Thyrotropin releasing hormone: development of inactivation system during maturation of the rat.

Whereas thyrotropin releasing hormone is rapidly and extensively degraded by plasma of adult rats, no appreciable loss of biological or immunological activity is caused by plasma from rats 4 or 16 days old. The plasma of neonatal rats does not appear to contain an inhibitor of thyrotropin releasing hormone peptidase or a peptidase with altered substrate affinity. The development of an active peptidase in rat plasma suggests a physiological role for inactivation of thyrotropin releasing hormone.

Metabolic clearance and production rates of human thyrotropin.

Metabolic clearance (MCR) and production rates (PR) of human thyrotropin (hTSH) were determined by the constant infusion to equilibrium method 57 times in 55 patients. 16 control patients had a mean hTSH MCR of 50.7 ml/min. The mean hTSH MCR was significantly (P < 0.02) higher in 19 euthyroid men (51.6 ml/min) than in 12 euthyroid women (43.0 ml/min), but this apparent sex difference disappeared when the MCR were corrected for surface area, 25.8 (men) versus 25.2 ml/min per m(2) (women). Hypothyroid patients had significantly (P < 0.005) lower hTSH MCR (30.9 ml/min), and hyperthyroid patients had significantly (P < 0.05) higher hTSH MCR (60.9 ml/min) than controls. The hTSH MCR in patients with "decreased thyroid reserve" (40.9 ml/min), hyperfunctioning thyroid nodule (53.8 ml/min), and "empty sella syndrome" (46.6 ml/min) were not significantly different from controls. The mean hTSH PR in controls (104.3 mU/day) was significantly (P < 0.005) different from that in patients with "decreased thyroid reserve" (956 mU/day), hypothyroidism (4,440 mU/day), hyperthyroidism (< 43.9 mU/day) and a hyperfunctioning thyroid nodule (< 38.7 mU/day). In primary hypothyroidism intravenous triiodothyronine therapy (50 mug/day) for 10 days decreased the hTSH PR (from 4,244 to 2,461 mU/day) before changes in the hTSH MCR (from 33.1 to 33.7 mU/day) were observed. These studies have demonstrated that changes in the serum concentration of hTSH are mainly due to altered pituitary hTSH secretion with only a minor contribution from the change in hTSH MCR.

Lower levels of thyrotropin-releasing hormone-degrading activity in human cord and in maternal sera than in the serum of euthyroid, nonpregnant adults.

Thyrotropin-releasing hormone (TRH)-degrading activity was investigated in human cord, maternal, and euthyroid adult sera by measuring (a) the rate of disappearance of TRH and (b) the rate of formation of degradation products. The rate of TRH degradation in cord and maternal sera was 25-33% of that in euthyroid adult serum. Concomitantly, in cord and maternal sera, the rate of formation of proline, a major TRH degradation product in serum, was one-quarter to one-third that in euthyroid adult sera. The differences were highly significant (P less than 0.001). The decreased levels of TRH-degrading activity in cord and maternal sera cannot be explained by (a) the presence of a dialyzable inhibitor, (b) the absence of an activator of TRH degradation, or (c) a reversal of the degradation process. There was no difference in the types of radioactive degradation products formed by cord, maternal, and euthyroid adult sera. The low level of TRH-degrading activity and its possible relationship to high thyrotropin-stimulating hormone levels in cord serum suggest that TRH-degrading activity may be a factor to consider in investigations of the perinatal pituitary-thyroid axis, but further studies are needed to determine the role of serum degradation of TRH in regulating physiological levels of TRH.

Metabolic clearance and production rates of prolactin in man.

Metabolic clearance rates (MCR) and production rates (PR) of prolactin (PRL) have been determined by the constant infusion to equilibrium technique in 11 normal subjects, 6 patients with hyperthyroidism, 4 patients with hypothyroidism, and 9 patients with hyperprolactinemia. PRL MCR was also determined tin four patients during dopamine infusion. Mean PRL MCR was 46 +/- 1 ml/min per m2 in women and 44 +/- 3 ml/min per m2 in men, and was significantly correlated with body mass (r = 0.84, P less than 0.001). In contrast with controls, PRL MCR was higher in hyperthyroidism (MCR = 52 +/- 8 ml/min per m2, P less than 0.05), was slightly lower in hypothyroidism (MCR = 38 +/- 10 ml/min per m2, P = NS), and was significantly correlated with serum thyroxine (r = 0.46, P less than 0.02). PRL MCR was lower than controls in hyperprolactinemia (MCR = 40 +/- 5 ml/min per m2, P less than 0.01) and was inversely correlated with serum PRL (r = -0.72, P less than 0.001). PRL MCR was not significantly changed by dopamine infusion. Mean PRL PR for women and men was 211 +/- 74 and 187 +/- 44 micrograms/d per m2, respectively (P = NS). In hyperthyroidism the PRL PR was elevated (PR = 335 +/- 68 micrograms/d per m2, P less than 0.02), but in hypothyroidism the increase (PR = 233 +/- 159 micrograms/d per m2) was not significant. In hyperprolactinemia the PRL PR was extremely high (PR = 31,000 +/- 29,000 micrograms/d per m2). Dopamine infusion decreased RPL PR from 270 to 66 micrograms/d per m2 indicating that its effect was on pituitary PRL secretion and not PRL metabolism. To evaluate possible circulating PRL heterogeneity that might arise during infusion, gel filtration of infusate and serum obtained during the MCR procedure was performed. Labeled monomeric PRL (peak III, Kav (partition coefficient) = 0.4) was partially converted to two larger forms (peaks I and II) in vivo. Peak I (Kav = 0) was 30--40% immunoprecipitable, although peak II (Kav = 0.2) was not immunoprecipitable. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of peak I resulted in greater than or equal to 90% conversion to peak III and restoration of full immunoactivity. Thus, peak I is a noncovalently linked aggregate that is partially immunoactive, and therefore able to alter MCR determinations. These studies demonstrate the impact of hormone heterogeneity on MCR estimations and suggest that gel filtration of immunoprecipitable material be an integral part of future MCR measurements.

Partial target organ resistance to thyroid hormone.

An 8-year old boy with a small goiter, normal basal metabolic rate (BMR), and elevated serum thyroid hormone levels (thyroxine [T(4)] 19.5 mug per 100 ml, free T(4) 4 ng per 100 ml, triiodothyronine [T(3)] 505 ng per 100 ml) was studied. He had measurable serum thyroid-stimulating hormone (TSH) levels (average 5.5 muU per ml), and the thyroxine-binding proteins, hearing, and epiphyseal structures were normal. There was no parental consanguinity nor were there thyroid abnormalities either in the parents or six siblings.Methimazole, 50 mg daily, depressed thyroxine synthesis (T(4) 10.5, free T(4) 2.5) and caused a rise in TSH to 11 muU per ml. After discontinuation of treatment, TSH declined to 4.2 muU per ml and chemical hyperthyroidism returned (T(4) 21.0 mug per 100 ml, free T(4) 4.2, and total T(3) 475 ng per 100 ml, radioactive iodine [RAI] uptake 68%), but studies of BMR and insensible water loss showed the patient to be clinically euthyroid. Thyrotropin-releasing hormone (TRH), 200 mug i.v., caused a brisk rise in TSH to 28 muU per ml, with T(4) rising to 28 mug per 100 ml, free T(4) to 5.6, and T(3) to 730 ng per 100 ml, thus indicating that the pituitary-thyroid system was intact and that the patient's TSH was biologically active. The unusual sensitivity of the pituitary cells to TRH in spite of the markedly elevated serum thyroid hormone levels also suggested that the pituitary was insensitive to suppression by T(3) or T(4). Serum dilution studies gave immunochemical evidence that this patient's TSH was normal. Neither propranolol, 60 mg, chlorpromazine, 30 mg, nor prednisone, 15 mg daily, influenced thyroid indices. Steroid treatment, however, suppressed the pituitary response to TRH, T(3) in doses increased over a period of 12 days to as much as 150 mug daily caused a rise in serum T(3) to above 800 ng per 100 ml, a decline of T(4) to euthyroid levels (T(4) 9.5 mug per 100 ml, free T(4) 1.6 ng per 100 ml), suppression of the RAI uptake from 68% to 35%, and marked blunting of the responses to TRH, but the BMR and insensible water loss remained normal. The data suggest that the patient's disorder is due to partial resistance to thyroid hormone.

Suppression of pituitary TSH secretion in the patient with a hyperfunctioning thyroid nodule.

10 patients with a single hyperfunctioning thyroid nodule each were studied for pituitary thyrotropin (TSH) suppression. They were judged to be euthyroid on clinical grounds. The total thyroxine (T(4)D), free thyroxine (FT(4)), total triiodothyronine (T(3)D), and free triiodothyronine (FT(3)) were normal in most of the patients. Incorporation of (131)I into the hyperfunctioning thyroid nodules was not suppressed by the administration of physiological doses of T(3). Basal serum TSH concentrations were undetectable (<0.5 - 1.0 muU/ml) in all patients. The metabolic clearance of TSH in one patient before and after excision of the thyroid nodule was unchanged (40 vs. 42 ml/min) whereas the calculated production rate was undetectable before the operation (<29 mU/day) and normal after (103 mU/day). These data, in one patient, suggest that the undetectable concentration of TSH in these patients is a result of suppressed TSH secretion rather than accelerated TSH clearance. In eight patients, basal serum TSH concentrations failed to increase after the intravenous administration of 200 mug of thyrotropin-releasing hormone (TRH); minimal increases in serum TSH concentrations were observed in two patients. The suppression of TSH was evident despite "normal" concentrations of circulating thyroid hormones. The observation that normal serum concentrations of T(4)D, FT(4), T(3)D, and FT(3) may be associated with undetectable basal serum TSH concentrations and suppressed TSH response to TRH was also found in four hypothyroid patients given increasing doses of L-thyroxine and sequential TRH stimulation tests.

Metabolic clearance and secretion rates of subunits of human thyrotropin.

Metabolic clearance rates (MCR) of the alpha and beta subunits of human thyrotropin (hTSH-alpha and hTSH-beta) were determined by a constant infusion to equilibrium method. In 15 normal individuals (six men, six premenopausal women, and three post-menopausal women), the mean MCR of hTSH-alpha (68 ml/min per m2) was significantly faster than that of hTSH-beta (48 ml/min per m2) was significantly faster than that of hTSH-beta (48 ml/min per m2); both were two to three times more rapid than the previously determined MCR of hTSH. In patients with primary hypothyroidism, MCR were significantly slower with a mean value of 55 ml/min per m2 for hTSH-alpha and 37 ml/min per m2 for hTSH-beta. However, MCR of subunits were not significantly faster than normal in hyperthyroid patients. Serum concentrations of alpha subunits and hTSH-beta were measured by radioimmunoassay, and secretion rates of alpha and hTSH-beta from the pituitary were calculated using hTSH-alpha and hTSH-beta MCR, respectively. In the normal individuals, alpha secretion rates averaged 91 mug/day per m2, greater than those previously determined for hTSH and human follicle-stimulating hormone. Alpha secretion rates were significantly elevated in the normal postmenopausal women (211 mug/day per m2) and in the premenopausal hypothyroid women (202 mug/day per m2); they were also elevated in the postmenopausal hypothyroid women (277 mug/day per m2). Alpha secretion rates were significantly decreased in the premenopausal hyperthyroid women (66 mug/day per m2). Usually, the secretion rates of hTSH-beta could not be calculated in normal individuals, and the rates in hyperthyroid patients could never be calculated because serum hTSH-beta was not detected. Six normals had detectable hTSH-beta secretion rates (17 mug/day per m2); hTSH-beta secretion rates were significantly increased in patients with primary hypothyroidism (28 mug/day per m2). Although we had previously demonstrated a 50-fold increase in hTSH secretion rates in primary hypothyroidism, there was only a 2-fold increase in alpha and hTSH-beta secretion rates. Thus, increased subunit synthesis appears to be utilized predominantly for production of complete hTSH.

Antineutrophil autoantibodies in Graves' disease. Implications of thyrotropin binding to neutrophils.

The hyperthyroidism of Graves' disease may be caused by autoantibodies to thyrotropin (TSH) receptors. We have found that patients with this disease have autoantibodies to neutrophils as well, which can be displaced by TSH. Using a radiochemical opsonic assay, we found serum antibodies against homologous neutrophils in 6 of 11 Graves' patients. With a staphylococcal protein A-binding assay, we detected circulating antibodies to homologous neutrophils in 10 of 20 patients, while finding cell-bound antibody on autologous neutrophils in 7 of 8 (including 2 with negative serum tests). Use of human 125I-TSH in a radioligand binding assay revealed that TSH bound to neutrophils rapidly (maximum binding within 10 min at 22 degrees C, pH 7.4), specifically (less than 20% nonspecific binding), and reversibly. Adding TSH to the radiochemical assay resulted in a dose-dependent inhibition of opsonic antibody activity in serum from patients with Graves' disease. In contrast, TSH did not inhibit antibody activity of serum from patients with immune neutropenia not associated with thyroid disease. Our findings suggest a basis for the association of Graves' disease with neutropenia. Furthermore, the discovery of such antineutrophil antibodies in Graves' disease permits detection of cell-bound antibody when free antibody is not present.

Cancer mortality in women with thyroid disease.

A retrospective follow-up study of 7338 women with either nontoxic nodular goiter, thyroid adenoma, hyperthyroidism, hypothyroidism, Hashimoto's thyroiditis, or no thyroid disease was conducted. All women patients at the Massachusetts General Hospital Thyroid Clinic who were seen between 1925 and 1974 and who were treated for a minimum of 1 year were traced. A total of 2231 women (30.4%) were dead and 2012 women (27.4%) were alive as of December 31, 1978. Partial follow-up information was available for the remaining 3095 women (42.2%). The average length of follow-up was 15.2 years. When losses to follow-up were withdrawn at the time of their loss, the standardized mortality ratios (SMR) for all causes of death were 1.2 [95% confidence interval (CI), 1.1-1.3] for women with nontoxic nodular goiter, 1.2 (95% CI 1.0-1.3) for those with thyroid adenoma, 1.4 (95% CI 1.3-1.5) for women with hyperthyroidism, 1.5 (95% CI 1.3-1.7) for hypothyroid women, 1.2 (95% CI 0.9-1.5) for those with Hashimoto's thyroiditis, and 1.5 (95% CI 1.4-1.6) for those without thyroid disease. For deaths from all cancers, the standardized mortality ratios were 1.5 (95% CI 1.2-1.8) for women with nontoxic nodular goiter, 1.5 (95% CI 1.1-1.9) for those with thyroid adenoma, 1.2 (95% CI 1.0-1.4) for women with hyperthyroidism, 1.0 (95% CI 0.7-1.4) for the hypothyroid women, 1.2 (95% CI 0.7-2.1) for those with Hashimoto's thyroiditis, and 1.3 (95% CI 1.0-1.5) for those women without thyroid disease. When specific cancer sites were studied, excess numbers of deaths were observed from breast cancer in women with nontoxic nodular goiter (SMR = 1.6, 95% CI 1.0-2.6) and from lymphatic and hematopoietic cancer in women with nontoxic nodular goiter (SMR = 2.4, 95% CI 1.2-4.3) and thyroid adenoma (SMR = 2.7, 95% CI 1.1-5.2). An increase in thyroid cancer risk was observed in women with thyroid adenoma (SMR = 11.7, 95% CI 1.3-42.1) but was based on only two deaths. In hyperthyroid women, statistically significant increases in the number of deaths were observed from pancreatic cancer (SMR = 2.6, 95% CI 1.4-4.3) and respiratory cancer (SMR = 2.2, 95% CI 1.3-3.5), but not breast cancer (SMR = 1.3, 95% CI 0.8-1.8). When the data were stratified by the time between the onset of thyroid symptoms and death, a nonsignificant excess number of cancer deaths was observed in hyperthyroid women who died 20 or more years after their symptoms began.(ABSTRACT TRUNCATED AT 400 WORDS)

Thyroid microsomal membrane proteins. Effects of solubilization on molecular size.

The molecular size of microsomal membrane proteins from frozen porcine thyroids before and after solubilization by proteolytic and non-proteolytic techniques has been investigated by means of polyacrylamide-gel electrophoresis in the presence of 1% sodium dodecylsulfate. When thyroid microsomal membrane proteins are solubilized by non-proteolytic methods such as high pH, n-butanol, or deoxycholate, no major change in the electrophoretic pattern compared to untreated microsomes has been observed, thereby suggesting that these non-proteolytic methods are capable of extracting membrane proteins from thyroid microsomes without altering their molecular size. However, treatment of microsomes with protein-solubilizing levels of trypsin (1-5 mug trypsin per mg thyroid protein) results in degradation of all major proteins with a molecular weight greater than 30 000. The high-molecular-weight proteins are particularly susceptible to attack by trypsin. Thus, these experiments indicate that the use of trypsin to solubilize thyroid microsomal membrane proteins, particularly thyroid peroxidase, will result in fragmented proteins and should be avoided if intact membrane proteins are desired.

Triiodothyronine-induced thyrotoxicosis in ophthalmic Graves disease.

A euthyroid woman with ophthalmic Graves disease developed endogenous hyperthyroidism coincident with T3 suppression test. There is a putative role of liothyronine administration in precipitating or activating hyperthyroidism. Aberrancies in T3 suppression testing in graves disease occur.

Myocardial function in hypothyroidism. Relation to disease severity and response to treatment.

Systolic time intervals (ST) were used to evaluate myocardial function prospectively in 29 hypothyroid patients. The patients were divided into three categories of disease severity: (1) severe hypothyroidism, (2) mild hypothyroidism, and (3) decreased thyroid reserve or "prehypothyroidism." Groups 1 and 2 showed decreased myocardial contractility with a prolonged preejection period (PEP), shortened left ventricular ejection time (LVET), and increase PEP/LVET, compared with normal controls. The STI were more abnormal (P less than .05) in group 1 than in group 2, suggesting that the severity of myocardial dysfunction correlates with the severity of the hypothyroidism. Group 3 had normal STI. Ten patients were restudied when euthyroid and showed complete normalization of their STI, supporting the thesis that hypothyroidism was the sole cause of the initial myocardial dysfunction.

The effect of amoxapine and imipramine on serum prolactin levels.

The effect of traditional tricyclic antidepressants on serum prolactin levels is controversial. In a five-week double-blind study of depressed outpatients, imipramine hydrochloride therapy did not lead to any significant change in serum prolactin levels. In contrast, amoxapine, a new antidepressant, produced significant elevations in serum prolactin levels in female and in male patients. Amoxapine may block dopamine receptors in central tuberoinfundibular pathways, which would account for its prolactin-elevating activity. On the other hand, imipramine and other traditional tricyclic antidepressants do not affect dopamine transmission, do not raise serum prolactin levels, and are not effective antipsychotic drugs.

Plasma TSH levels, by radioimmunoassay, during the estrous cycle of the rat.

Between 1000 and 1200 hr on any given morning, female rats displaying regular 4-day estrous cycles were bled by decapitation. Plasma TSH concentrations were determined by homologous radioimmunoassay (RIA), in terms of NIAMDD-Rat-TSH-I-1 standards. Mean plasma TSH concentrations plus or minus SE were 1.2 plus or minus 0.1, 1.5 plus or minus 0.2, 2.3 plus or minus 0.6, and 1.1 plus or minus 0.1 ng/ml for rats killed at proestrus, estrus, diestrus-1, and diestrus-2, respectively. Statistical analysis revealed no significant differences among the TSH levels, except for the elevation at diestrus-1, which is of borderline significance at the 0.05 probability level. Our findings, based on RIA, are in contrast to previous reports, based on bioassays, of an elevation of plasma TSH at estrus.

Pituitary-thyroid axis in neonatal and adult rats: comparison of the sexes.

Systematic comparisons have been made of the development of the pituitary-thyroid axes of male and female rats, by measuring plasma thyrotropin (TSH) and thyroxine (T4) concentrations in neonates and adults. Observations were made in untreated groups as well as in rats treated with various regimens of exogenous T4, thyrotropin-releasing hormone, or TSH. All hormone determinations were by radioimmunoassay (RIA). Salient findings include the following: 1) In early neonatal life, untreated rats showed no significant sex difference in the plasma concentrations of either TSH or T4. 2) In adulthood, the plasma TSH of untreated males attained levels strikingly higher than those of neonates-the differences averaged 5-fold more. For females, the increase in plasma TSH during development was less marked, averaging slightly less than 2-fold more. Thus, untreated adults exhibited a clear sex difference in circulating TSH concentrations; the male TSH levels averaged 2.8-fold higher than those of females. 3) Plasma T4 concentrations also increased markedly during development. For both sexes, adult T4 levels were approximately 3-fold greater than the T4 levels in early neonatal life. Among untreated adults, the female T4 concentrations averaged 28% greater than those of males. 4) Plasma TSH and T4 concentrations exhibited only minor fluctuations, of borderline statistical significance, during the female estrous cycle. 5) A significant reduction in responsiveness to exogenous TRH was observed in adult male rats which had been treated with high doses of T4 in neonatal life, although the effect was not completely consistent. No significant reduction was observed in females which received the same treatment. We have concluded that major changes occur in the circulating hormone levels of the pituitary-thyroid axis of the rat between birth and adulthood, and that such changes are not identical for the two sexes.

Decreased thyroid function and high plasma prolactin levels in rats of the Buffalo strain.

Rats of the inbred Buffalo strain have previously been reported to be susceptible to thyroiditis, as defined by histology. We have studied the endocrinology of the pituitary-thyroid axis of this strain by making direct measurements of the plasma concentrations of TSH and T4 in untreated, adult Buffalo rats of both sexes. Plasma PRL levels were also measured. All hormone determinations were by RIA. In addition, relative thyroid weights were noted and, in many cases, preliminary assessment of thyroid histology was made. Our principle findings were as follows. 1) Decreased thyroid function, in addition to the previously reported histological abnormalities, was found to occur spontaneously among the rats studied. Indications of decreased function included elevations of plasma TSH and thyroid weight and depressions of plasma T4. We estimated the incidence of unequivocal thyroid disease as approximately 3% in each sex. 2) Basal plasma PRL concentrations of Buffalo rats averaged three to four times higher than those of outbred CD rats. Our findings strongly suggest that rats of the Buffalo strain will provide a good model for the study of thyroid failure of varying degrees and concomitant changes in the circulating levels of pituitary hormones.

Thiourea and cyanamide as inhibitors of thyroid peroxidase: the role of iodide.

Thiourea, methylmercaptoimidazole, propylthiouracil, and thiouracil are all potent inhibitors of thyroid peroxidase (TPO)-catalyzed iodination. Unlike the cyclic thioureylenes, thiourea at 5 mM has no effect on guaiacol oxidation. If iodide is added to guaiacol assays containing thiourea, enzyme activity is lost. The latter observation may be explained as follows. In the presence of iodide, the iodinating species [TPO.Ioxid], oxidizes thiourea to formamidine disulfide. This product decomposes to cyanamide at neutral pH. We have shown cyanamide to be an inhibitor of the peroxidative and iodinating functions of TPO. Studies in rats demonstrate that doses of thiourea which completely inhibit in vivo protein-bound iodine formation have no irreversible effect on TPO, as measured by guaiacol peroxidation after removal of the thyroids. The major in vivo action of cyanamide is similar to that of thiourea. The data suggest that the primary in vivo and in vitro mode of action of thiourea is the reversible Ioxid-trapping mechanism. The anomalous inhibition of guaiacol peroxidation seen in the presence of thiourea plus iodide derives from the formation of formamide disulfide, followed by its nonenzymic decomposition to cyanamide.

The developmental pattern of thyrotropin-releasing hormone-degrading activity in the plasma of rats.

The developmental pattern of TRH-degrading activity in rat plasma was determined by measuring the ability of plasma from rats of various ages to degrade TRH into degradation products. From a rate of 0.519 pmol TRH degraded/microliter-1.h-1 on day 8, plasma TRH-degrading activity increased to 18.1 pmol TRH by day 90 in female rats. The increase in rate of formation of proline, a major plasma degradation product, was in very good agreement with the increase in rate of TRH degradation. The major increase in TRH-degrading activity occurred between the third and seventh week of life. A similar pattern of development was observed in male rats; the only significant difference was that plasma from day 90 male rats was approximately 30% more active than that from female rats (P less than 0.002). Daily T3 administration to rats from days 8--26 resulted in a 2-fold increase in plasma TRH-degrading activity (P less than 0.05). The contribution of plasma degradation to the physiological control of TRH activity is not clear, but the magnitude of the increase in plasma TRH-degrading activity (approximately 30-fold) during the maturation of the rat is suggestive of a mechanism of biological significance.

Methimazole pharmacology in the rat: studies using a newly developed radioimmunoassay for methimazole.

Methimazole [1-methyl-2-mercaptoimidazole (MMI)] was given to normal male rats in their drinking water in concentrations ranging from 0.0001-0.05% for either 1 week or 1 month. Serum MMI levels in the rats ranged from 0.008-19.6 micrograms/ml, and were similar after 1 week and 1 month of treatment. Serum MMI was linearly related to the MMI concentration in the drinking water (r = 0.98, P less than 0.001). In contrast, intrathyroid MMI content plateaued with increasing MMI concentrations in the water, and was linearly related to the logarithm of the MMI concentration. At the highest MMI concentration (0.05%), thyroid MMI contents were similar in the 1-week and 1-month groups (approximately 1 X 10(-4) M). Surprisingly, at lower MMI concentrations, thyroid MMI content was significantly higher in the 1-week group than the 1-month group. Thyroid function was inhibited by MMI with similar depression of serum T4 or T3 after 1 week or 1 month of MMI treatment. Although the MMI concentration for 50% suppression of thyroid PBI was 0.003% in both groups, thyroid MMI content at this MMI concentration was 97 microM after 1 week but only 15 microM after 1 month. The continued thyroid-inhibiting activity of MMI at 1 month, despite a striking decrease in thyroid MMI content, may relate to intrathyroid iodide depletion, which was more severe after 1 month (thyroid 127I = 40 microM) than after 1 week (thyroid 127I = 140 microM) or in controls (470 microM). Rats were given 0.05% MMI for either 1 week or 1 month, and the drug was then withdrawn. In the 1-week group, serum MMI disappeared biexponentially, with a rapidly declining phase (t1/2 = 3.2 h) and a second, slower disappearance phase (t1/2 = 47.7 h). Similar findings were noted after 1 month of treatment. The disappearance of thyroid MMI was also biexponential after 1 week, but this variable could not be evaluated after 1 month because thyroid MMI fell rapidly to undetectable levels. There was a highly significant correlation in the 1-week group between the disappearance of MMI from the thyroid and the recovery of thyroid function as assessed by thyroid PBI (r = 0.81, P less than 0.01). Despite the very rapid disappearance of MMI from the thyroid after 1 month of treatment, the recovery time of thyroid PBI was significantly longer than after 1 week of treatment (2.1 days vs. 1.4 days for 50% recovery, P less than 0.01).(ABSTRACT TRUNCATED AT 400 WORDS)

Production and release of thyrotropin and its subunits by monolayer cultures containing bovine anterior pituitary cells.

We have developed a dispersed cell monolayer system derived from bovine anterior pituitary glands. Fresh 1- to 6-week-old calf anterior pituitaries were mechanically and enzymatically dispersed and incubated with Dulbecco's Modified Minimal Essential Medium containing 10% hypothyroid goat serum. The media and cell extracts from confluent monolayers were analyzed for bovine TSH and free alpha, and TSH beta subunits by specific homologous RIAs. Basal levels of TSH, free alpha, and free TSH beta subunits in the media were 6.2 +/- 0.3, and 0.95 +/- 0.05 ng/10(6) cells . 24 h, respectively. Hence, an 8- to 10-fold excess of free alpha over free TSH beta subunits was released into the medium. Intracellular basal levels of TSH, free alpha, and free TSH beta subunits were 27.6 +/- 1.7, 10.7 +/- 0.2, and 2.6 +/- 0.3 ng/10(6) cells . 24 h, respectively, and indicated a 3- to 4-fold excess of free alpha over free TSH beta subunits within the cells. The total alpha-subunit to total beta-subunit ratio was 2:1. TRH stimulated release of TSH and its subunits in a dose-dependent fashion, with a half-maximal dose of 2 nM and a maximal response dose of 10 nM. Stimulation with 100 nM TRH increased the levels of TSH, free alpha, and free TSH beta subunits (450-900%, 180-200%, and 300-400%, respectively) in medium, with concomitant decreases within cells. Treatment with thyroid hormones decreased basal and blunted TRH-stimulated levels of TSH and its subunits in medium but had no effect on intracellular stores. However, large doses of T4 (25 nM) or T3 (1 nM) did not completely abolish the TRH (100 nM)-stimulated hormone response. TRH and thyroid hormones affect the release of TSH and TSH beta to a greater extent than they do the alpha-subunit. Finally, total alpha and TSH beta subunit production was increased with TRH stimulation and decreased with thyroid hormone exposure. Thus, an in vitro system to study the net production and secretion of TSH and its subunits in the normal pituitary thyrotrope has been established. (Endocrinology 108: 387, 1981)