生长激素和生长激素受体的多样性

生长激素及其受体对动物生长发育起着重要的作用。转录过程选择性剪接和存在多种降解途径可能是GH或GHR产生多样性的原因。随着GH结构形态的改变,其功能也在发生变化。GH基因的多样性对鸡的抗病选择性反应与产蛋性能有相关,GH和GHR基因的多样性会影响奶牛的产奶生产性能。GHR的分子多样性可能导致动物生长发育模式的变异,例如动物的矮小病。     

Interactions of gonadotropin-releasing hormone (GnRH) and gonadotropin-inhibitory hormone (GnIH) in birds and mammals

Gonadotropin-releasing hormone (GnRH) regulates secretion of both of the gonadotropins, luteinizing hormone (LH) and follicle-stimulating hormone. Thus, it is a key hormone for vertebrate reproduction. GnRH was considered to be unusual among hypothalamic neuropeptides in that it appeared to have no direct antagonist, although some neurochemicals and peripheral hormones (opiates, GABA, gonadal steroids, inhibin) can modulate gonadotropin release to a degree. Five years ago, a vertebrate hypothalamic neuropeptide that inhibited pituitary gonadotropin release in a dose-dependent manner was discovered in quail by Tsutsui et al. (2000. Biochem Biophys Res Commun 275:661-667). We now know that this inhibitory peptide, named gonadotropin-inhibitory hormone, or GnIH, is a regulator of gonadotropin release in vitro and in vivo. Its discovery has opened the door to an entirely new line of research within the realm of reproductive biology. In our collaborative studies, we have begun to elucidate the manner in which GnIH interacts with GnRH to time release of gonadotropins and thus time reproductive activity in birds and mammals. This paper reviews the distribution of GnIH in songbirds relative to GnRHs, and our findings on its modes of action in vitro and in vivo, based on laboratory and field studies. These data are simultaneously compared with our findings in mammals, highlighting how the use of different model species within different vertebrate classes can be a useful approach to identify the conserved actions of this novel neuropeptide, along with its potential importance to vertebrate reproduction.     

Pattern of luteinizing hormone (LH) and follicle stimulating hormone (FSH) in bovine blood plasma after injection of a synthetic gonadotropin-releasing hormone (Gn-RH)

A synthetic gonadotropin-releasing hormone (Gn-RH) was administered to female and male adult bovines in order to study the release of luteinizing and follicle-stimulating hormones into blood by the pituitary gland. Plasma LH and FSH were determined by means of a radioimmunological method. In females as well as in males, increasing doses of Gn-RH (range 50 to 1500 μg) administered i.v. or i.m. caused a linear increase in plasma LH. The release of FSH evidently was curvilinear over the same dosage range.After 2 or 3 injections of Gn-RH every 3 hours, or every 24 hours or more, smaller amounts of LH were released; repeated treatment did not result in reduction of FSH. Thus pituitary depletion of LH occurred more readily than FSH. The effect of Gn-RH on plasma levels of LH and FSH at various stages of the estrous cycle shows a tendency for an increasing release of both gonadotropins on Days 17 – 18 in comparison to Days 4 – 5 or Days 11 – 12.The results suggest that, within the limits allowed by the heterogenous FSH assay and the method of administration used in these experiments, synthetic Gn-RH does not evoke completely normal physiological responses. Therefore, further work is needed to determine its role in improving reproductive function.     

Regulation of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) gene expression by gonadotropin-releasing hormone (GnRH) and sexual steroids in the Mediterranean Sea bass

The secretion of gonadotropins, the key reproductive hormones in vertebrates, is controlled from the brain by the gonadotropin-releasing hormone (GnRH), but also by complex steroid feedback mechanisms. In this study, after the recent cloning of the three gonadotropin subunits of sea bass (Dicentrarchus labrax), we aimed at investigating the effects of GnRH and sexual steroids on pituitary gonadotropin mRNA levels, in this valuable aquaculture fish species. Implantation of sea bass, in the period of sexual resting, for 12 days with estradiol (E2), testosterone (T) or the non-aromatizable androgen dihydrotestosterone (DHT), almost suppressed basal expression of FSHbeta (four to 15-fold inhibition from control levels), while slightly increasing that of alpha (1.5-fold) and LHbeta (approx. twofold) subunits. Further injection with a GnRH analogue (15 microg/kg BW; [D-Ala6, Pro9-Net]-mGnRH), had no effect on FSHbeta mRNA levels, but stimulated (twofold) pituitary alpha and LHbeta mRNA levels in sham- and T-implanted fish, and slightly in E2- and DHT-implanted fish (approx. 1.5-fold). The GnRHa injection, as expected, elevated plasma LH levels with a parallel decrease on LH pituitary content, with no differences between implanted fish. In conclusion, high circulating steroid levels seems to exert different action on gonadotropin secretion, inhibiting FSH while stimulating LH synthesis. In these experimental conditions, the GnRHa stimulate LH synthesis and release, but have no effect on FSH synthesis.     

Effects of the gonadotropin-releasing hormone associated peptides (GAP) on the release of luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin (PRL) in vivo

Six peptide sequences residing between basic amino acid residues in GAP were tested for effects on the release of FSH, LH and PRL in vivo in ovariectomized, estrogen-progesterone-primed (OEP) rats. Synthetic GAP peptides (1–13, 1–23, 15–23, 25–36, 38–53 and 41–53) were injected intravenously (IV) into conscious OEP rats and plasma levels of FSH, LH and PRL were measured by RIA. The activity of GAP peptides in the control of PRL was further examined in ether-stressed male rats which were injected IV with GAP peptides just prior to a 1-min etherization. GAP(1–13) significantly stimulated FSH release at doses of 1, 10 and 100 μg, whereas it stimulated LH release only at the highest dose of 100 μg. GAP(1–23) elevated plasma levels of FSH and LH only at a dose of 100 μg. The other 4 peptides had no effect on the release of gonadotropins. Of these 6 peptides, only GAP(1–13) partially lowered the plasma levels of PRL at the high dose of 100 μg in OEP rats, but it had no effect on the ether-induced PRL surge at doses of 10 and 100 μg. In conclusion, both GAP(1–13) and GAP(1–23) stimulate FSH and LH release in vivo; these 2 peptides are much less potent in stimulating gonadotropin release than is LHRH. GAP(1–13) exerts a preferential FSH-releasing activity, but its PRL-inhibiting activity is minimal.     

Cellular characteristics of immunolabeled luteinizing hormone releasing hormone (LHRH) neurons

This study utilized the preembedding immunocytochemical technique in order to identify LHRH-containing neurons in rat brain and define their ultrastructural characteristics. LHRH-containing neurons in the vertical limb of the diagonal band of Broca, medial septum, triangular nucleus of the septum and other regions were studied by taking ultrathin serial sections. These neurons had scant cytoplasm surrounding a centrally-located, spheroid, euchromatic nucleus. Neurosecretory granules were evenly distributed throughout the cell, but many tended to lie directly under the plasmalemma. The cytoplasm was organized in such a way that the most extensive portion of the rough endoplasmic reticulum was polar opposite to areas having high concentrations of Golgi complex, lysosome-like bodies, smooth endoplasmic reticulum and ribosomes. The perikarya had no axosomatic synapses but functional interaction via unspecialized appositions to the plasmalemma cannot be discounted. Many of the perikarya bore at least one cilium. Processes from immunonegative cells were occasionally observed to penetrate the cytoplasm of the LHRH perikaryon or its processes. At their points of origin, dendrites were found to be broadened processes containing many elements common to the cytoplasm: ribosomes, smooth endoplasmic reticulum, cristal and lamellar mitochondria, neurotubules, and an occasional alveolate caveola. Infrequently, some of the LHRH axons were partially myelinated. This method of studying serial-sectioned immunocytochemically-identified cells is suggested as a means of describing the cellular and subcellular characteristics of other specific peptide-containing cells.     

Ultrasonography and hormone profiles of adrenocorticotrophic hormone (ACTH)-induced persistent ovarian follicles (cysts) in cattle

The objective of this study was to develop a model for the study of abnormal ovarian follicles in cattle by treating heifers with adrenocorticotrophic hormone (ACTH) (100 iu at 12 h intervals for 7 days, beginning on day 15 of the oestrous cycle). Cortisol concentrations increased (P < 0.05) within 24 h after beginning ACTH treatment and cortisol and progesterone concentrations remained elevated after cessation of ACTH treatment for 8 and 4 days, respectively. The pulses and surges of LH decreased during ACTH treatment, but FSH profiles were similar to those in controls and persistent or prolonged follicles were eventually observed in all heifers. In five heifers, prolonged dominant follicles ovulated after 10 days, whereas in six heifers, persistent follicular structures were present for 20 days, but ceased to secrete oestradiol after approximately 12 days. In the heifers with persistent follicular structures, new follicles emerged when the persistent follicle became non-oestrogenic. During the last 2 days of normal follicular growth, the concentration of oestradiol was greater than it was during prolonged or persistent follicle development (P < 0.05). There were no differences in the growth rates or maximum diameters of abnormal follicles that had different outcomes, but oestradiol concentrations were greater in prolonged follicles that ovulated compared with those follicles that persisted (P = 0.06). In conclusion, stimulation with ACTH resulted in a marked deviance from normal follicular activity. The aberrations were probably caused by the interruption of pulsatile secretion of LH (but not FSH) leading to decreased but prolonged oestradiol secretion.     

Modulation of long-term memory and extinction responses induced by growth hormone (GH) and growth hormone releasing hormone (GHRH) in rats

The purpose of this work is to study the participation of growth hormone (GH) and growth hormone releasing hormone (GHRH) in the modulation of long-term memory and the extinction response of a passive avoidance task in rats. However, the effect on memory vary according to the age of the animals due to plasma levels of either hormone being modified during the aging process. Male Wistar rats were divided according to age into two experimental blocks (young rats 3 months old and aged rats 24 months old at the start of the experiment) where each block received the same treatment. Each experimental block was then divided randomly into three groups where two were experimental and the other served as control. The animals were then submitted to a one-trial passive avoidance conditioning and tested for memory retention 24 hrs after as well as twice a week until the extinction response occurred. The control group received an isotonic saline solution and the other two groups received 0.8 U.I. Of GH or 4 mcg of GHRH respectively. All substances were in a 0.08 ml volume and applied 24 hrs before training as well as 24 hrs before each retention session. The results indicate that GH and GHRH modulate longterm memory as well as the extinction response and in either case the response seems to vary with age. GH and GHRH facilitates long-term memory in young rats but not in aged rats. Finally, whereas GH delays the extinction response in both groups, GHRH retards the extinction only in aged rats.     

Ontogeny of luteinizing hormone releasing hormone (LHRH) and somatostatin (SRIF) in the hypothalamus of the sheep

Using the immunoperoxidase method, luteinizing hormone releasing hormone (LHRH) and somatostatin (SRIF) were demonstrated in the hypothalamus of fetal sheep. Both hormones were found in the perikarya at about day 60 of fetal life, i.e., at the end of the first half of pregnancy. Immunoreactive LHRH (irLHRH) perikarya were situated in the vicinity of the organum vasculosum of the lamina terminalis (OVLT), i.e., in the medial preoptic nucleus and in the nucleus of the diagonal band of Broca. They were scattered and generally sparse in these areas. In the earliest stages of fetal life (60, 75, 90 days of gestation) irSRIF perikarya grouped in the ventromedial nucleus and in the lateral preoptic nucleus, were very numerous. In the oldest fetuses (120 and 135 days of gestation) they had disappeared from these nuclei but could be found in some extrahypothalamic regions--the amygdala, septo-olfactory area and sometimes in the anterior periventricular zone of the hypothalamus. Neither irLHRH nor irSRIF material were stored in the nerve terminals of the external layer of the median eminence (ME) before day 75 of gestation. In all developmental stages examined, irLHRH material in the ME was very scarce whereas irSRIF material very aboundant.     

Gonadotropin-releasing hormone (GnRH) inhibits ovulation induced with luteinizing hormone (LH) in proestrous hypophysectomized rats

In this paper we present evidence that a single low dose of the natural synthetic gonadotropin-releasing hormone (GnRH), inhibits ovulation induced by LH in proestrous-hypophysectomized rats. Rats hypophysectomized by the parapharyngeal route in the morning of proestrus received an intravenous injection of 100 or 300 ng GnRH at 1400 h immediately followed by 1.0 microgram LH per 100 g bw. In control groups, either one or both hormones were replaced with 0.9% NaCl. Ovulation was assessed the following morning by counting the ova present in oviductal flushings. All the rats treated with LH alone ovulated, and the addition of GnRH reduced significantly the number of ovulating rats and the number of ova per ovulating rat. In other groups of rats hypophysectomized in the morning of proestrus and treated in the same way, ovarian or adrenal secretory rates of estradiol and/or progesterone were measured after cannulation of the corresponding vein, in the afternoon of proestrus. In these animals, GnRH failed to inhibit either the ovarian progesterone surge observed 2 h after LH administration, or the adrenal progesterone secretion. All hypophysectomized rats showed lower ovarian secretory rate of estradiol than intact rats; this rate was not affected by treatment with LH or LH plus GnRH. The systemic estradiol levels in plasma of hypophysectomized rats were distributed within a range of 20 pg/ml to 50 pg/ml. The number of rats whose levels were above 21 pg/ml on estrus day was significantly higher in rats receiving 300 ng GnRH as compared to those receiving 100 ng GnRH, reaching values that surpassed the concentration found in intact, untreated animals at the same time of estrus. This effect did not depend on LH administration.     

Gonadotropin-releasing hormone (GnRH) pharmacokinetics: peptide hormone pharmacokinetics needs clarification

The plasma level curves of the peptide hormone gonadotropin-releasing hormone (GnRH) after its intravenous, intramuscular, and intraperitoneal administration into rats were fitted according to a two- (i.v.) and one-compartment model (i.m., i.p.), respectively. From the pharmacokinetic parameters it is concluded that urinary excretion and proteolytic degradation by kidney and liver are not sufficient to fully account for the clearance of the hormone and that, therefore, proteolytic degradation by tissues may play a role for the elimination of GnRH. This may be generally true with other short peptide hormones. The GnRH pharmacokinetics is shown as an example to underline that there presently exist problems of interpreting pharmacokinetic data of peptide hormones and that there is a need for a close interplay between biochemical and pharmacokinetic studies on peptide hormones for their pharmacokinetic behaviour to be understood.     

Dual intracellular pathways in gonadotropin releasing hormone (GNRH) induced desensitization of luteinizing hormone (LH) secretion.

The mechanisms of GnRH-induced desensitization of LH secretion are poorly understood. Protein kinase C (PKC) and protein kinase A (PKA) desensitize some receptors of the 7-membrane type, and the GnRH receptor has consensus phosphorylation sites for PKC in the first and third intracellular loops, and a site for PKA in the first intracellular loop. In the first set of experiments we determined whether synthetic peptides representing the three intracellular loops of the receptor could be phosphorylated in vitro by purified PKC and PKA. As compared with a model substrate peptide for PKC, the third intracellular loop was phosphorylated 74% and the first intracellular loop 21%; PKA-phosphorylated the first intracellular loop peptide 17% as well as a model peptide substrate. In the second set of experiments, we used phorbol 12-myristate 13 acetate (PMA), an established PKC stimulator, and cholera toxin (CTX), established to activate the Gs protein and presumed to activate PKA, to treat cultured rat pituitary cells followed by LH measurements. Treatment with both drugs severely impaired GnRH-stimulated LH secretion whereas neither drug alone reduced LH secretion. Dibutyryl cAMP did not duplicate the effects of cholera toxin suggesting that the CTX action could not be explained by an increase in cAMP. These results suggest that more than one intracellular signaling pathway requires activation in order to induce desensitization; one pathway involves PKC and the other involves a pathway stimulated by cholera toxin, presumably Gs protein, which does not involve PKA.     

alpha-MSH (melanocyte stimulating hormone) and MCH (melanin concentrating hormone) actions in Bufo ictericus ictericus melanophores

1. The darkening actions of MCH (melanin concentrating hormone), alpha-MSH and the synthetic analog [Nle4, D-Phe7]-alpha-MSH on the toad, Bufo ictericus ictericus, melanophores were studied regarding the role of calcium in the hormone receptor coupling, signal transduction and intracellular pigment translocation. 2. In the absence of external calcium, MCH and both melanotropins still elicit maximal skin darkening. 3. Verapamil, a calcium-channel blocker, completely abolishes the alpha-MSH-induced response and partially inhibits MCH-induced darkening, although the calcium carrier, ionophore A23187, was unable to promote any pigment translocation. 4. Since darkening responses promoted by cyclic nucleotides proceeded normally in the presence of verapamil and extracellular calcium was not necessary for melanotropin dispersing action, it is suggested that the blocking activity obtained with verapamil is probably due to an impairment of the Ca2+-dependent adenylate cyclase activity. 5. Reversal of melanotropin-induced darkening could be obtained with melatonin, in both normal and Ca2+-free Ringer, whereas MCH darkening is reversed by melatonin only in the absence of calcium. 6. The results seem to indicate that calcium is not required for hormone receptor binding and pigment migration, whereas it is specifically needed for signal transduction.     

Robust growth hormone (GH) secretion in aged female rats co-administered GH-releasing hexapeptide (GHRP-6) and GH-releasing hormone (GHRH)

Aging is associated with a blunted growth hormone (GH) secretory response to GH-releasing hormone (GHRH), in vivo. The objective of the present study was to assess the effects of aging on the GH secretory response to GH-releasing hexapeptide (GHRP-6), a synthetic GH secretagogue. GHRP-6 (30 micrograms/kg) was administered alone or in combination with GHRH (2 micrograms/kg) to anesthetized female Fischer 344 rats, 3 or 19 months of age. The peptides were co-administered to determine the effect of aging upon the potentiating effect of GHRP-6 on GHRH activity. The increase in plasma GH as a function of time following administration of GHRP-6 was lower (p less than 0.001) in old rats than in young rats; whereas the increase in plasma GH secretion as a function of time following co-administration of GHRP-6 and GHRH was higher (p less than 0.001) in old rats than in young rats (mean Cmax = 8539 +/- 790.6 micrograms/l vs. 2970 +/- 866 micrograms/l, respectively; p less than 0.01). Since pituitary GH concentrations in old rats were lower than in young rats (257.0 +/- 59.8 micrograms/mg wet wt. vs. 639.7 +/- 149.2 micrograms/mg wet wt., respectively; p less than 0.03), the results suggested that GH functional reserve in old female rats was not linked to pituitary GH concentration. The differential responses of old rats to individually administered and co-administered GHRP-6 are important because they demonstrate that robust and immediate GH secretion can occur in old rats that are appropriately stimulated. The data further suggest that the cellular processes subserving GH secretion are intact in old rats, and that age-related decrements in GH secretion result from inadequate stimulation, rather than to maladaptive changes in the mechanism of GH release.     

Chemiluminescent immunoassay (CLIA) for salmon growth hormone (GH)

A highly sensitive and specific chemiluminescent immunoassay (CLIA) was developed for quantification of growth hormone (GH) in salmonid species. The CLIA for salmon GH was performed using the sandwich method with anti-GH IgG as the first antibody and chemiluminescent acridinium ester-labelled specific anti-GH F(ab′)₂ as the second antibody. The measurable range of salmon GH in the CLIA was 39–1250 pg/mL using a short assay (1 day) protocol and 3.9–125 pg/mL in a longer (2-day) assay. The dilution curve in the CLIA of serum from masu salmon (Oncorhynchus masou) was parallel to the standard curve of recombinant chum salmon (Oncorhynchus keta) GH. Seasonal changes of serum GH levels were measured in 1 year-old masu salmon cultivated in a pond from March to November. Their serum GH levels increased during smoltification from March to April, achieved a maximum level of 21 ng/mL in August, and then declined gradually to 11 ng/mL in October. © 1997 John Wiley & Sons, Ltd.