an estradiol-induced LH surge, ewes (n = 4/group) were administered 10 to 250 mg ENC followed 30 min later by 25 micrograms estradiol. Ten or 25 mg ENC suppressed the estradiol-induced LH surge in one of four ewes, whereas 100- or 250-mg doses suppressed the LH surge in three and four of four ewes, respectively. In ewes that received a single treatment of 100 mg ENC plus 25 micrograms estradiol, serum concentrations of LH remained below 1 ng/ml for 3 wk. Compared with untreated ewes, the number of pituitary GnRH receptors was elevated (P less than .05) at 12 d and 28 d, but pituitary content of LH had decreased (P less than .05) by 28 d in ewes treated with 100 mg ENC. To determine whether ENC could block the inhibitory effects of estradiol on serum concentrations of LH, ewes received injections of .03, .1, 1 or 10 mg ENC every 4 d. Half the ewes treated with each dose also received estradiol implants. Injection of .03, .1 or 1 mg ENC alone did not affect serum concentrations of LH, whereas the 10-mg dose decreased serum concentrations of LH below 1 ng/ml by wk 1 of treatment. No dose prevented the inhibition of serum concentrations of LH caused by estradiol implants. In ovariectomized ewes, ENC was antagonistic to estradiol; it prevented the positive effects of estradiol required to induce an LH surge.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of dexamethasone, feeding time, and insulin infusion on leptin concentrations in stallions

Three experiments tested the hypotheses that daily cortisol rhythm, feeding time, and/or insulin infusion affect(s) leptin secretion in stallions. Ten mature stallions received ad libitum hay and water and were fed a grain concentrate once daily at 0700. In Exp. 1, stallions received either a single injection of dexamethasone (125 microg/kg BW i.m.; n = 5) or vehicle (controls; n = 5) at 0700 on d -1. Starting 24 h later, blood samples were collected every 2 h for 36 h via jugular venipuncture. Cortisol in control stallions varied (P < 0.01) with time, with a morning peak and evening nadir; dexamethasone suppressed (P < 0.01) cortisol concentrations. Leptin and insulin were greater (P < 0.01) in the treated stallions, as was the insulin response to feeding (P < 0.01). Leptin in control stallions varied (P < 0.01) in a diurnal pattern, peaking approximately 10 h after onset of eating. This pattern of leptin secretion was similar, although of greater magnitude (P < 0.01), in treated stallions. In Exp. 2, five stallions were fed the concentrate portion of their diet daily at 0700 and five were switched to feeding at 1900. After 14 d on these regimens, blood samples were collected every 4 h for 48 h and then twice daily for 5 d. Cortisol varied diurnally (P = 0.02) and was not altered (P = 0.21) by feeding time. Insulin and leptin increased (P < 0.01) after feeding, and the peaks in insulin and leptin were shifted 12 h by feeding at 1900. In Exp. 3, six stallions were used in two 3 x 3 Latin square experiments. Treatments were 1) normal daily meal at 0700; 2) no feed for 24 h; and 3) no feed and a bolus injection of insulin (0.4 mIU/kg BW i.v.) followed by infusion of insulin (1.2 mIU.kg BW(-1).min(-1)) for 180 min, which was gradually decreased to 0 by 240 min; sufficient glucose was infused to maintain euglycemia. Plasma insulin increased (P < 0.01) in stallions when they were meal-fed (to approximately 150 microIU/mL) or infused with insulin and glucose (to approximately 75 microIU/mL), but insulin remained low (10 microIU/mL or less) when they were not fed. The increases in insulin were paralleled by gradual increases (P < 0.01) in leptin concentrations 3 to 4 h later in stallions fed or infused with insulin and glucose. When stallions were not fed, leptin concentrations remained low. These results demonstrate that feeding time, and more specifically the insulin increase associated with a meal, not cortisol rhythm, drives the postprandial increase in plasma leptin concentrations in horses.     

Bovine C-terminal octapeptide of RFamide-related peptide-3 suppresses luteinizing hormone (LH) secretion from the pituitary as well as pulsatile LH secretion in bovines

Gonadotropin-inhibiting hormone (GnIH), observed in quail as a member of the RFamide neuropeptide family, suppresses luteinizing hormone (LH) secretion from the avian pituitary. Rats and cattle have an active gene of another member of the RFamide neuropeptide family, termed RFamide-related peptide-3 (RFRP-3), although bovine RFRP-3 is different from that of rats in both length and amino-acid sequence. A single injection of GnIH or RFRP-3 inhibited LH secretion in rodents, which continued for various periods. This study was conducted to evaluate the effects of bovine C-terminal octapeptide of RFRP-3 (RFRP-3-8) on LH secretion from cultured anterior pituitary (AP) cells of cattle, and the effects of RFRP-3-8 injections on pulsatile LH secretion in castrated male calves. The suppressive effect of RFRP-3-8 on LH secretion from AP cells was observed in the presence of gonadotropin-releasing hormone (GnRH), but not in the absence of GnRH in culture media. In another experiment collecting blood samples serially from castrated male calves with repeated intravenous injections of RFRP-3-8 (n = 6) or saline (n = 6), the RFRP-3-8 group showed suppressed LH pulse frequency during the injection period (P < 0.05); however, the RFRP-3-8 group showed no difference from the saline group in all measures of LH secretion in the postinjection period. In conclusion, our results suggested that RFRP-3-8 suppresses LH secretion from cultured AP cells, as well as LH pulse frequency in cattle.     

日粮能量对育成柴鸡外周血浆促性腺激素与垂体分泌促性腺激素的影响

采用单因素方差随机试验,选用14周龄育成柴鸡36只,随机的分成3个处理,每个处理3个重复,每个重复4只鸡,分别饲喂低能、对照、高能的日粮,试验期为28d。结果显示,育成柴鸡外周血浆FSH浓度,低能、对照与高能差异极限著（P〈0.01）,并且随着日龄增加FSH浓度逐渐增加。育成柴鸡外周血浆LH浓度第1周时低能组与对照、高能组差异极显著（P〈0.01）,对照与高能组差异不显著;第2、3周处理低能组、对照、高能组差异极显著（P〈0.01）,并且随着日龄增加浓度增加。育成柴鸡体外培养垂体细胞分泌FSH、LH高能组与低能、对照组差异极显著（P〈0.01）,而低能组与对照组差异不显著。结果表明,日粮能量通过下丘脑-垂体-性腺轴影响育成柴鸡生殖机能。     

Effects of estradiol on secretion of LH, hypothalamic function and testicular development in bull calves

Two experiments were conducted in order to determine the effects of estradiol (E₂) on the development of the hypothalamic-pituitary-testicular axis in bull calves. In experiment 1, calves were assigned randomly to one of the following groups: 1) intact, 2) intact E₂-treated, 3) castrated, or 4) castrated E₂-treated. Treatments began when the calves were 7.5 wk of age and continued for 16.5 wk. Samples of blood were collected once a week from 3 to 14 wk of age and every 10 min for 6 hr at 8, 12 and 16 wk of age. Concentrations of E₂ in plasma decreased between 3 and 4 wk of age and were further reduced by castration. Maximum concentrations of E₂ (24.3 pg/ml) were observed 72 hr after insertion of E₂ implants, however, plasma E₂ stablized at 5–9 pg/ml by 2 wk after insertion of implants. Treatment with E₂ eliminated the pulsatile secretion of LH in intact and castrated calves and retarded testicular growth. In experiment 2, calves were assigned to a control (n=4) or E₂-treated (n=6) group. Implants of E₂ were inserted at 7.5 wk of age. At 24 wk of age, calves were bled and then sacrificed to collect hypothalamic and pituitary tissues. Age-related changes in testicular weight and secretion of LH were blocked by E₂. Neither the morphology nor the intensity of immunostaining of GnRH nerve cell bodies in the preoptic area (POA) were affected by E₂. However, the density of GnRH fibers and beads in the stalk median eminence (SME), and concentrations of pituitary GnRH receptors were greater (P<.01) in E₂-treated compared to control calves. In addition, concentrations of norepinephrine (NE) in the SME were lower in E₂-treated calves when compared to controls. Based on these observations, it is concluded that administration of E₂ at 7.5 wk of age causes profound alterations in hypothalamic function including, changes in metabolism of NE and suppression of GnRH release.     

Preliminary report on the expression of leptin and leptin receptor (ObR) in normal, hyperplastic and neoplastic canine mammary tissues

Leptin and its receptor (ObR) expression were investigated by immunohistochemistry in normal, hyperplastic and neoplastic canine mammary tissues and related to clinical-pathological features. Leptin expression was detected in healthy mammary tissues, adenosis and in benign mammary tumours and was lower in ductal hyperplasias and malignant tumours. A high percentage of ObR-positive cells were present in adenosis, benign tumours and in complex carcinomas, while ObR expression was lower in healthy mammary tissues, in ductal hyperplasias and in a large part of invasive mammary carcinomas. Our data demonstrated that cancer cells expressed at low level leptin and ObR in canine mammary tumours with a more aggressive behaviour, as well as in lymph node metastases. Consequently, leptin and ObR expressions in this species resulted to be not associated with a reduced overall survival.     

Influence of abomasal infusion of glucose or partially hydrolyzed starch on pancreatic exocrine secretion in beef steers

Five crossbred steers (348 +/- 12 kg) fitted with a pancreatic pouch draining the main pancreatic duct and duodenal re-entrant and abomasal infusion cannulas were used in a 5 x 5 Latin square design to determine the influence of postruminal carbohydrate source and level on pancreatic exocrine secretion in beef steers. Abomasal infusion treatments (250 mL infused/h) were water (control), 20 g/h glucose, 40 g/h glucose, 20 g/h starch hydrolysate (SH), and 40 g/h SH. Infusion periods were 8 d with 3 to 4 d of rest between periods. Pancreatic juice was collected for 6 h on d 8 of each collection period. Every 30 min a 10% subsample was composited and frozen and the remainder was infused into the duodenum via the reentrant cannula. Abomasal infusion of glucose or SH increased (P < 0.10) total secretion of pancreatic juice and decreased (P < 0.10) secretion of alpha-amylase activity. Abomasal carbohydrate infusion did not influence total secretion of protein, trypsin activity, or chymotrypsin activity. This experiment indicates that increasing postruminal glucose or SH decreases pancreatic alpha-amylase secretion.     

Effect of pyloric blockade and infusion of histamine or pentagastrin on gastric secretion in horses

OBJECTIVE: To determine the origin of the nonacid (nonparietal) component of gastric secretions in horses induced by pentagastrin infusion. ANIMALS: 6 horses. PROCEDURE: A Latin square design was used, involving 6 horses, 3 treatments, and 2 duodenal intubation conditions (catheter with balloon to obstruct pylorus [B] or without balloon allowing movement of contents between stomach and duodenum [NB]). Each horse had an indwelling gastric cannula and a catheter positioned in the duodenum. Gastric and duodenal contents were collected during 15-minute periods. Each experiment consisted of serial collection periods: baseline; infusion of pyrilamine maleate (1 mg/kg of body weight, IV); not treated; and IV infusion of saline (0.9% NaCl) solution alone, saline solution containing pentagastrin (6 microg/kg x h), or saline solution containing histamine (30 microg/kg x h). Volume of samples was recorded, and electrolyte concentrations were measured. RESULTS: Pentagastrin and histamine stimulated maximal acid output; however, during NB conditions, pentagastrin-induced concentration of hydrogen ions was significantly less than during histamine or pentagastrin infusions during B conditions. The large volume produced in response to pentagastrin during NB conditions was accompanied by increased sodium ion output that was greater than for pentagastrin during B conditions, but both values were significantly greater than values for histamine during B or NB conditions. CONCLUSIONS AND CLINICAL RELEVANCE: Nonparietal secretions collected during IV infusion of pentagastrin are duodenal in origin. Reflux of duodenal contents into the stomach of horses is enhanced by pentagastrin. Flow of duodenal contents into the stomach could have implications in the pathogenesis of ulcers in horses.     

日粮能量对乌鸡外周血清促性腺激素与垂体分泌促性腺激素的影响

采用体外细胞培养和放射免疫测定法（RIA）测定日粮能量对鸟鸡外周血清促性腺激素与垂体分泌促性腺激素的影响。日粮能量处理：低能Ⅱ组、低能I组、对照组、高能I组、高能Ⅱ组;体外培养垂体细胞组分：高能组、对照纽、低能组。结果显示,血清中FSH含量,对照组与高能Ⅱ组相比差异显著（P〈0．05）,与低能I、Ⅱ组相比差异极显著（P〈0．01）;血清中LH含量,对照组与高能I组、高能Ⅱ组组间差异显著（P〈0．05）;低能Ⅱ组与高能I组、低能Ⅱ组与高能Ⅱ组、低能I组与高能I组、低能I组与高能Ⅱ组组间差异极显著（P〈0．01）;单层垂体细胞中FSH含量,高能组和对照组与低能组比差异显著（P〈0．05）;单层垂体细胞中LH含量,高能组与低能组比较差异极显著（P〈0．01）,对照组与低能组比较差异不显著。结果表明,日粮能量对处理体外培养乌鸡垂体细胞分泌FSH、LH有促进作用。     

三价铬对肥育猪生长激素分泌及垂体mRNA表达的影响

将96头体质量约65 kg的杜长大三元杂交猪,按体质量相近、公母各半的原则随机分成4组,每组3个重复.试验猪1组设为对照,饲喂基础饲粮,其余3组为试验组,分别饲喂在基础饲粮基础上添加来源于氯化铬、吡啶羧酸铬,纳米铬的含200 μg/kg铬试验饲粮.试验猪充分饲喂,自由饮水,试验为期40 d.饲养试验结束前1 d,从每组中选择6头猪,间隔15 min颈静脉采血1次,连续3 h,制备血清用于生长激素动态分泌模式研究.饲养试验结束后,按体质量相近原则从每组中选择8头猪进行屠宰,测定背膘厚和背最长肌面积;并各采取3头猪的脑垂体,RT-PCR法测定生长激素mRNA水平.结果表明,饲粮中添加200μg/kg三价纳米铬使肥育猪日增重提高了6.31%(P<0.05),料重比降低了4.61%(P<0.05),背膘厚降低了24.32%(P<0.05),背最长肌面积提高了20.22%(P<0.05).生长激素动态模式分析结果显示,纳米铬组试验猪生长激素总体水平、最低值、峰值和峰持续时间分别提高了42.62%(P<0.05)、87.94%(P<0.05)、26.60%(P<0.05)和17.19%(P<0.05),吡啶羧酸铬组试验猪生长激素总体水平、峰值分别提高了36.58%(P<0.05)和27.18%(P<0.05).垂体生长激素基因RT-PCR分析结果显示,纳米铬组试验猪垂体生长激素mRNA水平提高了27.63%(P<0.05).研究结果提示,三价纳米铬可提高肥育猪垂体生长激素mRNA水平,促进机体生长激素分泌,从而促进生长和改善胴体特性.     

Expression of the cannabinoid receptor type 1 in the pituitary of rabbits and its role in the control of LH secretion

The aim of this study was to elucidate the possible direct regulatory role of the endocannabinoids in the modulation of LH secretion in rabbits, a reflex ovulator species. The cannabinoid receptor type 1 (CB1) was characterized by RT-PCR techniques in the anterior pituitary of intact and ovariectomized does treated with GnRH and primed with estrogen and CB1 antagonist, rimonabant. Cannabinoid receptor type 1 immune reaction was evidenced by immunohistochemistry in the cytoplasm of approximately 10% of the pituitary cells with a density of 8.5 ± 1.9 (per 0.01 mm²), both periodic acid–Schiff positive (30%) and negative (70%). All CB1-immunoreactive cells were also immune reactive for estrogen receptor type 1. Ovariectomy, either alone or combined with estrogen priming, did not modify the relative abundances of pituitary CB1 mRNA, but decreased (P < 0.01) the expression of estrogen receptor type 1 mRNA. Treatment with CB1 antagonist (rimonabant) inhibited (P < 0.01) LH secretory capacity by the pituitary after GnRH injection, and estrogen priming had no effect. The present findings indicate that the endocannabinoid system is a potential candidate for the regulation of the hypothalamic-pituitary-ovarian axis in reflex ovulatory species.     

Effect of GnRH and its antagonist (Antarelix) on LH release from cultured bovine anterior pituitary cells

In the following investigations, the LH secretion of cells from pituitaries in heifers on days 16-18 of their oestrous cycle (n = 14) was analysed. Cells were dissociated with trypsin and collagenase and maintained in a static culture system. For the estimation of LH release, the cells were incubated with various concentrations of mammalian GnRH (Lutrelef) for 6 h. To determine the action of Antarelix (GnRH antagonist), the cells were preincubated for 1 h with concentrations of 10(-5) or 10(-4) M Antarelix followed by 10(-6) M GnRH coincubation for a further 6 h. At the end of each incubation, the medium was collected for LH analysis. Parallel, intracellular LH was qualitatively detected by immunocytochemistry. Changes in the intensity of LH staining within the cells in dependence of different GnRH concentrations were not observed, but a significant increase LH secretion in pituitary cells was measured at 10(-6) M GnRH. Antarelix had no effect on basal LH secretion at concentrations of 10(-4) and 10(-5) M. After coincubation of pituitary cells with Antarelix and GnRH, Antarelix blocked the GnRH-stimulated LH secretion with a maximal effect of 10(-4) M, but the staining of immunoreactive intracellular LH was detected at approximately the same level compared to the pituitary cells treated with exogenous GnRH alone. These data demonstrate that Antarelix is effective in influencing the GnRH-stimulated LH secretion of pituitary cells in vitro. After administration of Antarelix in vivo, the GnRH-stimulated LH secretion of cultured pituitary cells was not inhibited.     

Effect of the preovulatory LH surge on bovine follicular progesterone receptor mRNA expression

A growing body of evidence indicates that intrafollicular progesterone receptor signaling pathways are obligatory for follicle rupture. However, the intrafollicular localization and regulation of progesterone receptor expression during the periovulatory period in cattle are not known. In this study, we determined the effect of the preovulatory gonadotropin surge on localization and expression of progesterone receptor mRNA in bovine periovulatory follicular and luteal tissue. Ovaries containing preovulatory follicles or new corpora lutea (CL) were collected at approximately 0, 6, 12, 18, 24 (preovulatory follicles) and 48 h (CL) after a GnRH-induced LH surge (n=5-8 per timepoint). Expression of progesterone receptor mRNA was detected in periovulatory follicular and luteal tissue at all timepoints examined. Relative levels of progesterone receptor mRNA were dramatically upregulated within 6h after the LH surge compared to all other time points (P<0.0001). In situ hybridization analysis revealed that the significant increase in progesterone receptor mRNA expression was localized to the granulosal layer of preovulatory follicles. Our results indicate that progesterone receptor mRNA expression is upregulated specifically in the granulosal layer of bovine preovulatory follicles following the LH surge. Progesterone receptor signaling pathways may help mediate the effects of the preovulatory LH surge on follicle rupture in cattle.     

Basal and GnRH induced secretion of FSH and LH in ovariectomized ewes treated with bovine follicular fluid

Ovariectomized (OVX) ewes were injected with 5 ml of either bovine serum, charcoalextracted bovine follicular fluid (FF), or whole bovine FF. Five hours after this pretreatment, ewes on each pretreatment were injected with either 0, 1, or 5 μg of GnRH. Ewes that were pretreated with either type of FF had decreased concentrations of FSH regardless of dose of GnRH when compared to ewes pretreated with bovine serum. There was no effect of charcoal extraction. There were no differences among the pretreatment groups in LH response to GnRH. In a second experiment, OVX ewes were pretreated (4 ml) with either bovine serum or bovine FF 5 hr prior to GnRH or with bovine FF 42, 30 and 18 hr prior to GnRH. Ewes were injected with either 0 or 5 μg of GnRH. Pretreatment with FF for 5 or 42 hr prior to GnRH resulted in significantly decreased concentrations of FSH both at the time of GnRH treatment and during the following 2 hr. Concentrations of LH did not differ among pretreatment groups. In a third experiment, OVX ewes were pretreated with either bovine serum or bovine FF 30, 18 and 5 hr prior to GnRH. Ewes were injected with either 0, 5 or 50 μg of GnRH. Pretreatment with FF resulted in decreased concentrations of FSH both at the time of GnRH treatment and during the following 2 hr. Concentrations of LH were also decreased at the time of GnRH treatment.     

Effect of epinephrine, norepinephrine and(or) GnRH on serum LH in prepuberal beef heifers

Forty prepuberal Simmental X Brahman-Hereford heifers were utilized to determine the effects of epinephrine (E), norepinephrine (NE), gonadotropin releasing hormone (GnRH) or combinations of GnRH + E and GnRH + NE on serum luteinizing hormone (LH) concentrations. Animals were assigned randomly to one of five treatments with four replicates/treatment. Treatments consisted of I) 100 micrograms GnRH at time 0 (n = 8); II) 50 mg NE at time -15 and 0 (n = 8); III) 50 mg E at time -15 and 0 (n = 8); IV) 100 micrograms GnRH at time 0, plus 50 mg NE at time -15 and 0 (n = 8) and V) 100 micrograms GnRH at time 0, plus 50 mg E at time -15 and 0 (n = 8). All treatment compounds were administered im in 2 ml physiological saline and blood samples were collected via tail vessel puncture at -30, -15, 0, 15, 30, 45, 60, 90, 120, 180, 240, 300 and 360 min from GnRH injection. Treatment with NE or E alone had no effect (P greater than .10) on serum LH during the sampling period. The initial LH release to GnRH was altered (P less than .05) by concomitant treatment with NE (treatment IV) or E (treatment V). Magnitude of the LH release was reduced (P less than .01) by treatment V. Area under the LH surge was reduced (P less than .05) by treatment IV (NE) and V (E).