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Effects of Metyrapone Administration on Thyrotropin Secretion in Healthy Subjects–A Clinical Research Center Study¹

Division of Endocrinology, Diabetes, and Clinical Nutrition, Oregon Health Sciences University, Portland, Oregon 97201

Address all correspondence and requests for reprints to: Dr. M. H. Samuels, Division of Endocrinology, Oregon Health Sciences University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97201. E-mail: samuelsm{at}ohsu.edu

	Abstract

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Although pharmacological doses of glucocorticoids suppress TSH secretion, less is known regarding the effects of physiological variations in cortisol levels on TSH. To study this issue, 12 healthy subjects each underwent 2 studies, in random order: 1) each subject received an infusion of saline for 48 h; and 2) each subject received an infusion of saline and oral administration of metyrapone (500 mg every 4 h) for 48 h. Cortisol and TSH levels were measured every 15 min during the final 24 h of each study, and resulting mean hormone levels during the 24-h periods were compared between the two studies. Metyrapone administration reduced serum cortisol levels by 39% between 0800 and 1345 h and by 47% between 0200 and 0745 h, with no significant changes during other time periods. Metyrapone increased daytime (0800–1945 h) mean TSH levels by 35%, with no change in nocturnal (2000–0745 h) TSH levels. This led to equalization of daytime and nocturnal TSH levels and abolition of the usual circadian variation in TSH. TSH pulse frequency was no different between the two studies, whereas daytime TSH pulse amplitude increased 33% during metyrapone administration. There were no changes in TSH responses to TRH, or in serum T₃ or free T₄ levels, at the end of the studies. These results suggest that the early morning increase in endogenous cortisol levels in healthy subjects causes the daytime decrease in TSH levels. In addition, these results show that very mild changes in cortisol levels within the physiological range are sufficient to affect TSH secretion.

	Introduction

Top Abstract Introduction Subjects and Methods Results Discussion References

 
SUPRAPHYSIOLOGICAL doses of glucocorticoids or elevated endogenous cortisol levels suppress serum TSH levels in humans (1, 2, 3, 4), but the effects of physiological cortisol levels on TSH secretion are less clear. To adequately investigate this phenomenon, frequent measurements of cortisol and TSH must be made, given the pulsatile and circadian nature of secretion of both hormones. Recent data obtained in this fashion from subjects with primary adrenal insufficiency suggest that cortisol levels within the physiological range control TSH secretion, and that minor changes in serum cortisol levels have significant effects on serum TSH levels (5, 6, 7). These studies compared TSH secretion in the complete absence of cortisol to TSH secretion during cortisol regimens that replicated normal serum cortisol levels. Based on these results, we were interested in the question of whether even more subtle variations in serum cortisol levels affect TSH secretion. To address this question, we designed a study to partially suppress endogenous cortisol levels using metyrapone in healthy subjects and performed frequent measurements of serum cortisol and TSH levels over 24 h. Thus, this study extends our earlier observations on the significant effects of varying cortisol levels within the normal range on dynamic TSH secretion.

	Subjects and Methods

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Subjects

Twelve normal subjects (10 women and 2 men), ages 18–40 yr, with no active medical problems, of normal weight, and taking no medications, were recruited for this study. Each subject underwent a screening visit, including a complete history, physical examination, and laboratory testing (complete blood count, chemistry panel, and pregnancy test in women) to exclude underlying medical or endocrine problems or medication use that would preclude entrance into the study. All subjects had normal serum TSH and free T₄ levels and normal thyroid gland examinations before the study. The study was approved by the Oregon Health Sciences University institutional review board, and each subject gave informed consent before starting the study.

Experimental design

Each subject underwent two studies as in-patients at the Oregon Health Sciences University General Clinical Research Center, in random order. The two studies were spaced at least 1 month apart.

Baseline study. The subjects were admitted to the General Clinical Research Center for 50 h, starting at 0800 h the first day and ending at 1000 h the third day, during which time they were free to move about and eat normal meals. During the first day, a venous catheter was placed in a forearm vein, and 5% glucose in 0.9% saline was infused at 100 cc/h for the rest of the study. Starting at 0800 h on the second day, 2 cc blood were withdrawn from the catheter every 15 min for 24 h. At 0800 h on the third day, 250 µg TRH were injected iv, and 2 cc blood were withdrawn 20, 30, 60, and 120 min later.

Metyrapone study. This study was identical to the baseline study, but the subjects were given metyrapone, an 11ß-hydroxylase inhibitor thatblocks cortisol synthesis, at 500 mg every 4 h, orally, during the 2-day study. This dose was chosen in an attempt to reduce serum cortisol levels without reaching undetectable levels and without causing significant side-effects that could lead to stress-induced changes in TSH secretion (8). The first dose of metyrapone was given at 0800 h on the first day, and the last dose was given at 0400 h on the third day, for a total of 12 doses. Two subjects had mild nausea during metyrapone administration that was controlled by increasing food intake at the time of drug administration. One subject developed mild peripheral edema during metyrapone administration that resolved within 1 day of drug discontinuation. No other side-effects were noted.

Laboratory methods

All samples were analyzed for cortisol and TSH levels by two-site chemiluminescent assays (Nichols Institute Diagnostics, San Juan Capistrano, CA). Coefficients of variation were 3–5% (intraassay) and 6–10% (interassay) at serum hormone levels measured in the subjects. Assay sensitivity was 22.1 nmol/L (0.8 µg/dL) for cortisol and 0.02 mU/L for TSH. Free T₄ levels were measured at the end of each study by equilibrium dialysis (Nichols Institute Diagnostics), and total T₃ levels were measured at the end of each study by in-house RIA. All samples from an individual were run in duplicate, and all samples from a single study were run in the same assay.

TSH pulses were located by Cluster analysis, using dose-dependent coefficients of variation calculated from sample replicates in each subject’s hormone series. Cluster parameters were two points for test nadirs and one point for test peaks. The t statistics were 2.0 for up-strokes and down-strokes. These parameters yield false positive and negative peak detection rates of less than 5%, determined by analysis of pooled serum samples and simulated hormone series (9).

Paired t tests were used to compare hormone levels during the following time periods: 1) daytime (0800–1945 h), nocturnal (2000–0745 h), and 24-h mean TSH levels and TSH pulse parameters; 2) mean serum cortisol levels over 24 h and divided into 6-h time blocks (0800–1345, 1400–1945, 2000–0145, and 0200–0745 h); and 3) mean serum free T₄ and total T₃ levels at the end of each study.

	Results

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Serum cortisol levels during the two studies

Figure 1 (top panel) and Table 1 show mean cortisol levels measured every 15 min over 24 h during the second day of each study. Twenty-four-hour mean serum cortisol levels decreased 26% during metyrapone administration, from 229 ± 28 nmol/L (8.29 ± 1.01 µg/dL) to 171 ± 27 nmol/L (6.20 ± 0.97 µg/dL), although this did not quite reach statistical significance (P = 0.08). When the mean cortisol levels were divided into four 6-h time blocks, it was apparent that metyrapone caused a significant decrease in cortisol levels from 0800 to 1345 h [a 39% decrease, from 337 ± 44 to 205 ± 37 nmol/L (12.20 ± 1.58 to 7.44 ± 1.34 µg/dL)] and from 0200 to 0745 h [a 47% decrease, from 289 ± 50 to 154 ± 20 nmol/L (10.48 ± 1.81 to 5.58 ± 0.72 µg/dL)]. The 24-h serum cortisol profile during metyrapone administration was unchanged compared to baseline during the time of relative quiescence of the adrenal axis (late evening and nighttime), but was blunted during the time of adrenal activation (early morning), leading to abolition of the usual cortisol circadian rhythm (Fig. 1). Thus, metyrapone administration reached our goal of lowering cortisol levels without leading to complete suppression of serum cortisol.

View larger version (28K): [in this window] [in a new window]   Figure 1. Top panel, Mean serum cortisol levels measured every 15 min in healthy subjects during the final 24 h of the baseline study (•) and the metyrapone study (). Bottom panel, Mean serum TSH levels measured every 15 min in the subjects during the final 24 h of the baseline study (•) and the metyrapone study ().

Figure 1 (bottom panel) and Table 1 show mean TSH levels measured every 15 min over 24 h during the second day of each study. Twenty-four-hour mean TSH levels were no different during the two studies. However, daytime TSH levels (0800–1945 h) increased 35% during metyrapone administration (from 1.09 ± 0.10 to 1.47 ± 0.21 mU/L), closely following the time of maximal effect of metyrapone on cortisol levels. In contrast, there was no change in TSH levels at night (2000–0745 h), whereas cortisol levels were low during both studies. Thus, the normal circadian variation in TSH levels was lost during metyrapone administration due to an increase in daytime levels, in conjunction with blunting of the normal daytime increase in cortisol secretion. There were no differences in cortisol or TSH responses to metyrapone administration in the male subjects compared to the female subjects, although the number of male subjects was too small to draw any definite conclusions regarding possible gender differences.

Table 1 also summarizes the results of TSH pulse analysis during the two studies. There was no change in 24-h TSH pulse frequency or amplitude between the two studies, whereas metyrapone increased daytime TSH pulse amplitude by 33%, similar in magnitude to the changes in mean TSH levels. Serum TSH responses to TRH were not different between the two studies. There were no differences in serum T₃ or free T₄ levels measured at the end of the two studies.

	Discussion

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Many studies of TSH secretion in humans have shown that glucocorticoids suppress TSH levels (1, 2, 3, 4), but most of these studies involved supraphysiological doses of glucocorticoids. More recent studies by our group and by Hangaard et al. in patients with Addison’s disease have extended these findings into the physiological range of cortisol levels (5, 6, 7). These studies used graduated doses of hydrocortisone given to mimic the normal diurnal rhythm of cortisol (and, in the case of our studies, the normal pulsatile nature of cortisol secretion) and compared the resulting TSH levels to those obtained in the absence of cortisol. The main findings of these studies are that complete cortisol withdrawal increases TSH levels, and replacement doses of cortisol decrease TSH levels. Based on these results, we were intrigued by the idea that even more subtle changes in cortisol levels within the normal range could affect TSH secretion. To this end, in the current study we investigated TSH levels during partial lowering of endogenous cortisol levels, using metyrapone administration. Our protocol was successful in decreasing cortisol levels by 40–50% during the times when cortisol levels are usually highest, but without complete suppression of serum cortisol. The lack of significant side-effects ensured that TSH and cortisol levels would not be affected by nonspecific stress effects in the subjects.

Our main finding from this study is that daytime TSH levels are increased by this maneuver, with abolition of the usual TSH circadian rhythm. The degree of daytime TSH increase (35%) is impressive, given the fact that mean early morning cortisol levels did not decrease below 140 nmol/L (5 µg/dL). This suggests that the early morning increase in endogenous cortisol levels in healthy subjects causes the normal daytime decrease in TSH levels. In contrast, metyrapone has no effect on TSH levels during the time when cortisol secretion is normally low. These results also show that very mild changes in cortisol levels, smaller than those measured in previous studies, are sufficient to affect TSH secretion. It is possible that this fine-tuning of TSH levels within the normal range by physiological variation in cortisol levels is one way that cortisol affects intermediary metabolism and stress responses throughout the circadian time period. However, the biological relevance of this regulation has yet to be determined.

A few previous studies measured TSH levels in healthy subjects after the administration of metyrapone (10, 11, 12). Two studies reported no effect on TSH levels, whereas the third reported a doubling of 0800 h TSH levels. In these studies the degree of cortisol suppression was variable, single doses of metyrapone were used, less sensitive TSH assays were used, and/or only single measurements of TSH were made. Therefore, the current study represents the first documentation of the effect of partial cortisol blockade on 24-h dynamic TSH secretion.

Metyrapone lowers serum cortisol levels by inhibiting the final enzymatic step in cortisol biosynthesis, the conversion of 11-deoxycortisol to cortisol. Thus, decreased serum cortisol levels are associated with increased serum 11-deoxycortisol levels during metyrapone administration. We did not measure 11-deoxycortisol levels in our subjects, but we assume that they were elevated, given our success in lowering serum cortisol levels. In addition, we cannot completely rule out an independent effect of 11-deoxycortisol levels on TSH secretion. However, previous studies in subjects with adrenal insufficiency support the conclusion that low cortisol levels, rather than increased 11-deoxycortisol levels, increased TSH secretion in the present study (5, 6, 7), because subjects with adrenal insufficiency do not have significant 11-deoxycortisol production.

In another recent study our group investigated the effects of minor elevations in serum cortisol levels on TSH secretion in healthy subjects (13). In this study subjects underwent short-term (56 h) caloric deprivation. Fasting led to a 32% increase in 24-h serum cortisol levels, with more pronounced increases during the night. The same subjects in the fed state then underwent low dose pulsatile infusions of hydrocortisone that replicated the fasting-induced changes in serum cortisol levels. Serum TSH levels fell by over 50% during either fasting or hydrocortisone infusions, with more pronounced suppression at night and abolition of the normal circadian TSH rhythm. These changes occurred with very minor alterations in serum cortisol levels [mean nocturnal cortisol levels 140–190 nmol/L (5–7 µg/dL) during either fasting or hydrocortisone infusions]. Taken together with the current study, these results show that TSH secretion is extremely sensitive to small increments in cortisol levels. During the time when cortisol levels are usually low (at night), small elevations can profoundly suppress TSH secretion; during the time when cortisol levels are usually higher (early morning and daytime), small decreases can elevate TSH levels. In each case, the observed alterations in cortisol levels are within the normal range and would not be noted with less frequent measurements.

TSH responses to TRH did not change after metyrapone administration in the current study. This suggests that physiological cortisol levels act via the hypothalamus, rather than the pituitary, to suppress TSH secretion. This is in agreement with our previous studies in patients with adrenal insufficiency (7) and contrasts with studies in humans that report blunted TSH responses to TRH after supraphysiological doses of glucocorticoids (14, 15). Animal and in vitro studies have shown that glucocorticoids have effects on hypothalamic TRH production (16, 17, 18, 19), whereas rat pituitary cell culture studies report conflicting results on whether glucocorticoids directly suppress TSH secretion (20, 21). In almost all of these studies doses of glucocorticoids were supraphysiological. T₃ and free T₄ levels were no different at the end of the two studies. This is not surprising, given the short time course of the studies. It is possible that a longer period of slightly higher daytime TSH levels could increase serum thyroid hormone levels, although the more likely result would be increased negative feedback and reestablishment of lower TSH levels.

In summary, we administered metyrapone to healthy subjects in a dose that led to lowering of early morning and daytime serum cortisol levels and measured resulting changes in serum TSH levels. We found that this intervention significantly increased daytime serum TSH levels, thereby abolishing the normal circadian rhythm of TSH. This suggests that the normal endogenous diurnal rhythm of cortisol controls the 24-h TSH rhythm, and that minor changes in serum cortisol have important effects on TSH secretion.

	Acknowledgments

 
We thank the Oregon Health Sciences University General Clinical Research Center staff for excellent patient care, sample collection, and assay performance.

	Footnotes

 
¹ This work was supported by NIH Grant R29-DK-48366 and the Oregon Health Sciences University General Clinical Research Center (NIH Grant M01-RR-00334).

Received April 8, 2000.

Revised May 11, 2000.

Accepted May 17, 2000.

	References

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