Terry Taylor

Ontogeny of thyrotropin-releasing hormone gene expression in the rat diencephalon.

The development of thyrotropin-releasing hormone (TRH) gene expression in the rat diencephalon was studied using in situ hybridization histochemistry. The first neurons expressing the TRH gene were found on gestational day 14 (E14) in the lateral hypothalamus, shortly after completion of their last cell division. On E15 and E16, additional labeled cells appeared medially in the developing dorsomedial and paraventricular nuclei, respectively, followed on E17 by cells in the preoptic area. The number of labeled cells in the hypothalamus continued to increase during the last part of gestation. At birth, TRH mRNA neurons were present in all the locations seen in the adult hypothalamus. During the first week of life, the labeling intensity and number of neurons containing TRH mRNA continued to increase at the locations described above. TRH mRNA was not detected in the reticular thalamic nucleus until the 7th postnatal day when some labeled cells appeared in its dorsocaudal portion. Over the next 10 days, the number of labeled cells and the intensity of labeling in the thalamic reticular nucleus progressively increased. During the same period of time, only small changes in the number and intensity of labeled cells in the hypothalamus were seen. On the 21st postnatal day, after a decrease in labeling in the lateral hypothalamus had been noted, final adult patterns of expression were present. With the very sensitive and anatomically specific method of in situ hybridization histochemistry, the ontogeny of TRH gene expression in the rat diencephalon has been elucidated.

Glycosylation and posttranslational processing of thyroid-stimulating hormone: clinical implications.

Publisher Summary This chapter discusses clinical implications of glycosylation and posttranslational processing of thyroid-stimulating hormone (TSH). TSH is a glycoprotein hormone of molecular weight 28,000 which is composed of two noncovalently linked subunits, α and β . It is chemically related to the pituitary gonadotropins, luteinizing hormone (LH), follicle-stimulating hormone (FSH), and to chorionic gonadotropin (CG). Elevated serum α /TSH ratios have been described in patients with TSH-secreting pituitary tumors. The excess production from such tumors has been of value in differentiating patients with neoplastic from non-neoplastic causes of TSH-induced hyperthyroidism. In addition, isolated production of α subunit, without concomitant production of TSH or gonadotropins, by certain pituitary adenomas and other malignant tumors is demonstrated. Elevated serum TSH- β /TSH ratios are described in two patients, one with an enlarged thyroid and one with an enlarged pituitary. The TSH- β in both cases have large molecular weight, display immunologic properties different from standard TSH- β , and is unresponsive to thyrotropin releasing hormone. Although this unusual form of TSH- β has been partially characterized, its significance remains unknown. The chapter also describes unusual forms of human TSH with decreased bioactivity. One apparently normal subject was found to have a high-molecular-weight form of TSH with normal receptor-binding properties but decreased bioactivity. Such forms can result from aggregated or protein-bound TSH caused by abnormalities of glycosylation, similar to those noted after tunicamycin treatment.

Thyroid Hormone Regulation of TRH mRNA Levels in Rat Paraventricular Nucleus of the Hypothalamus Changes during Ontogeny

The changing roles of the hypothalamus and pituitary in regulating thyroid hormone levels in the rat during ontogeny has not been fully elucidated. It has been reported that endogenous TRH begins to stimulate TSH secretion at 5-8 days after birth but that the pituitary responds to hypothyroidism during late gestation. To determine the onset and extent of TRH response to low thyroid hormone levels during ontogeny, normal and hypothyroid rats treated with methimazole for 7 days were sacrificed at 16 days gestation (E16), 20 days gestation (E20), 7, 21 and 56 days after birth (n = 5/study group). Plasma hormones were assayed from pregnant mothers, pups (pooled) and adults. Levels of TRH mRNA were measured in the paraventricular nuclei (PVN) by in situ hybridization histochemistry. A labeled 48-base cDNA oligonucleotide for TRH was hybridized with brain slices (n = 6/animal) in the region of the medial parvocellular division of the PVN of the hypothalamus and the signal was quantitated by digitized computer analysis. Plasma-free T4 levels decreased and plasma TSH levels increased in the animals treated with methimazole as compared to the euthyroid controls. TRH mRNA was detected in the PVN at E16 after brain slices were dipped in emulsion and granules observed by dark-field microscopy. In the euthyroid animals, TRH mRNA increased from E20 (150 +/- 9 OD units) to 7 days (222 +/- 5 OD units) and remained unchanged at 21 days (252 +/- 27 OD units) and 56 days (244 +/- 6 OD units).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of in vivo bolus versus continuous TRH administration on TSH secretion, biosynthesis, and glycosylation in normal and hypothyroid rats ☆

The effects of in vivo TRH administered either as bolus or continuous doses on TSH secretion, synthesis, and glycosylation were studied in normal and hypothyroid rats. Nine-week-old normal or 3-week postthyroidectomy rats were administered bolus doses of saline or TRH (0.5 mg/kg) twice daily or continuous saline or TRH (1 mg/kg/day) via an osmotic pump. After 5 days, pituitaries were removed and incubated with [35S]methionine (MET) and [3H]glucosamine (GLCN), with or without 10(-8) M TRH, for 6 and 24 h. Samples were precipitated with anti-TSH beta sera and then analyzed by gel electrophoresis. In normal rats, plasma TSH, T4 and T3 increased with continuous in vivo TRH but not with bolus TRH; in hypothyroid rats, plasma TSH, T4 and T3 were not altered by continuous or bolus doses of TRH. Additionally, in normal rats, continuous in vivo TRH increased incorporation of MET in secreted TSH (477 vs. 212 X 10(3) dpm/mg DNA; P less than 0.05) and intrapituitary TSH (5035 vs. 2124 X 10(3) dpm/mg DNA; P less than 0.05), and GLCN in secreted TSH (148 vs. 50 dpm/mg DNA; P less than 0.05) and intrapituitary TSH (2344 vs. 744 X 10(3) dpm/mg DNA; P less than 0.05). In contrast, in hypothyroid animals, continuous in vivo TRH did not alter MET or GLCN incorporation in TSH. Bolus TRH did not alter secreted or intrapituitary MET or GLCN incorporation into TSH in the normal rat. However, bolus TRH in the intrapituitary MET or GLCN incorporation into TSH in the normal rat.(ABSTRACT TRUNCATED AT 250 WORDS)

Pre-Translational and Post-Translational Regulation of TSH Synthesis in Normal and Neoplastic Thyrotrophs

We are interested in the mechanisms by which endocrine and developmental factors regulate TSH synthesis at both pre-translational and post-translational levels. Thyroid hormone profoundly decreases transcription of the TSH-beta gene, while TRH and agents modifying cyclic AMP increase transcription. To elucidate the molecular mechanisms underlying these effects, human embryonal kidney cells were transfected with constructs of the human TSH-beta gene fused to the chloramphenicol acetyltransferase gene. The first exon of human TSH-beta, contains an element that increases basal expression and mediates T3-induced gene repression, probably through a direct interaction with c-erbA beta. This transcriptional repression by T3 appears aberrant in thyrotropic tumors. In contrast, TRH and agents modifying cyclic AMP mediate increased transcription of TSH-beta through interacting with upstream regulatory elements. Thyroid hormone, TRH and developmental factors also regulate the branching pattern and relative sialylation of TSH carbohydrate chains, which may affect TSH action in vitro and in vivo. Certain thyrotropic tumors produce TSH with more complex carbohydrate branching patterns, which may increase its biologic activity.

Comparison of a Pituitary TSH-Secreting Micro- Versus Macroadenoma

While more than 40 cases of TSH-secreting pituitary adenomas have been reported in the medical literature (1,2), little attention has been devoted to structural variability—in particular that due to carbohydrate microheterogeneity—of thyrotropin molecules derived from different tumors and distinct physiological states. We recently evaluated two patients representing extremes within the spectrum of TSH-secreting pituitary adenomas and have contrasted several biochemical features of these tumors.