Effect of growth hormone-releasing factor on phosphoinositide hydrolysis in somatotrophs

doi:10.1016/0303-7207(90)90146-Y

Molecular and Cellular Endocrinology

Volume 72, Issue 3, 10 September 1990, Pages 221-226

https://doi.org/10.1016/0303-7207(90)90146-Y Get rights and content

Abstract

We studied the role of the phosphatidylinositol system in the action of growth hormone-releasing factor (GRF). We asked whether GRF stimulates the activity of phospholipase C by determining GRF-induced changes in ³²P labeling of the individual phosphoinositides and inositol phosphates in purified rat somatotrophs. The somatotrophs were challenged with GRF (10⁻⁷M) for 0.33, 1, 3, 10, 30, and 90 min. GRF did not significantly or consistently alter ³²P incorporation into phosphatidylinositol bisphosphate (PIP₂), phosphatidylinositol monophosphate (PIP), or phosphatidylinositol (PI), except for a small reduction in PIP labeling at 90 min. In general the level of ³²P incorporation into the inositol phosphates did not increase but instead decreased with GRF. There was a small but significant reduction of labeling of inositol trisphosphate (IP₃) at 90 min of GRF incubation. There were also small but significant decreases in ³²P incorporation into inositol bisphosphate (IP₂) at 0.33, 3, and 30 min. GRF did not significantly alter ³²P labeling of inositol monophosphate (IP). These results indicate that GRF does not stimulate phospholipase C activity in somatotrophs. We conclude that the phosphatidylinositol second messenger system does not play an essential role in the action of GRF.

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Arginine (Arg) presents a potent growth hormone (GH) releasing activity. In vivo and in vitro studies carried out in our laboratory have demonstrated that acute treatment with Arg also increases GH gene expression. Taking into account the recognizable diabetogenic role of GH and that Arg increases insulin release, this study aimed at evaluating the effects of oral chronic administration of Arg on GH gene expression, by Northern blotting analysis, and on the insulin sensitivity, by means of the Insulin Tolerance Test (ITT), blood glucose decay rate (kitt) and insulin plasma concentration. We demonstrated that rats that consumed Arg (∼ 35 mg/day) in drinking water, during 4 weeks, presented an increase in GH mRNA content (p < 0.01), a decreased peripheral response to insulin, as shown by the reduced blood glucose decay rate (p < 0.05), and a higher insulin plasma concentration (p < 0.01) than control group. Arg treatment did not modify the animals' food and water intake, while it decreased the heart rate and the arterial blood pressure compared to control group (p < 0.05). According to the results presented herein we conclude that chronic oral administration of arginine increases GH gene expression and induces insulin resistance. The arterial blood pressure decrease has already been pointed out in the literature, and seems to occur in response to the dilating effect of nitric oxide generated from Arg, as well as from NO generation in central and peripheral neuronal populations that express NOS and are involved in the autonomic regulation of the cardiac function.
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This chapter describes growth hormone (GH) as a polypeptide secreted by the somatotroph cells of the anterior pituitary gland, is the primary stimulator of linear growth in vertebrates and is involved in the regulation of protein and fat metabolism. GH also plays an important role in the control of immune function, gonadal steroidogenesis, and bone development. The actions of GH are mediated via the GH receptor, and major target tissues include the liver, muscle, and adipose tissue, where GH acts directly to stimulate cell proliferation and differentiation. The chapter discusses that the synthesis and secretion of GH from pituitary somatotroph cells is tightly regulated by several factors, including neurotransmitters such as GABA, NPY, and the opiods; peptide hormones such as ghrelin and IGF-1; and steroid hormones including thyroid hormone and the glucocorticoids. The hypothalamic neuropeptides growth hormone-releasing hormone (GHRH) and somatostatin (SRIF) are the predominant stimulatory and inhibitory neuroendocrine signals, respectively, for GH secretion by pituitary somatotroph cells. Observations from both clinical studies and animal model systems confirm that GHRH is an important regulator of GH secretion and linear growth.
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Porcine somatotropes can be separated by Percoll density gradient centrifugation into low (LD) and high density (HD) subpopulations that differ ultrastructurally and functionally. Here, we report the effects of growth hormone-releasing factor (GRF) on the cytosolic free calcium concentration ([Ca²⁺]_i) of single LD and HD somatotropes. Resting [Ca²⁺]_i in LD somatotropes was 2-fold higher than in HD cells. GRF induced [Ca²⁺]_i increases in a similar percentage of somatotropes from both subsets. However, amplitude and kinetics of the responses were markedly different. In all responsive LD somatotropes, GRF evoked a rapid initial peak followed by a sustained plateau (plateau-type response). Blockade of extracellular Ca²⁺ entry by 3 mM EDTA, 2 mM CoCl₂, or 100 μM verapamil completely abolished the plateau phase without affecting the initial Ca²⁺ spike. Conversely, only the plateau phase was preserved in thapsigargin (TG)-treated LD cells. The vast majority of GRF-responsive HD somatotropes exhibited a transient [Ca²⁺]_i peak that returned gradually to baseline (transient-type response). This response was completely blocked by removal of extracellular Ca²⁺, whereas TG treatment had no effect. Taken together, our results indicate that the response of LD somatotropes to GRF depends on mobilization of Ca²⁺ of both extra- and intracellular origin, whereas that of HD somatotropes seems to be exclusively dependent on extracellular Ca²⁺ entry through L-type voltage sensitive Ca²⁺ channels (VSCC). These findings are the first to demonstrate a differential effect of GRF on Ca²⁺ mobilization in two somatotrope subpopulations, and suggest the existence of differences in the GRF receptor(s) expressed in each subpopulation and/or in the intracellular signalling pathways activated upon GRF binding.
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The purpose of this study was to investigate the involvement of protein kinase C in growth hormone-releasing factor (GRF) action by directly measuring the effect of GRF on protein kinase C activity in purified male rat somatotrophs. Somatotrophs were incubated with GRF (10⁻⁷ M) for 0.33, 1, 3, 10, 30 and 90 min. Protein kinase C present in soluble and paniculate fractions was partially purified using DEAE-cellulose chromatography, and protein kinase C activity was assayed. In control experiments, to insure protein kinase C activity could be activated, two known protein kinase C activators, phorbol 12-myristate 13-acetate (PMA) and dioctanoyl-rac-glycerol (diC₈) were added for 3 min.
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^☆: This work was supported by MRC of Canada Grant MT-1634.

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Effect of growth hormone-releasing factor on phosphoinositide hydrolysis in somatotrophs☆

Abstract

Life Sci.

Methods Enzymol.

Mol. Cell. Endocrinol.

Mol. Cell. Endocrinol.

Mol. Cell. Endocrinol.

Eur. J. Pharmacol.

Endocrinology

Endocrinology

Nature

Nature

Am. J. Physiol.

Endocrinology

Acta Endocrinol.

Endocrinology

Endocrinology

Endocrinol. Jpn.