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ABSTRACT

Pyroglutamylglutamylprolineamide (pGlu-Glu-ProNH₂) is a tripeptide with structural and immunological similarities to thyrotrophin-releasing hormone (TRH; pGlu-His-ProNH₂). Since TRH stimulates GH secretion in domestic fowl, the possibility that pGlu-Glu-ProNH₂ may also provoke GH release was investigated. Unlike TRH, pGlu-Glu-ProNH₂ alone had no effect on GH release from incubated chicken pituitary glands and did not down-regulate pituitary TRH receptors. However, pGlu-Glu-ProNH₂ suppressed TRH-induced GH release from pituitary glands incubated in vitro and competitively displaced [³H]methyl³-histidine²-TRH from pituitary membranes. Systemic injections of pGlu-Glu-ProNH₂ had no significant effect on basal GH concentrations in conscious birds, but promptly lowered circulating GH levels in sodiumpentobarbitone anaesthetized fowl. Submaximal GH responses of conscious and anaesthetized birds to systemic TRH challenge were, however, potentiated by prior or concomitant administration of pGlu-Glu-ProNH₂. These results demonstrate, for the first time, that pGlu-Glu-ProNH₂ has biological activity, with inhibitory and stimulatory actions within the avian hypothalamo-pituitary axis. These results indicate that pGlu-Glu-ProNH₂ may act as a TRH receptor antagonist within this axis.

Journal of Endocrinology (1993) 138, 137–147

ABSTRACT

Pyridoxine-deficient young rats (3 weeks old) had significantly reduced levels of pituitary TSH, serum thyroxine (T₄) and tri-iodothyronine (T₃) compared with pyridoxine-supplemented rats. The status of the pituitary-thyroid axis of normal, pyridoxine-supplemented and pyridoxine-deficient rats was evaluated by studying the binding parameters of [³H](3-methyl-histidine²)TRH in the pituitary of these rats. The effects of TRH and T₄ injections on pituitary TSH and serum TSH, T₄ and T₃ of these two groups were also compared. The maximal binding of TRH receptors in the pituitary of pyridoxine-deficient rats was significantly higher than that of pyridoxine-supplemented control and normal rats, but there was no change in the binding affinity. Treatment with TRH stimulated TSH synthesis and release. It also increased serum T₄ and T₃ in both pyridoxine-supplemented and pyridoxine-deficient rats. Treatment with T₄ decreased serum and pituitary TSH in both pyridoxine-supplemented and pyridoxine-deficient rats, compared with saline-treated rats. The increased pituitary TRH receptor content, response to TRH administration and the fact that regulation at the level of the pituitary is not affected in the pyridoxine-deficient rat indicates a hypothalamic origin for the hypothyroidism of the pyridoxine-deficient rat.

J. Endocr. (1986) 109, 345–349

Abstract

1,25-Dihydroxyvitamin D₃ (1,25-(OH)₂D₃) is active in primary dispersed and clonal pituitary cells where it stimulates pituitary hormone production and agonist-induced hormone release. We have studied the effect of 1,25-(OH)₂D₃ on thyrotropin-releasing hormone (TRH) binding in clonal rat pituitary tumour (GH₃) cells. Compared with vehicle-treated cells, 1,25-(OH)₂D₃ (10 nmol/l) increased specific [³H]MeTRH binding by 26% at 8 h, 38% at 16 h, 35% at 24 h and reached a maximum at 48 h (90%). In dose–response experiments, specific [³H]MeTRH binding increased with 1,25-(OH)₂D₃ concentration and reached a maximum at 10 nmol/l. Half-maximal binding occurred at 0·5 nmol 1,25-(OH)₂D₃/l. The vitamin D metabolite, 25-OH D₃, increased [³H]MeTRH binding but was 1000-fold less potent than 1,25-(OH)₂D₃. In equilibrium binding assays, treatment with 10 nmol 1,25-(OH)₂D₃/l for 48 h increased the maximum binding from 67·4 ± 8·8 fmol/mg protein in vehicle-treated cells to 96·7 ± 12·4 fmol/mg protein in treated cells. There was no difference in apparent K_d (1·08 ± 0·10 nmol/l for 1,25-(OH)₂D₃-treated and 0·97 ± 0·11 nmol/l for vehicle-treated cells). Molecular investigations revealed that 10 nmol 1,25-(OH)₂D₃/l for 24 h caused an 8-fold increase in TRH receptor-specific mRNA. Actinomycin D (2 μg/ml, 6 h) abrogated the 1,25-(OH)₂D₃-induced increase in [³H]MeTRH binding. Cortisol also increased [³H]MeTRH binding but showed no additivity or synergism with 1,25-(OH)₂D₃. TRH-stimulated prolactin release was not enhanced by 1,25-(OH)₂D₃. We conclude that the active vitamin D metabolite, 1,25-(OH)₂D₃, caused a time- and dose-dependent increase in [³H]MeTRH binding. The effect was vitamin D metabolite-specific and resulted from an upregulation of the TRH receptor. Further studies are needed to determine the functional significance of this novel finding.

Journal of Endocrinology (1995) 147, 397–404

ABSTRACT

The effects of tri-iodothyronine (T₃) and TRH on prolactin mRNA accumulation in monolayer pituitary cell cultures prepared from both euthyroid and hypothyroid rats were investigated. Basal prolactin mRNA concentrations and prolactin release into culture medium were increased in hypothyroid cultures, the increase being related to the duration of hypothyroidism in vivo. The inhibitory effects of T₃ seen in euthyroid cells were preserved in cells derived from hypothyroid animals, and the degree of inhibition was greater in cells from the most severely hypothyroid rats. However, the stimulation of prolactin synthesis and secretion induced by TRH in euthyroid cultures was not found in the hypothyroid cells. Hypothalamic and anterior pituitary TRH content were measured in similarly hypothyroid and euthyroid rats. A large hypothalamic pool of TRH was found, which was unchanged in hypothyroidism, whereas anterior pituitary TRH content was increased in the hypothyroid rats. The consequent down-regulation of anterior pituitary TRH receptors may explain the poor response of prolactin to TRH seen in vitro.

J. Endocr. (1987) 115, 497–503

Abstract

Pyroglutamylglutamylprolineamide, which was first discovered in mammalian prostate, differs from thyrotrophin-releasing hormone (TRH) by substitution of glutamic acid for histidine at position two of the tripeptide. Recently, the newly discovered peptide has been identified in substantial concentrations in the rat anterior pituitary gland and, in this study, we have investigated the effects of the peptide on rat anterior pituitary cells in culture. GH₃ cells were chosen to examine the possible effects of the new peptide, particularly in relation to its effects on the TRH receptor. This cell-type was deficient, in comparison with normal rat pituitary cells, in the new TRH-related peptide and appeared to be an ideal model cell in which to study the effects of pGlu-Glu-ProNH₂. TRH (0·01–100 nm) was found to stimulate the secretion of both GH and prolactin from GH₃ cells whereas pGlu-Glu-ProNH₂ had no effect within the same concentration ranges. In contrast, at micromolar concentrations pGlu-Glu-ProNH₂ exhibited intrinsic TRH-like activity causing stimulation of both GH and prolactin release from GH₃ cells. Both TRH and pGlu-Glu-ProNH₂ appeared to act through the same intracellular signalling mechanism, causing significant increases in intracellular inositol phosphate within the expected concentration ranges. However, pGlu-Glu-ProNH₂ (up to 1 mm) displaced neither [³H]TRH nor [³H]MeTRH from membrane-binding sites on GH₃ cells, suggesting that the effects of the new peptide were mediated through a second receptor. The physiological relevance of these effects of pGlu-Glu-ProNH₂ requires further investigation.

Journal of Endocrinology (1994) 142, 111–118

ABSTRACT

The activity of ornithine decarboxylase (ODC) in the rat anterior pituitary gland varies during the oestrous cycle, with a rise in activity seen at pro-oestrus. This enzyme, which is rate-limiting for the synthesis of the polyamines, can be specifically and irreversibly blocked by α-difluoromethylornithine (DFMO). A previous study showed that when this drug was administered to rats in vivo on the afternoon of pro-oestrus, it suppressed the normal surge in plasma prolactin levels that occurred later that day. The effect of DFMO was associated with reduced levels of putrescine in the anterior pituitary gland, suggesting that ODC activity in the lactotroph might be involved in the prolactin surge. We have examined the effects of DFMO on the secretion of prolactin from anterior pituitary cells, isolated either from male rats or from females at different stages of the oestrous cycle. The drug was found to reduce prolactin secretion stimulated by thyrotrophin-releasing hormone (TRH), but only in cells isolated from pro-oestrous animals and only for 2 days after cell isolation. Basal secretion was unaffected by DFMO. The results imply that ODC is important for TRH-stimulated prolactin secretion at pro-oestrus, and it is specific for pro-oestrus. The prolactin surge could therefore be influenced by this ODC-dependent effect of TRH. The pro-oestrous-specific response to TRH may be a consequence of the increased ODC activity seen at this time. Alternatively, the increased ODC activity could be a consequence of coupling to TRH receptors, which are known to increase in number at pro-oestrus.

Journal of Endocrinology (1993) 137, 133–139

Abstract

The reduced thyroid activity during short-term starvation is associated with a lowered hypothalamic synthesis and secretion of TRH. However, little is known about the cause of the reduced thyroid function during prolonged malnutrition. We have therefore studied the effects of food reduction to one-third of normal (FR33) on the hypothalamus-pituitary-thyroid axis of male and female Wistar rats. After 3 weeks body weights of FR33 rats were almost 50% lower than those of controls. In both sexes, FR33 caused marked increases in serum corticosterone, and decreases in serum TSH, thyroxine (T₄), free T₄, tri-iodothyronine (T₃) and free T₃. While the free T₃ fraction (FFT₃) in serum decreased, the free T₄ fraction (FFT₄) tended to increase. Electrophoretic analysis indicated that decreased FFT₃ was correlated with an increased thyroxine-binding globulin, while the increase in FFT₄ seemed due to a decreased thyroxine-binding prealbumin binding capacity. Total RNA and proTRH mRNA in the hypothalamus were not affected by FR33. Median eminence and posterior pituitary TRH content tended to increase in FR33 rats, suggesting that hypothalamic TRH release is reduced in FR33 rats. Anterior pituitary TSH content was decreased by FR33 in both sexes, but pituitary TSHβ mRNA and TRH receptor status were not affected except for increased pituitary TSHβ mRNA in female FR33 rats. Although FR33 had no effect on pituitary weight, pituitary RNA and membrane protein content in FR33 rats were 50–70% lower than values in controls.

In conclusion, prolonged food reduction suppresses the pituitary-thyroid axis in rats. In contrast to short-term food deprivation, the mechanism whereby serum TSH is suppressed does not appear to involve decreases in proTRH gene expression, but may include effects on pituitary mRNA translation. Our results further support the hypothesis that TSH release may be lowered by increased corticosterone secretion, although the mechanism of this effect may differ between acute starvation and prolonged food reduction.

Journal of Endocrinology (1996) 150, 169–178