Rapid down regulation of pyroglutamyl peptidase II activity by arachidonic acid in primary cultures of adenohypophyseal cells

Thyrotropin releasing hormone (TRH; pglu-his-proNH2) is inactivated, in the extracellular space, by pyroglutamyl aminopeptidase II (PPII), a narrow specificity ectopeptidase. In adenohypophysis, multiple hormones regulate PPII surface activity. The intracellular pathways of regulation are still poorly understood. Since some of the neurohormones which regulate PPII activity, including TRH and dopamine, transduce in part their effect through modulation of arachidonic acid (AA) mobilization, we have tested its role in regulation of PPII activity in primary cultures of rat adenohypophyseal cells. Melittin concentrations from 0.25 to 1 ug/ml induced a rapid decrease of PPII activity; 0.5 ug/ml caused a maximum effect (38-45% inhibition) at 20-30 min. AA (0.5 or 5 uM) also inhibited PPII activity (42-72%, maximum at 20 min); AA effect was reversible, with values approaching control at 1 h. The inhibitory effect of AA was blocked by lipoxygenase (10 uM nordihidroguaiaretic acid) but not ciclooxygenase inhibitors (10 uM indomethacin) suggesting the involvement of the lipoxygenase pathway. These data show that production of arachidonic acid by adenohypophyseal cells can rapidly but transiently down regulate surface PPII activity. This is the first evidence that AA mobilization can regulate the activity of an ectopeptidase.     

Localization of TRH-sensitive sites in rat brain mediating intestinal transit

Stereotaxic microinjection of thyrotropin releasing hormone (TRH) into 16 brain areas revealed that only three sites, the medial septum and the lateral and anterior hypothalami, were sensitive to a 1.0 ug dose in stimulating intestinal transit in anesthetized rats. The medial septum and anterior hypothalamus also responded to 0.1 ug, but not to 0.01 ug, of TRH. Because TRH and its receptors are distributed in these brain areas, the present results suggest a possible role for this peptide in the central regulation of gastrointestinal activity.     

Influence of synthetic thyrotropin-releasing hormone tartrate monohydrate on plasma gonadotropin concentration of normal males.

Synthetic thyrotropin-releasing hormone (TRH) tartrate monohydrate was administered by rapid intravenous injection to nine normal males. Plasma thyroid-stimulating hormone (TSH), luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were measured before and at selected periods after TRH injection. The mean plasma TSH value immediately prior to TRH injection was 3.5 muU/ml and the level 15 min after injection was 14.8 muU/ml. The mean plasma LH value immediately prior to TRH injection was 8.0 mIU/ml and the level 15 min after injection was 15.0 mIU/ml. The latter elevation was statistically significant (p less than 0.01), although it was just above the upper normal range. The mean plasma FSH value immediately prior to TRH injecion was 7.7 mIU/ml, and a significant difference was not observed after TRH administration. These results revealed that synthetic TRH tartrate monohydrate influenced the release of LH from the anterior pituitary.     

Evaluation of thyrotropin-releasing hormone as a potential antidepressant agent in the conscious dog

Results from preliminary clinical reports have indicated that thyrotropin-releasing hormone (TRH) produces improvement in depressed patients. In the present study, doses of TRH at least 25 times greater than those reported as clinically effective on a mg/kg basis were evaluated for antidepressant activity in the conscious dog. Clinically effective antidepressants, such as MAO inhibitors and tricyclics like imipramine, potentiate certain behavioral, autonomic, and cardiovascular responses produced by the indole alkaloid yohimbine, whereas general CNS stimulants such as amphetamine or cocaine do not potentiate these responses. Both 50 and 100 ug/kg doses of TRH failed to potentiate yohimbine effects. Certain gross similarities in effects produced by TRH and amphetamine observed in this study support the view that beneficial effects of TRH in depression may be related to general sympathetic activation produced by this hormone.     

A new antibiotic combination for frozen bovine semen, 3. Evaluation of fertility

Field fertility (nonreturn rate) studies were performed independently by three artificial insemination organizations to evaluate bovine semen processed for freezing using the antibiotics gentamicin, tylosin and Linco-Spectin at concentrations of 500 ug, 100 ug, and 300/600 ug, respectively, per milliliter of neat semen and per milliliter of nonglycerol portion of the extender. The antibiotic combination including penicillin, dihydrostreptomycin, polymyxin B sulfate, with/without Linco-Spectin (500 units/ml, 2000 ug/ml, 1000 units/ml and 300/600 ug/ml, respectively) was used as the control treatment. Results indicated no significant effect on seminal quality as measured by field fertility under the conditions of these experiments using heated whole milk or egg yolk-sodium citrate seminal extenders. Use of the new antibiotic combination has been adopted by Certified Semen Services.     

Effect of 3-hydroxy-4-1(H)pyridone on the basal and thyrotropin-releasing hormone-stimulated release of thyroid-stimulating hormone from perifused anterior pituitary fragments

This study investigated the direct effect of 3-hydroxy-4-1(H)-pyridone (DHP), the breakdown product of mimosine in the rumen, on thyroid-stimulating hormone (TSH) secretion by perifusion of rat anterior pituitary fragments. During a 2-h perifusion with thyrotropin-releasing hormone (TRH), the total release of TSH increased linearly (P less than 0.05, r = 0.966) with increasing concentration of TRH from 1 to 100 ng/ml. The release was maximal at 100 ng/ml. There were no differences in total basal TSH release among control and DHP-treated pituitary fragments. DHP at concentrations of 1, 10, and 100 micrograms/ml had no significant effect on the TSH response to TRH. However, DHP at the concentration of 1 mg/ml significantly suppressed the TSH response to TRH administered continuously or as a 10-min pulse. These results suggest that DHP modulates the pituitary thyrotroph's response to TRH.     

Indirect stimulation by thyrotropin-releasing hormone of canine gallbladder motility

Thyrotropin-releasing hormone (TRH) was unable to induce any noticeable contraction of canine isolated gallbladder strips up to the dose of 10(-4) g/ml, while caerulein (CAER) was spasmogenic in a dose-related manner beyond 10(-11) g/ml. This effect of CAER was unaffected by either atropine or tetrodotoxin. In conscious dogs, the intravenous bolus of TRH (20 micrograms/kg) or CAER (0.2-2.0 micrograms/kg) caused gallbladder emptying. The TRH response, unlike that of an equipotent dose of CAER, was prevented by atropine. In experiments on electrical activity of the digestive tract in conscious dogs, both TRH or CAER induced a concomitant increase on the myoelectrical activity in the proximal part of the small intestine. The excitatory effects were prevented by atropine only in the case of TRH. These results demonstrate that TRH stimulates indirectly the gallbladder and proximal duodenum of the dog. They suggest the involvement of a cholinergic pathway in this excitatory action.     

Stimulation of growth hormone release in dwarf and normal chickens by thyrotrophin releasing hormone (TRH) or human pancreatic growth hormone releasing factor (hpGRF)

The effect of thyrotrophin releasing hormone (TRH) or human pancreatic growth hormone releasing factor (hpGRF) on growth hormone (GH) release was studied in both dwarf and normal Rhode Island Red chickens with a similar genotype except for a sex-linked dw gene. Both TRH (10 micrograms/kg) and hpGRF (20 micrograms/kg) injections stimulated plasma GH release within 15 min in young and adult chickens. The increase in GH release was higher in young cockerels than that in adult chickens. The age-related decline in the response to TRH stimulation was observed in both strains, while hpGRF was a still potent GH-releaser in adult chickens. The maximal and long acting response was observed in young dwarf chickens, suggesting differences in GH pools releasable by TRH and GRF in the anterior pituitary gland. The pituitary gland was stimulated directly by perifusion with hpGRF (1 microgram/ml and 10 micrograms/ml) or TRH (1 microgram/ml). Repeated perifusion of GRF at 40 min intervals blunted further increase in GH release, but successive perifusion with TRH stimulated GH release. The results suggest the possibility that desensitization to the effects of hpGRF occurs in vitro and that the extent of response depends on the number of receptors for hpGRF or TRH and/or the amount of GH stored in the pituitary gland.     

Synergism of cortisol and thyrotropin-releasing hormone in lung maturation in fetal sheep

The effects of fetal infusions of cortisol and thyrotropin-releasing hormone (TRH) singly and together on pressure-volume relationships and saturated phosphatidylcholine (SPC) concentrations in the lungs were studied in 28 fetal sheep delivered at 128 days of gestation. Four groups each of 7 fetuses were infused with either saline (for 156 h), TRH (25 micrograms/h in 60-s pulses for 156 h), TRH (for 156 h) combined with cortisol (1 mg/h for 84 h), or cortisol (for 84 h). Cortisol had no effect on SPC concentrations, whereas both TRH and cortisol plus TRH increased the concentration of SPC in lavage fluid but not lung tissue. Neither cortisol nor TRH significantly affected lung distensibility [V40; 0.64 +/- 0.04 and 0.57 +/- 0.10 (SE) ml/g, respectively, vs. 0.41 +/- 0.03 ml/g in controls] or stability (V5; 0.24 +/- 0.01 and 0.35 +/- 0.07 ml/g vs. 0.24 +/- 0.03 ml/g), whereas treatment with a combination of the two hormones was associated with a fourfold increase in V40 (1.70 +/- 0.16 ml/g) and V5 (1.03 +/- 0.15 ml/g). Since raised concentrations of cortisol, triiodothyronine, and estradiol-17 beta (treatment with cortisol) had no effect on V40 and V5, whereas similar hormonal changes associated with elevated prolactin levels (treatment with cortisol plus TRH) had marked effects, we conclude that prolactin plays an essential part in the synergism of cortisol and TRH.     

Effect of thyrotropin releasing hormone thyroxine on the activity of cytochrome oxidase in rat adenohypophysis]

Thyrotropin releasing-hormone (TRH) increased the activity of cytrochrome C oxidase in a concentration of 0.01 and 1.0 microgram/ml in the adenohypophysis of rats fed methylthiouracil for 6 weeks. This effect of TRH on the activity of the enzyme was blocked with T4 added to the incubation medium in a concentration of 20 microgram/ml. Actinomycin D (20 MICrogram/ml) prevented the block of the enzyme with thyroxin. In a concentration of 0.01 microgram/ml TRH, and in a concentration of 2.0 microgram/ml T4 failed to change the activity of cytochrome oxidase in the adenohypophysis of normal and partially thyroidectomized rats.     

Effects of dopamine on the release of thyrotropin-releasing hormone from the rat retina in vitro.

The effects of dopamine on the release of thyrotropin-releasing hormone (TRH) from the rat retina in vitro were studied. The rat retina was incubated in the medium 199 (pH 7.4) with 1.0 mg/ml of bacitracin and 100 micrograms/ml of ascorbic acid. The amount of TRH release into the medium was measured by radioimmunoassay. The TRH release from the rat retina was inhibited significantly in a dose-related manner with the addition of dopamine, but not with pimozide. The inhibitory effects of dopamine on TRH release from the rat retina were blocked with an addition of pimozide to the medium. The elution profile of methanol-extracted rat retina on sephadex G-10 was identical to that of synthetic TRH. From these findings it is concluded that the dopaminergic system inhibits TRH release from the rat retina in vitro.     

Effects of the dopamine agonist cabergoline on the pulsatile and TRH-induced secretion of prolactin, LH, and testosterone in male beagle dogs

In the present study, the pulsatile serum profiles of prolactin, LH and testosterone were investigated in eight clinically healthy fertile male beagles of one to six years of age. Serum hormone concentrations were determined in blood samples collected at 15 min intervals over a period of 6 h before (control) and six days before the end of a four weeks treatment with the dopamine agonist cabergoline (5 microg kg(-1) bodyweight/day). In addition, the effect of cabergoline administration was investigated on thyrotropin-releasing hormone (TRH)-induced changes in the serum concentrations of these hormones. In all eight dogs, the serum prolactin concentrations (mean 3.0 +/- 0.3 ng ml(-1)) were on a relatively constant level not showing any pulsatility, while the secretion patterns of LH and testosterone were characterised by several hormone pulses. Cabergoline administration caused a minor but significant reduction of the mean prolactin concentration (2.9 +/- 0.2 ng ml(-1), p < 0.05) and did not affect the secretion of LH (mean 4.6 +/- 1.3 ng ml(-1) versus 4.4 +/- 1.7 ng ml(-1)) or testosterone (2.5 +/- 0.9 ng ml(-1) versus 2.4 +/- 1.2 ng ml(-1)). Under control conditions, a significant prolactin release was induced by intravenous TRH administration (before TRH: 3.8 +/- 0.9 ng ml(-1), 20 min after TRH: 9.1 +/- 5.9 ng ml(-1)) demonstrating the role of TRH as potent prolactin releasing factor. This prolactin increase was almost completely suppressed under cabergoline medication (before TRH: 3.0 +/- 0.2 ng ml(-1), 20 min after TRH: 3.3 +/- 0.5 ng ml(-1)). The concentrations of LH and testosterone were not affected by TRH administration. The results of these studies suggest that dopamine agonists mainly affect suprabasal secretion of prolactin in the dog.     

In vitro release of prolactin by thyrotropin-releasing hormone: influence of dopamine, thyroxine, and cycloheximide.

An acute incubation procedure, using explanted normal rat hemipituitaries pretreated with fresh plasma obtained from pituitary donor animals, was employed to further investigate the in vitro stimulation of prolactin (PRL release by thyrotropin-releasing hormone (TRH). Pretreatment with dopamine (0.1 microgram/ml) caused a 30-50% decrease in the amount of PRL released into incubation media; the inhibitory effect of dopamine was not reversed by treatment with 0.5-6.0 ng. TRH, although these TRH concentrations consistently stimulated PRL release from pituitaries not exposed to dopamine. Treatment with thyroxine (10(-6) to 10(-5) M) showed a competitive inhibition of thyrotropin release by TRH (0.5 ng), but was without effect on TRH-stimulated PRL release. Cycloheximide (100 microgram/ml) blocked a net increase in PRL levels. TRH, nevertheless, significantly increased PRL release in the presence of cycloheximide. The results indicate that neither dopamine nor thyroxine compete with TRH in causing PRL release, and that the TRH stimulation of PRL release is unrelated to ongoing levels of hormone synthesis.     

Effects of acetylcholine on thyrotropin-releasing hormone release from the rat caecum in vitro

Effects of acetylcholine on the release of thyrotropin-releasing hormone (TRH) from the rat caecum in vitro were studied. The rat caecum was incubated in medium 199 with 1.0 mg/ml of bacitracin and 100 micrograms/ml of ascorbic acid (pH 7.4) (medium). The amount of TRH release into the medium was measured by radioimmunoassay. The immunoreactive TRH (ir-TRH) release from the rat caecum was enhanced significantly in a dose-related manner with the addition of acetylcholine, but not changed with atropine. The stimulatory effect of acetylcholine on ir-TRH release from the rat caecum was blocked with an addition of atropine. Elution profile of acid-methanol-extracted rat caecum on Sephadex G-10 was identical to that of synthetic TRH. The findings suggest that the cholinergic system stimulates TRH release from the rat caecum in vitro.     

High concentrations of p-Glu-His-Pro-NH2 (Thyrotropin-releasing hormone) occur in rat prostate

Thyrotropin-releasing hormone (TRH) immunoreactivity occurs in high concentration within the rat prostate. Previous studies have shown that the immunoreactive species consists of more than one TRH-like tripeptide which cross-reacts in the TRH radioimmunoassay. The component which was highly retained during cation exchange chromatography was subjected to a preparative scale isolation, purification and structural analysis. The methods used included methanol extraction, waterethyl ether partitioning, cation exchange chromatography, affinity chromatography, high pressure liquid chromatography, TRH radioimmunoassay, in vitro pituitary bioassay, TRH receptor assay, and amino acid analysis. The mean concentration of the predominant amino acids (Glu, His, Pro), 344 pmoles/ml, and the TRH concentration measured by TRH radioimmunoassay prior to acid hydrolysis, 372 pmoles/ml, were nearly identical. Because the material analyzed cochromatographed with synthetic TRH in several chromatographic systems, had a radioreceptor potency which was indistinguishable from that for synthetic TRH, and released TSH and prolactin but not growth hormone from rat pituitaries in vitro, it is concluded that pGlu-His-Pro-NH₂ is one of the TRH-like peptides in the rat vental prostate.     

Effect of thyrotropin-releasing hormone on immune functions of peripheral blood mononuclear cells

The tripeptide thyrotropin-releasing hormone (TRH) works as a hypothalamic hormone, but is found also outside the brain in intrinsic nerve fibers of the gastrointestinal tract. There is evidence that TRH modulates the activity of immunocompetent cells, although there are only very few data on TRH-mediated immune effector functions. Since we could recently show that TRH inhibits monocyte activities we were also interested in other possible TRH modulated immune functions. Peripheral blood mononuclear cells (PBMC) from ten healthy subjects were cultured for 7 days and pulsed with 0.125 and 0.250 microgram/ml Pokeweed mitogen (PWM). 10(-12) to 10(-6) M TRH was added simultaneously with PWM. Lymphocyte proliferation [(3H]thymidine incorporation), interferon-gamma (IFN-gamma) activity (RIA) and immunoglobulin activities (IgG, IgM, IgA; ELISA) were determined in the supernatants. We could demonstrate a TRH-dependent decrease in PWM-pulsed IgG activity with significant (alpha = 0.05) values at 10(-8) and 10(-10) M (-29 +/- 6%/-16 +/- 3% for PWM 0.125 microgram/ml and -17 +/- 9%/-11 +/- 9% for PWM 0.250 microgram/ml). This inhibitory effect could be abolished by an anti-TRH antiserum. There was no TRH effect on IgM and IgA activities, IFN-gamma activity and lymphocyte proliferation compared with the PWM stimulated values alone. The described TRH effect on the polyclonal IgG response by PBMC gives further evidence for a functional link between the immune system and the endocrine system, although its underlying mechanism is not yet clear.     

人参皂苷Rb1、Rc对大鼠胚胎脑发育及GPx基因表达的影响

目的观察人参皂苷Rb1、Rc对sD大鼠胚胎脑发育及谷胱甘肽过氧化物酶（61utathione peroxidase,GPx）基因表达的影响。方法实验选用全胚胎体外培养方法,根据Van Maele-Fabry等制定的形态学分析系统进行形态学检测,实验随机分为对照组、人参皂苷Rbl（10,45和90ug／ml）组和人参皂苷Rc（10,45和90ug／ml）组,采用半定量RT—PCR分析人参皂苷Rb1和人参皂苷Rc对胚胎谷胱甘肽过氧化物酶基因表达的影响。结果与对照组比较,人参皂苷Rb1（10,45和90ug／ml）组胚胎脑、前脑、中脑与后脑的长度值没有统计学意义（P〉0．05）;与对照组比较,人参皂苷Rc（10,45和90ug／m1）组前脑、中脑与后脑的长度值显著增加（P〈0．05）。RT—PCR结果显示,人参皂苷Rb1（10,45和90ug／ml）组cGPx与phGPx mRNA的表达水平与对照组比较均没有统计学意义（P〉O．05）,而pGPxmRNA的表达水平在浓度50和100ug／ml时显著增加（P〈0．05）。人参皂苷Rc（10,45和90ug／ml）组cGPx与phGPxmRNA的表达水平与对照组比较均没有统计学意义（P〉0．05）,而pGPxmRNA的表达水平在各种浓度的Rc作用后都增加,且在溶度10和45ug／ml浓度时增加显著（P〈0．05）。结论一定剂量的人参皂苷Rc能促进胚胎脑的发育,一定剂量的人参皂苷Rb1、Rc能明显增强胚胎脑pGPx mRNA的表达水平。     

Effects of femto- and picomolar concentrations of thyroliberin and tuftsin on the contractile activity of lymphatic vessels of the rat mesentery

Thyrotropin-releasing-hormone (TRH) in concentration from I g/ml to 1.10(-10) micrograms/ml and tuftsin (T) (10 micrograms/ml to 1.10(-6) micrograms/ml) biomicroscopy were used in rats to show the ability to activate the contraction of lymphatic microvascular walls. TRH in used concentrations stimulated spontaneously contractility of lymphatic vessel's walls or evoked its contractility when it was absent. T stopped the contraction of lymphatic microvascular walls and caused the reaction of vasodilatation.     

A new antibiotic combination for frozen bovine semen. 2. Evaluation of seminal quality

Amikacin, clindamycin, gentamicin, Linco-Spectin, minocin and tylosin were added individually and in combinations at various concentrations to bovine neat semen and to egg-yolk citrate, egg yolk-tris or heated whole milk extenders (nonglycerol fractions) prior to final processing for freezing to -196 degrees C. After thawing samples, seminal quality was measured by progressive motility and acrosomal integrity evaluations. Studies were performed in parallel with microbiological efficacy studies of Shin et al. (7). The antibiotic combination including gentamicin, tylosin and Linco-Spectin at 500 ug/ml, 100 ug/ml and 300/600 ug/ml, respectively, was not detrimental to seminal quality and, in the parallel studies, was most efficacious in controlling microorganisms potentially present in bovine semen.     

Dopamine inhibits thyrotropin-releasing hormone release from rat adrenal gland in vitro

The effects of dopamine on the release of thyrotropin-releasing hormone (TRH) from the rat adrenal gland were studied in vitro. The rat adrenal glands were incubated in medium 199 with 1.0 mg/ml of bacitracin and 100 micrograms/ml of ascorbic acid (pH 7.4) (medium) for 20 min. The amount of TRH release into the medium was measured by radioimmunoassay. The immunoreactive TRH (ir-TRH) release from the rat adrenal gland was inhibited significantly in a dose-related manner with the addition of dopamine and enhanced with the addition of pimozide or domperidone to the medium. Dopamine's effects on ir-TRH release from the adrenal gland were blocked with the addition of pimozide or domperidone. The elution profile of methanol-extracted rat adrenal gland was identical to that of synthetic TRH. The findings suggest that the dopaminergic system inhibits TRH release from the rat adrenal gland.