Inhibition of growth hormone synthesis by somatostatin in cultured pituitary of rainbow trout

Summary When the pituitary of rainbow trout (Oncorhynchus mykiss) was incubated in a serum-free medium, a high level of growth hormone release as well as an activation of growth hormone synthesis were observed, suggesting the existence of hypothalamic inhibitory factor(s) on growth hormone synthesis. Although an inhibitory effect of somatostatin on growth hormone release is well established in both mammals and teleosts, an effect on growth hormone synthesis has not been demonstrated. In this study, we examined the effect of somatostatin on growth hormone synthesis in organ-cultured trout pituitary using immunoprecipitation and Northern blot analysis. Somatostatin inhibited growth hormone release from the cultured pituitary within 10 min after addition without affecting prolactin release. Incubation of the pituitary with somatostatin also caused a significant reduction in newly-synthesized growth hormone in a dose-related manner, as assessed by incorporation of [³H]leucine into immunoprecipitable growth hormone. There were no changes in the level or molecular length of growth hormone mRNA after somatostatin treatment, as assessed by Northern slot blot and Northern gel blot analyses. Human growth hormone-releasing factor stimulated growth hormone release, although the spontaneous synthesis of growth hormone was not augmented. However, somatostatin-inhibited growth hormone synthesis was restored by growth hormone-releasing factor to the control level. The spontaneous increase in growth hormone synthesis observed in the organ-cultured trout pituitary may be caused, at least in part, by the removal of the inhibitory effect of hypothalamic somatostatin.Abbreviations GH growth hormone - GHRF GH-releasing factor - PRL prolactin - SDS sodium dodecyl sulphate - SRIF somatostatin (somatropin release-inhibiting factor)     

Effects of TRH and somatostatin on releases of prolactin and growth hormone in vitro by the pituitary of Poecilia latipinna

Summary Pituitary glands from a teleost fish were incubated in the presence of the synthetic hypophysiotropic peptides, thyrotrophin-releasing hormone and somatostatin, in two media of different osmotic pressure.The effects on prolactin and growth hormone cells were detected by electron-microscopic morphometry with the aid of an image analyser. Thyrotrophin-releasing hormone caused changes in prolactin cell ultrastructure consistent with stimulated hormone release and, in the low osmotic pressure medium, appeared to increase synthetic activity. There was no effect on growth hormone cells. After somatostatin treatment, both synthesis and release in prolactin cells appeared to be inhibited, and there was an obvious inhibition of synthesis and release in growth hormone cells. The response of both cell types to somatostatin did not appear to be dependent on the osmotic pressure of the medium.     

Dopamine-stimulated parathyroid hormone release in vitro: further evidence for a two-pool model of parathyroid hormone secretion

Bovine parathyroid tissue was placed in an in vitro perifusion system for the study of parathyroid hormone secretion stimulated by low calcium and dopamine. Dopamine caused a transient increase in parathyroid hormone release, while low calcium caused a sustained increase in parathyroid hormone secretion. The dopamine response was similar to that caused by isoproterenol. After parathyroid hormone release had been stimulated by dopamine there was no response to isoproterenol, suggesting they cause the release of the same cellular pool of hormone. Inhibition of protein synthesis with cycloheximide eliminated the response to low calcium, with no effect on dopamine-stimulated parathyroid hormone release. These studies suggest dopamine stimulates the release of a limited quantity storage pool of parathyroid hormone, while low calcium causes a sustained release of hormone by stimulating secretion of newly synthesized hormone. Low calcium has little or no effect on release of the storage granule pool of parathyroid hormone.     

Effect of zinc on the binding and action of growth hormone in isolated rat adipocytes

Ionic zinc has been shown to exert a dose-dependent (250-500 microM) stimulation of 125I-human growth hormone specific binding to isolated rat adipocytes. The effect was rapid, being observed in less than 15 min exposure to zinc, and was due to an increased number of available binding sites as determined by Scatchard analysis. The well-known insulin-like effects of both zinc and growth hormone on lipogenesis in this tissue were additive at submaximal doses of each agonist, but zinc did not appear to potentiate growth hormone action. A similar, but smaller effect of zinc on growth hormone specific binding was observed in rabbit liver membranes at lower zinc concentrations (5-100 microM). At higher zinc doses a marked increase in non-specific binding caused a reduction of specific binding in liver membranes. These studies raise the possibility that zinc may be a modulator of growth hormone binding in vivo. Although our initial experiments suggest that this may be independent of an effect on growth hormone action, further studies are required to assess the possibility that zinc's effect at the receptor level may be to increase the sensitivity of tissues to growth hormone, thereby promoting a full manifestation of effects at lower growth hormone concentrations.     

H9c2 cardiomyoblasts produce thyroid hormone

Thyroid hormone acts on a wide range of tissues. In the cardiovascular system, thyroid hormone is an important regulator of cardiac function and cardiovascular hemodynamics. Although some early reports in the literature suggested an unknown extrathyroidal source of thyroid hormone, it is currently thought to be produced exclusively in the thyroid gland, a highly specialized organ with the sole function of generating, storing, and secreting thyroid hormone. Whereas most of the proteins necessary for thyroid hormone synthesis are thought to be expressed exclusively in the thyroid gland, we now have found evidence that all of these proteins, i.e., thyroglobulin, DUOX1, DUOX2, the sodium-iodide symporter, pendrin, thyroid peroxidase, and thyroid-stimulating hormone receptor, are also expressed in cardiomyocytes. Furthermore, we found thyroglobulin to be transiently upregulated in an in vitro model of ischemia. When performing these experiments in the presence of 125 I, we found that 125 I was integrated into thyroglobulin and that under ischemia-like conditions the radioactive signal in thyroglobulin was reduced. Concomitantly we observed an increase of intracellularly produced, 125 I-labeled thyroid hormone. In conclusion, our findings demonstrate for the first time that cardiomyocytes produce thyroid hormone in a manner adapted to the cell's environment.     

An electrostatic model for the interaction between growth hormone and its receptor involving chelation of Ca2+ to the human growth hormone molecule

We previously postulated the local involvement of cations in the complex between human growth hormone and its receptors in the liver. The original electrostatic model involved convergence of unit negative charges on the hormone and receptor, towards an interposed Ca2+ ion. That model was consistent with (a) the Ca2+ dependence of human growth hormone binding, (b) the magnitude of the Ca2+ mediated increase in the affinity of human growth hormone binding, and (c) could also explain the relative affinities of human and non-primate growth hormones for growth hormone receptors. In the present report, the original electrostatic model is revised with the postulate that Ca2+ is chelated to the human growth hormone molecule. The consequences of this postulate are explored mathematically with the result that it becomes necessary to propose an additional unique hindrance determinant (positive residues in helices one and four are good candidates) to account for the lower affinity of non-primate growth hormones relative to human growth hormone. Predictions are made regarding the effect of a particular point mutation (at position 34) on the affinity of hormone binding.     

NMDA Receptors in the Medial Zona Incerta Stimulate Luteinizing Hormone and Prolactin Release

1. The aim of the present work is to demonstrate the interaction between the glutamatergic/NMDA and dopaminergic systems in the medial zona incerta on the control of luteinizing hormone and prolactin secretion and the influence of reproductive hormones. 2. Proestrus and ovariectomized rats were primed with estrogen and progesterone to induce high or low levels of luteinizing hormone and prolactin. 2-Amino-7-phosphonoheptanoic acid, an NMDA receptor antagonist, and dopamine were injected in the medial zona incerta. Blood samples were withdrawn every hour between 1,600 and 2,000 hours or 2,200 hours via intracardiac catheter from conscious rats. Additional groups of animals injected with the NMDA receptor antagonist were killed 1 or 4 h after injection. Dopamine and its metabolite 3,4-dihydroxyphenylacetic acid were measured in different hypothalamic regions. 3. 2-Amino-7-phosphonoheptanoic acid blocked the ovulatory luteinizing hormone surge in proestrus rats. 2-Amino-7-phosphonoheptanoic acid also blocked the increase in luteinizing hormone induced by ovarian hormones in ovariectomized rats, an effect that was partially reversed by dopamine injection. Conversely, the increased release of luteinizing hormone and prolactin induced by dopamine was prevented by 2-amino-7-phosphonoheptanoic acid. We found that the NMDA antagonist injection decreased the dopaminergic activity--as evaluated by the 3,4-dihydroxyphenylacetic acid/dopamine ratio--in the medio basal hypothalamus and increased in the preoptic area. 4. Our results show an stimulatory role of NMDA receptors on the ovulatory luteinizing hormone release and on luteinizing hormone release induced by sexual hormones and demonstrate that the stimulatory effect of dopamine on luteinizing hormone and prolactin is mediated by the NMDA receptors. These results suggest a close interaction between the glutamatergic and dopaminergic incertohypothalamic systems on the control of luteinizing hormone and prolactin release.     

Growth hormone acts at a pretranslational level in hepatocyte cultures

We have examined the effects of ovine growth hormone and recombinant DNA synthesized human growth hormone on hepatocytes maintained in serum free cultures. Both growth hormone preparations augmented or attenuated 3 specific mRNA sequences as revealed by two-dimensional gel electrophoresis of [35S] methionine labeled products synthesized in vitro in an mRNA dependent rabbit reticulocyte lysate system. The results clearly indicate that growth hormone, free of potential pituitary contaminants, acts directly on hepatocytes at a pretranslational level.     

Studies on the Movement of Water through the Isolated Toad Bladder and Its Modification by Vasopressin

Measurements of diffusion permeability and of net transfer of water have been made across the isolated urinary bladder of the toad, Bufo marinus, and the effects thereon of mammalian neurohypophyseal hormone have been examined. In the absence of a transmembrane osmotic gradient, vasopressin increases the unidirectional flux of water from a mean of 340 to a mean of 570 µl per cm² per hour but the net water movement remains essentially zero. In the presence of an osmotic gradient but without hormone net transfer of water remains very small. On addition of hormone large net fluxes of water occur; the magnitude of which is linearly proportional to the osmotic gradient. The action of the hormone on movement of water is not dependent on the presence of sodium or on active transport of sodium. Comparison of the net transport of water and of unidirectional diffusion permeability of the membrane to water indicates that non-diffusional transport must predominate as the means by which net movement occurs in the presence of an osmotic gradient. An action of the hormone on the mucosal surface of the bladder wall is demonstrated. The effects of the hormone on water movement are most simply explained as an action to increase the permeability and porosity of the mucosal surface of the membrane.     

Juvenile hormone esterase is a biochemical anti-juvenile hormone agent

Juvenile hormone esterase, purified by affinity chromatography from the larval hemolymph of Manduca sexta in the fifth stadium, was injected into larvae of the same species in the earlier stadia resulting in a blackening of the cuticle following ecdysis to the next larval stadium. This anti-juvenile hormone response was dose-dependent for an injection in the second, third or fourth stadium. Cuticular blackening was prevented by treating larvae with the juvenoid epofenonane. Larval response to injected juvenile hormone esterase also varied with the time of injection within a single stadium, having a maximum effect for injections at the time of head capsule slippage. Juvenile hormone esterase activity measured from the hemolymph after injection of larvae in the second stadium decreased over an 11 h time-course. Because the anti-juvenile hormone effects resulting from a single injection of juvenile hormone esterase were dependent on the time of injection, it appears that when juvenile hormone biosynthesis is active in the insect, the duration of enzyme activity limits the anti-juvenile effects that can be induced.     

Hypophysiotropic activity of histone H3 in vitro

To assess the effect of histone H3 on pituitary hormone secretion, rat anterior pituitary (AP) cells were used and growth hormone, prolactin, thyrotropin, luteinizing hormone and follicle stimulating hormone measured by radioimmunoassay. Incubation of cells with H3 (1, 6, and 30 microM) stimulated the release of all five hormones in a dose-dependent manner. This effect was blocked by preincubation of H3 with an anti-H3 antibody. Incubation of AP cells with 6 microM H3 in the presence of specific AP hormone secretagogues (GRP-6, thyrotropin-releasing hormone (TRH), gonadotropin-releasing hormone (GnRH)) showed additive effects on hormone secretion. Pharmacological experiments suggested that calcium- and diacylglycerol- (DAG) associated pathways, but not cAMP, participate in the hypophysiotropic activity of H3. Our results confirm previous evidence that histones may act as hypophysiotropic signals.     

Acromegaly—The Effects of Various Steroid Hormones on the Insulin-Induced Growth Hormone Response

The availability of a sensitive assay for human growth hormone has made it possible to directly measure the effects of various agents purported to alter growth patterns. Acromegalic patients present a special problem both in early diagnosis and in therapy. Being able to measure growth hormone in these patients provides an accurate index of activity and a precise measure of therapeutic effectiveness.In an attempt to determine whether a pituitary block of growth hormone secretion is feasible in this condition, a study was made of the effects of estrogen, androgen and glucocorticoid administration on growth hormone response to a standard insulin tolerance test in a patient with active acromegaly. In the dosage schedules used in this study, it was not possible to suppress either basal growth hormone secretion or blunt its responsiveness to the normal physiologic stimulus of hypoglycemia.     

The effect of luteinizing hormone releasing hormone and anti-luteinizing hormone releasing hormone antibodies on chorionic gonadotropin and progesterone secretion by human placental villiin vitro

Gonadotropin releasing hormone has been located and found to be secreted by the human placenta in culture. Addition of the releasing hormone upto 1μg concentration in the placental cultures brings about stimulation of chorionic gonadotropin and progesterone secretion. Higher amounts of the decapeptide has an inhibitory influence on both the gonadotropin and the steroid production. The action of the releasing hormone on the placenta could be blocked by the anti-luteinizing hormone releasing hormone monoclonal antibodies indicating a possible site of action of the antibodies for control of fertility     

Development of a long acting human growth hormone analog suitable for once a week dosing

Human growth hormone was conjugated to a carrier aldolase antibody, using a novel linker by connecting a disulphide bond in growth hormone to a lysine-94 amine located on the Fab arm of the antibody. The resulting CovX body showed reduced affinity towards human growth hormone receptor, reduced cell-based activity, but improved pharmacodynamic properties. We have demonstrated that this CovX-body, given once a week, showed comparable activity as growth hormone given daily in an in vivo hypophysectomized rat model.     

Functional parathyroid hormone receptors are present in an umbilical vein endothelial cell line

Acute parathyroid hormone exposure induces vascular smooth muscle relaxation. In contrast, continuous infusion of parathyroid hormone leads to vasoconstriction and an elevation of blood pressure. Despite the known effects of parathyroid hormone on vascular smooth muscle, possible direct effects on the vascular endothelium have not previously been investigated. Using a human umbilical vein endothelial cell line, we found that parathyroid hormone increased both intracellular calcium and cellular cAMP content in these endothelial cells. Furthermore, exposure of these cells to increasing concentrations of parathyroid hormone stimulated both [(3)H]thymidine incorporation and endothelin-1 secretion. Parathyroid hormone/parathyroid hormone-related peptide receptor mRNA could be detected at low levels in these cells. In summary, these data demonstrate that endothelium-derived cells contain functional parathyroid hormone receptors. The potential physiological role of these receptors remains to be determined.     

Possible evidence for the existence of a growth hormone fragment in porcine pituitary

In an attempt to search for growth hormone fragments in the pituitary, a radioimmunoassay was developed for a 55 residue S-amino-ethylated CNBr fragment (fragment B) of porcine growth hormone corresponding to residues 126–180 of human growth hormone. The assay was sensitive to 50 pg of fragment B whereas displacement of ¹²⁵I-labelled fragment B porcine growth hormone required a 10³ M excess and was non-parallel. In a homogolous porcine growth hormone radioimmunoassay, fragment B was non-reactive. Gel filtration of an extract of porcine pituitary on Sephadex G-75 revealed three peaks of fragment B immunoreactivity: peak I (29% of total immunoreactivity) eluted in the void volume, peak II (49%) eluted in the position of growth hormone, and peak III (12%) was more retarded than fragment B. Nearly all of the growth hormone immunoreactivity eluted as a single peak in the position of ¹²⁵I-labeled porcine growth hormone. The dilution curve of peak III but not of peaks I or II was parallel to that of fragment B. The results indicate the existence within porcine pituitary of material cross-reactive with a portion of the growth hormone molecule, possibly representing a growth hormone fragment.     

Psychosocial adaptation to short stature - an indication for growth hormone therapy?

Although growth hormone does not clearly improve final height in non-growth-hormone-deficient children with short stature, it leads to a temporary acceleration of growth velocity. It is an ongoing discussion whether this effect supports psychosocial adaptation to short stature and therefore could be an indication for growth hormone treatment in children with short stature without growth hormone deficiency. We have reviewed recent literature concerning psychosocial consequences of short stature. Together with own data we can demonstrate that short people regularly adapt well to their height and have a good self-esteem. On the other hand, we focus on the problem that most studies on this subject suffer from methodical problems. A growth-related questionnaire that evaluates subjective and objective perceptions of being short in patients and peers is not at hand. As a consequence, psychosocial problems due to short stature have not been exactly classified yet and therefore do not represent an indication for growth hormone therapy.     

Pre-proparathyroid hormone: fidelity of the translation of parathyroid messenger RNA by extracts of wheat germ.

Pre-proparathyroid hormone is the major protein synthesized in wheat-germ extracts in response to addition of an 8-15S fraction of parathyroid RNA. The accuracy of the translation of the mRNA from parathyroid tissue was examined by analysis of the carboxyl-terminal tryptic peptide and the amino-terminal amino acid of the protein, by analysis of the size distribution of the mRNA, and by translation of the mRNA in a second cell-free extract. When 8-15S RNA was fractionated on a sucrose gradient containing formamide, RNA that supported the synthesis of pre-proparathyroid hormone was present in a single symmetrical peak, suggesting that it was homogeneous. Analyses by paper chromatography and electrophoresis of the proline-containing tryptic peptides of pre-proparathyroid hormone indicate that they are identical with the corresponding proline-containing peptides of parathyroid hormone. Because the COOH-terminal tryptic peptide of parathyroid hormone contains proline, the data indicate that the COOH termini of pre-proparathyroid hormone and parathyroid hormone are identical. Methionine from initiator [35S]Met-tRNAfMet was rapidly incorporated into pre-proparathyroid hormone by the wheat-germ extract, and a single-step Edman degradation selectively removed almost all of the initiator [35S]methionine present in pre-proparathyroid hormone. Translation of the 8-15S RNA in a cell-free extract from Krebs-II ascites cells resulted in a protein that comigrated with pre-proparathyroid hormone on sodium dodecyl sulfate-acrylamide gel electrophoresis. These data support the conclusion that the wheat-germ system accurately translates the mRNA for parathyroid hormone, and they strengthen the contention that pre-proparathyroid hormone is the initial biosynthetic product.     

Hormone-induced changes in the in vitro DNA-binding activity of the chicken progesterone receptor

Previous analyses have indicated that steroid hormone receptors undergo an allosteric change in structure upon binding by the steroid ligand. This structural change was envisioned as an intramolecular unmasking of the protein's DNA-binding domain, thus allowing the receptor to function in gene regulation. We report an analysis of the effect of hormone on the DNA-binding activity of the chicken progesterone receptor. Using an isocratic elution of DNA affinity columns we show that unliganded receptor (aporeceptor) can bind a 23-basepair progesterone response element with high affinity and a high degree of sequence preference. Hormone causes a 1.5-fold increase in affinity for the PRE sequence and a 2-fold decrease in affinity for non-specific DNA. Kinetic analysis of the off-rate of receptor-DNA complexes is consistent with this minor effect of hormone. In addition, gel retardation analysis of receptor-progesterone response element complexes further substantiates that hormone is not required for sequence-specific DNA binding. These results indicate that hormone is not necessary for the progesterone receptor to fold into a conformation that recognizes specific gene regulatory sequences.