Methimazole interferes with the progression of experimental autoimmune myocarditis in rats

We produced experimental autoimmune hypophysitis (EAH) in rats and investigated its characteristics. Female Lewis rats were immunized by two injections with homologous pituitary homogenate and complete Freund's adjuvant. Blood was collected serially from the rats, and serum antibodies to pituitary antigens were examined. The rats were sacrificed 2 or 4 weeks after the final immunization, and histological examinations of the endocrine organs were carried out. Histological examination revealed slight, focal infiltration of mononuclear cells in the pituitary gland only in the rats immunized with the pituitary homogenate. Infiltration of mononuclear cells was not observed in the thyroid gland, pancreas, adrenal gland, or ovary. In the serological examination, antibodies to both cytosolic antigens and cytoplasmic particle antigens from the pituitary gland were detected by enzyme-linked immunosorbent assay (ELISA), and these antibody levels increased with time. Western blotting using the serum antibodies identified an immunoreactive protein of approximately 21.5 kDa among these antigens, and we confirmed that this protein was rat growth hormone (GH). Furthermore, antibodies to GH, thyrotropin (TSH), and luteinizing hormone (LH) were detected by ELISA. Antibodies to follicule stimulating hormone, prolactin, or adrenocorticotropin were not detected. These data suggest that several antigens from the pituitary gland are involved in EAH in rats, and that GH, TSH, and LH are major antigens among the pituitary antigens in this model.     

Effect of diphenylhydantoin on hypothalamic-pituitary-thyroid function in the rat

Chronic diphenylhydantoin (DPH) administration (5 mg x 100 g body wt-1 x day-1) to the normal rat is associated with a decrease in the serum thyroxine (T4) and triiodothyronine (T3) concentrations without an appropriate rise in the serum thyrotropin (TSH) concentration, suggesting a possible direct effect of DPH on TSH secretion. To further study this possibility, DPH was administered chronically to thyroidectomized, hypothyroid rats. In the hypothyroid rats treated chronically with DPH, serum TSH did not increase, pituitary TSH content was significantly decreased, and the serum TSH response to thyrotropin-releasing hormone (TRH) was decreased compared to that of diluent-treated, hypothyroid rats. Hypothalamic TRH content was similar in DPH and diluent-treated rats. These findings suggest that DPH suppresses pituitary TSH secretion, probably as a thyroid hormone agonist. The effect of a single large dose of DPH (20 mg/100 g body wt) administered to thyroidectomized rats also decreased serum tSH but, in contrast to the findings in chronically treated rats, hypothalamic TRH and pituitary TSH content and the serum TSH responses to TRH were increased. These differences may be due to the acute inhibitory effect of a large dose of DPH on hypothalamic TRH release. Furthermore, because the effect of thyroid hormone on regulating pituitary TSH synthesis and release is dose and time dependent, the effect of DPH as a thyroid hormone agonist on pituitary TSH dynamics may also be variable.     

Hypothalamus-pituitary-thyroid axis interactions with pineal gland in the rat.

We examined in the rat several possible relationships between the pineal gland and the hypothalamus-pituitary-thyroid axis. The pineal gland, the retina, and the hypothalamus exhibited a diurnal rhythm in thyrotropin-releasing hormone (TRH) content with peak values occurring around 1200 h. This rhythm in the hypothalamus was abolished by constant light but was not affected by pinealectomy. Nor did pinealectomy affect hypothalamic TRH content, pituitary content of thyroid-stimulating hormone (TSH) or prolactin; serum levels of (TSH), triiodothyronine (T3), or thyroxine (T4), or serum free-thyroxine index; or free-triiodothyronine index. Melatonin did not affect TSH or prolactin release from the anterior pituitary or TRH release from the hypothalamus in vitro. Isoproterenol did not affect the TRH content of pineal glands in vitro; nor did TRH or T3 affect basal or stimulated activities of serotonin N-acetyltransferase, the presumed controlling enzyme in melatonin production. We found no evidence for significant interactions between the pineal gland and the hypothalamus-pituitary-thyroid axis.     

Cytodifferentiation and immunocharacteristics of adenohypophysial cells in the toad, Bufo melanostictus

Summary Employing the unlabelled antibody enzyme technique cytodifferentiation, immunocharacteristics and topographical distribution of melanotropic (MSH), adrenocorticotropic (ACTH), thyrotropic (TSH), prolactin (PRL), gonadotropic (GTH) and growth hormone (GH) secreting cells in the embryonic/larval as well as adult pituitary gland of the common Indian toad, Bufo melanostictus, have been studied by using antisera raised in rabbit against mammalian hypophysial hormones. Immunoreactive MSH and ACTH cells appear first in the dorsocaudal and rostral regions of the pituitary anlage (PA) at stage 21 (Gosner's classification) of the embryonic development. This is followed by the differentiation of TSH and PRL cells at stage 22 in the midventral and central regions of the PA respectively. Finally, at stage 23 the GTH cells appear in the fostral and the GH cells in the caudal regions of the PA. With the progress of the development, cells showing immunoreactivity to various antisera gradually increase in number, size, granular content and finally occupy the characteristic adult disposition. The MSH cells comprise the pars intermedia. In the pars distalis (PD) the ACTH cells are localized in the rostroventral region, TSH cells in the central region and the GH cells in the dorsocaudal region. However, GTH and PRL cells are distributed throughout the PD.Dedicated to Late Dr. Maurice P. Dubois     

In Situ Hybridization Method Reveals (Pro)renin Receptor Expressing Cells in the Pituitary Gland of Rats: Correlation with Anterior Pituitary Hormones

Expression of (pro)renin receptor ((P)RR), a specific receptor for renin and prorenin, was studied in rat pituitary gland. In situ hybridization showed that cells expressing (P)RR mRNA were widely distributed in the anterior lobe and intermediate lobe of the pituitary gland. Double-staining using in situ hybridization for (P)RR mRNA and immunohistochemistry for the pituitary hormones showed that (P)RR mRNA was expressed in most of the GH cells and ACTH cells in the anterior lobe. (P)RR mRNA was also expressed in a few prolactin cells and TSH cells, but not in LH cells. The present study has shown for the first time the distribution of (P)RR mRNA expressing cells in the rat pituitary gland. These findings suggest that (P)RR plays physiological roles in the pituitary gland, such as the modulation of the pituitary hormone secretion.     

In vitro TSH and PRL secretion from eutopic and heterotopic rat pituitaries: effects of hypothyroidism

Five adenohypophyses from donors of the same strain, age, and sex were transplanted under the renal capsule of young adult female rats. At least 3 wk later, enzymatically dispersed cells from eutopic or heterotopic adenohypophyses from the same rat were perifused in vitro in a small chamber. Thyrotropin (TSH) and prolactin (PRL) secretion per 10(6) cells were significantly less from heterotopic than from eutopic cells under all conditions. In cells from euthyroid animals, TRH induced TSH secretion only in the eutopic cells but induced PRL secretion in both eutopic and heterotopic cells. Hypothyroidism increased TRH-induced TSH secretion and content in the cell lysate in both eutopic and heterotopic cells but increased TRH-induced PRL secretion only in the eutopic cells. The increase in TSH secretion induced by hypothyroidism in the heterotopic cells was of borderline statistical significance. The inability of TRH to induce TSH secretion in heterotopic pituitary cells from euthyroid rats may be due to a lower set point for thyroid hormone inhibition of TSH secretion in the heterotopic thyrotrophs. Heterotopic pituitary TSH secretion is probably suppressed by the normal plasma thyroid hormone concentration maintained by the eutopic pituitary and may be stimulated by TRH only in the presence of a subnormal plasma thyroid hormone concentration.     

Immunoreactive TSH cells in juvenile and peripubertal rats after estradiol and human chorionic gonadotropin treatment

Different hormones and growth factors control the homeostasis of the anterior pituitary gland. We examined the morphological features of pituitary thyroid stimulating hormone (TSH)-producing cells in juvenile and peripubertal female rats after treatments with estradiol-dipropionate (EDP), human chorionic gonadotropin (hCG) and a combination of both hormones (EDP+hCG). TSH-producing cells were labelled using a peroxidase-antiperoxidase immunohistochemical procedure for binding of a rabbit anti-rat beta-thyrotropic polyclonal antisera. Morphometric differences in the cytoplasmic and nuclear volume densities in thyrotropes after the treatments were determined using a stereological method. The relative weights of the pituitary glands were significantly higher in the EDP- and EDP+hCG-treated juvenile and peripubertal rats than in untreated age-matched controls. Treatment with EDP promoted a decrease, and treatment with hCG an increase, of the cellular and nuclear volumes in TSH cells in both juvenile and peripubertal females in comparison with the respective controls. Treatment with a combination of EDP+hCG did not induce any significant changes. The cytoplasmic and nuclear volume densities in TSH cells in the EDP+hCG-treated group were significantly higher than in the EDP-treated, and significantly lower than in hCG-treated rats at both growth stages. These findings suggest that estradiol and hCG exerted opposite effects on pituitary TSH-immunoreactive cells. The observed effects on thyrotrope morphology were apparently independent of the stage of development.     

Pituitary adenomas. An immunohistochemical study of hormone production and chromogranin localization.

Tumors from 42 surgically resected pituitaries and from 13 autopsy cases were studied immunohistochemically with polyclonal antisera to 7 anterior pituitary hormones and with a newly developed monoclonal antibody directed against human chromogranin for evaluation of the distribution of chromogranin in normal and neoplastic pituitaries. In addition, a prospective study was done for assessment of the prevalence, morphology, and endocrine cell types of pituitary tumors in 100 autopsy subjects. When these 55 pituitary adenomas were examined with monoclonal antibody (LK2H10) directed against human chromogranin, selective staining of normal adenohypophyseal cell types and pituitary tumors was observed. Most null-cell adenomas (12/14) were positive for chromogranin, whereas all prolactin (PRL)-producing adenomas (19/19) were negative. Growth hormone (GH) adenomas were focally positive (9/9). All oncocytomas (2/2), 1 thyrotropin (TSH) adenoma, and a follicle-stimulating hormone/luteinizing hormone adenoma were positive for chromogranin. One or more adenomas were present in 14% of the autopsy cases. The tumors occurred most frequently in patients in the fifth through the seventh decades of life. Immunohistochemical staining of 13 adenomas revealed 1 TSH, 1 ACTH, and 4 PRL-producing tumors, whereas 7 other tumors, which were null-cell or undifferentiated adenomas, failed to stain for any of the seven principle pituitary hormones. These results indicate that antibody LK2H10 to human chromogranin is useful in the immunohistochemical characterization of pituitary adenomas. Incidental pituitary microadenomas from autopsy-derived pituitaries most commonly produce PRL, or they belong to the null-cell or undifferentiated tumor group.     

Okadaic acid inhibits the release of TSH in response to TRH and K+ from rat anterior pituitaries.

The effects of okadaic acid, a non-phorbol-12-tetradecanoate-13-acetate (non-TPA)-type tumor promoter and a potent inhibitor of protein phosphatases, on thyroid-stimulating hormone (TSH) secretion from the rat anterior pituitary were examined. Preincubation of anterior pituitaries with okadaic acid caused a time- and concentration-related decrease in a subsequent thyrotropin-releasing hormone (TRH)-stimulated TSH secretion, whereas it did not cause any changes in basal secretion of TSH. In addition, okadaic acid inhibited a subsequent high K(+)-induced TSH secretion. In contrast, ionomycin-induced TSH secretion was not inhibited by pretreatment with okadaic acid. The present results suggest that okadaic acid may block the release of TSH by inhibition of Ca2+ influx through voltage-sensitive and/or receptor-operated Ca2+ channels.     

Transdifferentiation of pituitary thyrotrophs to lactothyrotrophs in primary hypothyroidism: case report

Primary hypothyroidism causes adenohypophysial hyperplasia via stimulation by hypothalamic thyrotropin-releasing hormone (TRH). The effect was long thought to simply result in thyroid-stimulating hormone (TSH) and prolactin (PRL) cell hyperplasia, an increase in TSH and PRL blood levels with resultant pituitary enlargement, often mimicking adenoma. Recently, it was shown that transformation of growth hormone (GH) cells into TSH cells takes place in both clinical and experimental primary hypothyroidism. Such shifts from one cell to another with a concomitant change in hormone production are termed "transdifferentiation" and involve the gradual acquisition of morphologic features of thyrotrophs ("somatothyrotrophs"). We recently encountered a unique case of pituitary hyperplasia in a 40-year-old female with primary hypothyroidism wherein increased TSH production was by way of PRL cell recruitment. The resultant "lactothyrotrophs" maintained TSH cell morphology (cellular elongation and prominence of PAS-positive lysosomes) but expressed immunoreactivity for both hormones. No co-expression of GH was noted nor was thyroidectomy cells seen. This form of transdifferentiation has not previously been described.     

Sensitizing effect of treatment with estrogens on TSH response to TRH in male rats.

Daily administration of estradiol benzoate (10 microgram/100 g body wt) to intact male rats led to a twofold increase of the plasma TSH (thyroid-stimulating hormone) response to thyrotropin-releasing hormone (TRH) after 4 and 7 days of treatment whereas the basal plasma TSH level was not affected. The basal plasma PRL concentration and the PRL response to TRH were both markedly increased by estrogen treatment. The TSH pituitary content remained unchanged, whereas the PRL pituitary content increased in parallel with the effect on PRL secretion. Treatment with estrogens for 1 wk sensitized the TSH secretory response to low doses of TRH (10 ng), whereas no significant effect on the response was found at high doses of the neurohormone. The present data show that the stimulatory effect of estrogens on the TSH response to TRH is due to true sensitization of the thyrotrophs to the action of the neurohormone, whereas that on prolactin secretion can result partly from increased pituitary prolactin content.     

Suppression of maternal pituitary thyroid-stimulating hormone during pregnancy

Maternal serum thyroid-stimulating hormone (TSH) has been estimatedin 38 pregnant women with three new specific non-radioactivemonoclonal immunoassays which can measure very low concentrations.Values for the Pharmacia-LKB ‘DELFIA’ fluoroimmunoassayand for the Amersham ‘Amerlite’ luminescence immunoassaywere statistically identical over the range 0.15–1.5 uIU/mlWHO 2nd IRP 80/558, but thereafter the Amersham values wereslightly lower. The Abbott ‘IMx’ assay system gaveslightly higher results commensurate with the quoted highernon-pregnant normal values. The Amersham assay was preferredon practical grounds. There was no relationship of TSH levelsto HCG levels, or to the length of pregnancy. Four women hadapparently zero TSH levels by the Amersham and Pharmacia-LKBassays, although the Abbott assay could detect very low concentrations.Together with women whose TSH levels were below the normal non-pregnantrange for each assay, there were a total of eight women (21%)with TSH levels below ‘normal’. This suppressionof maternal pituitary TSH levels during pregnancy was consideredto be due to the central feedback inhibitory thyrotrophic activityof HCG, as a separate placental thyrotrophic hormone is believednot to exist. The enlargement of the maternal thyroid glandand the increased production of thyroid hormones during normalpregnancy is likewise to be attributed to HCG rather than toan increase in TSH production.     

Immunocytochemical detection of corticotropin-releasing factor: multiple cross-reactions of a widely used carboxy-terminally directed corticotropin-releasing factor antiserum (code rC70) in rat hypothalamus

Forty-one-residue corticotropin-releasing factor is a physiologically significant mediator of the hypothalamic control of corticotropin secretion by the anterior pituitary gland. This releasing hormone is produced by parvicellular neurons in the hypothalamic paraventricular nucleus that project to the external zone of the median eminence. Recent immunocytochemical evidence based on work with a rabbit antiserum against rat corticotropin-releasing factor (code rC70) suggests that about half of the parvicellular corticotropin-releasing factor-containing neurons in the hypothalamic paraventricular nucleus synthesize vasopressin, another potent corticotropin secretagogue, while the rest of the cells do not. If this is indeed the case, the neurohumoral control of corticotropin release may be mediated via distinct hypothalamic effector pathways utilizing releasing hormone cocktails of varying composition. In the present study we have examined the specificity of various antisera against rat corticotropin-releasing factor in immunocytochemical staining. Male Wistar rats pretreated with colchicine were used throughout. The brain was fixed by perfusion with a Zamboni type fixative solution. Vibratome sections of the hypothalamus were immunostained with three different primary antisera (codes rC70, rCRF-3, oCRF-N) using the peroxidase-antiperoxidase or avidin-biotin complex methods. All three antisera stained cell groups previously described to be immunopositive for corticotropin-releasing factor. Most notably, however, rC70 labelled a significant number of additional cells, most readily identified in the arcuate and suprachiasmatic nuclei, as well as in the dorsolateral hypothalamic area caudal to the paraventricular nucleus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Standardisation des immunodosages de la TSH : production de nouveaux calibrateurs et harmonisation des tests

Discordances among TSH measurements have been long documented over the past two decades but are still poorly understood at the molecular level. This review presents an overview of TSH structural polymorphism in the pituitary stock compared to various recombinant preparations and the circulating hormone. Comparative immunochemical analysis of pituitary TSH and a recombinant hormone produced in mammalian cells demonstrate that several epitopes are differentially expressed as a result of alteration in glycosylation. Consequently, variable antibody recognition may be generated between blood samples and internal calibrators among assays. There is thus a need for designing new calibrators which best mimick circulating TSH for developing harmonization of TSH tests.     

In vitro effects of endogenous opiate peptides on thyrotropin function: inhibition of thyrotropin-releasing hormone release and absence of effect on thyrotropin release

Thyrotropin-releasing hormone (TRH) or thyroid-stimulating hormone (TSH) was measured by radioimmunoassay in the incubation medium of rat hypothalami or anterior pituitary halves, respectively. We studied the effect of opioid peptide addition (10(-8) to 10(-6) M) on TRH or TSH release. alpha- or beta-Endorphin decreased TRH release in a dose-dependent manner while only 10(-6) M Leu- or Met-enkephalin decreased TRH release. These inhibitory effects were prevented by addition of naloxone (10(-5) M). In the dose range used none of the opioid peptides modified TSH release. These results indicate that opioid peptides may play a role in the regulation of thyrotropin secretion via a hypothalamic action on TRH release.     

GnRH 相关肽在啮齿类前脑及下丘脑分布的免疫组织化学研究

人和大鼠的促性腺激素释放激素原(ProGnRH),由 GnRH 和 GnRH 相关肽(GAP)组成。本文应用3种分别抗人和大鼠 GAP N-末端、中段和 C-末端的抗血清和 ABC 免疫酶法,对大鼠、小鼠和豚鼠脑 GAP 神经元进行了观察。3种动物的 GAP 神经元分布相似,GAP 神经元胞体主要存在于隔-视前区,以终板血管器附近的斜角带最为集中。视上核附近亦有较多的 GAP神经元。GAP 纤维广泛存在于前脑和下丘脑,终止于终板血管器和正中隆起。3种 GAP 抗血清中,以抗 N-末端者染出的免疫反应阳性成分多,染色深。GAP 神经元与相邻切片上所染的 GnRH神经元的胞体、纤维和终末的形态和分布相同。本文的观察和其他有关的研究提示,哺乳类具有相同的 ProGnRH,其加工产物 GAP(或其裂解片段)与 GnRH 共同分泌到垂体门脉,调节垂体前叶的激素分泌。     

Development and validation of a radioimmunoassay for beta-endorphin-related peptides

A radioimmunoassay method suitable for measuring levels of B-endorphin, B-lipotropin and proopiocortin in tissue and plasma extracts was developed and the method was evaluated by using 3 independently prepared antisera. Of the several variables tested the choice of assay buffer and test tubes and the purification of tracer were found to be the most critical in the successful performance of B-endorphin radioimmunoassay. The prevention of degradation of tracer during incubation was also necessary when crude tissue extracts or plasma were assay. The sensitivities of the assays obtained with the 3 antisera utilized (Bendo 2, K2 and RB 100) were 1, 2.8 and 4 fmol B-endorphin per tube. All the antisera crossreacted equimolarly with B-lipotropin. The method was employed to measure the levels of B-endorphin immunoreactivity in rat pituitary and plasma by separating the different immunoreactants by gel filtration. It was found that both pituitary and plasma contain significant amounts of proopiocortin, B-lipotropin and B-endorphin, the molar proportions being 10:33:57 in pituitary and 15:15:17 in plasma, respectively. Both anterior and posterior lobes of rat pituitary were found to contain all the three immunoreactants. However, anterior lobe contained mostly the larger molecules, while posterior lobe was rich in B-endorphin. No absolute levels of the immunoreactants could be obtained due to the heterologous system used. Moreover the elution pattern of the immunoreactivity was found to be dependent on the conditions used for elution in gel filtration: higher proportion of the immunoreactivity eluted like B-endorphin when the elution was done in dissociating conditions (6 M urea) compared to elution with ordinary buffers.     

Hypothalamic control of thyrotrophin secretion

1. By means of the haemagglutination-inhibition technique, it has been possible to measure the plasma and pituitary levels of thyroid-stimulating hormone (TSH) following unilateral and bilateral electrolytic lesions placed in the supraoptic area of the hypothalamus of female rats.

2. Unilateral lesions after 8 and 48 hr caused a temporary decrease in the percentage of circulating TSH followed by a return to normal.

3. Bilateral lesions caused a fall in plasma TSH to a level 41% below normal at 10 days whereas the pituitary level increased 70%.

4. Rats with bilateral symmetrical supraoptic lesions, kept for 3 days at 4° C, had a lower plasma TSH content (23%) and a higher pituitary TSH content (16%) than the sham operated control animals but the levels of pituitary TSH did not show any significant difference (0·100 > P > 0·050).

5. Thyroidectomized rats with bilateral lesions kept at 26° C for 3 days had a much lower plasma TSH (39%) and a slightly higher pituitary TSH as compared to the normal intact animals, suggesting that the hypothalamus also influences the synthesis of TSH in the pituitary.

6. Thyroidectomized rats with bilateral lesions kept at 4° C for 3 days showed both a plasma and pituitary TSH increase compared to controls at 26° C, suggesting that when a higher demand for thyroid function is present the pituitary gland has some autonomy for both secretion and release of TSH. This autonomy appears to be slight, as there was no statistically significant difference between the pituitary TSH levels of the thyroidectomized animals bearing similar supraoptic lesions and exposed to 4 and 26° C.

     

Physiological Relevance of Thyroid Stimulating Hormone and Thyroid Stimulating Hormone Receptor in Tissues other than the Thyroid

Decades of research have provided strong evidence for a reciprocal relationship between the immune system and hormones of the hypothalamus–pituitary–thyroid (HPT) axis. Thyroid stimulating hormone (TSH), in particular, has been shown to have a variety of immune-regulating cytokine-like activities that can influence the outcome of T cell development in the thymus and intestine, and can affect the magnitude of antibody and cell-mediated responses of peripheral lymphocytes. Production of TSH and the expression of the TSH receptor are widely but selectively distributed across many different types of hematopoietic cells in the bone marrow, as well as among subsets of dendritic cells, monocytes and lymphocytes in the spleen and lymph nodes. In addition to their role in immunity, the involvement of TSH-producing hematopoietic cells in the microregulation of thyroid hormone activity represents a novel and potentially important aspect of the TSH-mediated immune-endocrine circuit.     

Thyrotropin-releasing hormone receptors in gut tissues resemble pituitary receptors

The relative order of activity of thyrotropin-releasing hormone (TRH) and various analogs in contracting the isolated guinea pig antrum and duodenum correlated with their potencies in activating thyroid-stimulating hormone (TSH) release. The action of TRH in both tissues was selectively antagonized by the putative pituitary TRH receptor antagonist, chlordiazepoxide (10 microM). The data indicate that the contractions produced by TRH in these gut tissues are mediated by TRH receptors with similar characteristics as the pituitary TRH receptors responsible for TSH release.