Klinische

Klin. Wschr. 56, 31-35 (1978)

W°ch%nhrift {) Springer-Verlag 1978

Thyroid-Stimulating Hormone (TSH), Triiodothyronine (T3) and Thyroxine (T4) Response to Intravenous and Oral Stimulation with Synthetic Thyrotropin-Releasing Hormone (TRH) in Young Healthy Adults P. Vogt, J. Girard and J.J. Staub Department of Endocrinology,UniversityChildrens"Hospital and Nuclear Medicine,UniversityMedicalClinicBasel, Switzerland

Plasma-TSH (Thyreoidea-stimulierendes Hormon), T 4 (Thyroxin) und T3 (Trijodothyronin)-Reaktion nach oraler und intraven6ser Stimulation mit TRH (thyrotropin-releasing-hormon)

Sehliissdwiirter: TSH - Thyroxin - Trijodothyronin - Hypothalamus - Hypophysen-Schilddrfisenachse - TRH.

Summary. TSH, Zusammenfassung. Die Plasma TSH, T3 und T4 Reaktion auf Stimulation mit dem synthetischen thyrotropin-releasing-hormone (TRH) wurde bei 24 jungen gesunden Erwachsenen nach intraven6ser Stimulation mit 200 lag TRH und bei 25 gesunden jungen Erwachsenen nach oraler Stimulation mit 1 - 3 maliger Applikation yon 40 mg TRH untersucht. Nach intraven6ser Stimulation steigt die TSH Konzentration innert 30 min auf einen Maximalwert von im Mittel 11,7 gE/ ml an. Der T3 Anstieg yon 1,17auf 1,62 ng/ml nach 2 h ist statistisch signifikant aber im Einzeltest diagnostisch nicht verwertbar. Ein signifikanter Geschlechtsunterschied konnte in der vorliegenden Untersuchung weder ffir die TSH noch ftir die T3 Reaktion gefunden werden. Nach oraler Stimulation wird ein etwas h6herer maximaler TSH Wert von 13;2 laE/ml 3 h nach der Applikation gefunden. Der T3 Anstieg auf 2,19 ng/ml ist statistisch signifikant und diagnostisch verwertbar. Die Zunahme der Thyroxinkonzentration nach 8 h oder 3 h nach der dritten TRH Dosis ist statistisch signifikant. Die nach intraven6ser Stimulation gefundenen Werte stimmen gut mit den bisherigen Ergebnissen in der Literatur tiberein. Wfihrend die Untersuchungen an gr6Beren gesunden Kollektiven ffir die intraven6se Stimulation eher sp~rlich sind, fehlen sie ffir die orale Stimulation praktisch ganz. Dreimalige repetierte Stimulation mitje 40 mg TRH fiber 24 h ergeben nach der dritten Dosis eine geringere TSH Stimulation auf nur 5,8 laE/ml bei einer vergleichbaren Zunahme der T3 Konzentration. Offprint requests to: Prof. Dr. J. Girard (address see above)

T 3 and T 4 response to stimulation with thyrotropin releasing hormone (TRH) has been investigated in 24 young healthy adults after intravenous injection and in 25 young healthy adults upon oral application of 40 mg of TRH. After intravenous injection the TSH concentration raises from a mean ofl.6 to a mean maximum of ll.7laU/ml. A statistically significant sex difference could not be found. T3 shows a statistically significant increase which is however too small to be of diagnostic value in an individual test. After oral stimulation with 40 nag of TRH, TSH rises to a slightly higher maximum of 13.2laU/ml after 3 h. The T3 increase from 1.5 to 2.19 ng/ml is significant and considerably higher than after intravenous stimulation. The thyroxin increase is statistically significant. The present results compare well with previously published data for intravenous stimulation. The oral route of TRH application has not yet been widely used and the present series establishes the normal response in young healthy adults. Repetitive stimulation with three times 40 mg of TRH leads to a decrease in TSH stimulation which reaches 5.8 laU/ml 3 h after the third dose. This is in contrast to a comparable increase in plasma Ta.

Key words: TRH - TSH - Thyroxin - Triiodothyronine - Hypothalamo-pituitary-thyroid-axis.

TSH, T3 and T 4 response to intravenous and oral stimulation with TRH had added very much to the understanding of thyroid physiology and pathophys-

32

P, Vogt et al.: Oral and Intravenous TRH-Test

iology. TSH and prolactin response to the releasing hormone has been widely applied in diagnostic endocrinology of the hypothalamo-pituitary and thyroid systems. The TSH response to TRH has proved to be the most sensitive test of thyroid function. Reports on the intravenous application of TRH in various pathological states are numerous [10, 16, 19, 33, 1, 4, 5, 7, 29, 2, 14, 6, 8, 17, 24]. Investigations on normal responses in larger series are comparatively rare [3, t8, 31]. The oral route of stimulation has not yet been widely applied. The TSH, T3 and T4 response to either intravenous or oral stimulation with TRH was evaluated in the present investigation in two groups of euthyroid healthy young adults.

2. Oral Test. After an overnight fast a blood sample was taken at 8.00 a.m. 1 tablet of T R H = 4 0 mg was swallowed with water. Further blood samples were taken 3 and 8 h later, A second dose of 40 mg TRH was taken at 9 p.m. (13 h after starting the investigation) and the third dose was taken the following morning 23 h after starting the investigation. A last blood sample was taken 3 h after the third dose of TRH (=26 h after starting the experiment). All plasma samples were collected on EDTA, centrifuged and plasma frozen in aliquots until assayed. TSH was assayed with a radioimmunoassay using human TSH (NIH-TSH-HS-2 or Kabi) for iodination with chloramine-T. The labelled preparation was purified by adsorption to 5 mg Quso (Philadelphia Quartz Co,) and the eluted protein fraction chromatographed on Sephadex G-75, H-TSH MRC 68/38 was used as a reference preparation. Antiporcine-TSH (gift from Dr, G. Hennen, Liege) was used at a final dilution of t :960,000. Phase separation was performed by immuno-precipitation with goat antirabbit-antiserum (antibodies inc:). Plasmas were assayed in volumes of 200, 100, 50 and 25/al respectively. Intraassay coefficient of variation was 11% interassay coefficient of variation 13% at a TSH concentration of 5 gU/ml. Thyroxin was assayed using a commercial kit (Pantex). T4 intraassay variation was 5%, interassay variation 11%. Triiodothyronine was assayed using a 12S!_Ta T3 (Farbwerke Hoechst AG) and an anti T3 antiserum and T3 standard preparation from Henning (Berlin), ANS was used for blocking TBG and charcoal dextran for separation.

Materials and Methods 24 young healthy adults (medical students and hospital staff) were investigated after the intravenous application of TRH, 4 out of the 9 female individuals took oral contraceptives containing estrogens, 25 male individuals 18 to 20 years of age were investigated after oral administration of TRH.

Results Test Procedure

1. Intravenous Test 1. Intravenous Test. After an overnight fast an indwelling needle was placed in a cubital vein and a blood sample taken before injection of 200 ~tg of TRH (injected over 30 s). Blood samples were taken 20, 30, 60 and t20 rain later.

Basal TSH concentrations are at or below the sensitivity of the method (1.6 gU/ml). The mean maximum

Table 1

Ta~e3

Minutes gU/ml TSH

0 20 30 60 120

Hours #E/ml TSH

T3 ng/ml

x

SD

SEM

1.6 11.13 11.72 8.2t 4.49

5.15 5.87 4.t2 3.40

1.05 t.22 0.85 0.72

x

SD

SEM

t.17

0.25

0.05

1.62

0.36

x 0 3 8 26

SD

2.31 1.95 13.16 6.12 3.50 3.10 5.82 2.85

ng/ml T3

gg/100 ml T 4

SEM

x

SD

SEM

x

SD

0.39 1.22 0.62 0.58

1.52 2.19 1.91 2.15

0.35 0.62 0.37 0.47

0.07 7.53 1.tl 0.I2 0.07 9.15 1.44 0 . 0 9 10.15 1.60

SEM 0.22 0.28 0.32

0.07

Table 2

Minutes

0 20 30 60 120

Female. group (gU/ml TSH)

Male group (~tU/ml TSH)

Female group (ng/ml T3)

Male group (ng/ml T3)

)2

SD

SEM

2

2

SD

SEM

.~

SD

SEM

1.6 13.44 12.83 10.25 5.25

1.13

0.36

0.12

1.29

0.15

0.04

5.45 4.43 5.15 5.01

1.81 1.56 1.71 1,77

1.69

0.53

0.18

1.55

0.23

0,06

1.6 9.75 11.12 7.09 3.89

SD

SEM

4,59 6.55 2.77 2.16

1.18 1.69 0,71 0.55

P. Vogt et al. : Oral and Intravenous TRH-Test

33

Table 4. Intravenoustest with TRH-Response of TSH (gU/ml) and Ta (ng/ml) Dose (gg TRH)

Basal

20'

30'

This study Fagtia [10]

200 100

11.1 12.2 5.1

Torjesen [32]

500

Malvaux [23] Shenkman [29] Job [20] Fleischer [12]

200 400 200 500

1.6 1.6 1.8 2.3 2.1 7,1 1,3 5.4 2.1

PateI [25]

200

Lawton [21]

200

36.5 9.1 18.9

60'

I20'

TSH

Basal

120'

Ta

11.7 11.6 4.7 6.2 4.2

8.2 6.7 3.2 4.6 2,8 26.2

4.5 2.9 2.5

1.17

1.62

0.45

1.12

1.64

0.52

17.9 15.0

12.5

I0.1 10.6 3.3 3.9 2.1 29.4 7.8 13.5 12.9

1.28

1.99

0.71 0,59

of 11.7 ( +_5.87) gU/ml is reached 20-30 min after injection. At 2 h the mean TSH concentration is still slightly elevated (Table 1). The 9 female individuals show a somewhat higher increase of TSH compared to the male group. The sex difference is however statistically insignificant (Table 2). Comparison of the TSH response in 5 females taking estrogen containing contraceptive does not differ from the response in the 4 other females. The mean T3 concentration shows a significant increase 2 h after injection of T R H (Table 1). The female group shows a somewhat more pronounced increase than the male group. The difference is not statistically significant (Table 2). Estrogen containing drugs do not influence the T3 response. T4 does not change significantly over 2 h. 2. Oral Test

Oral stimulation with 40 mg of T R H : TSH rises from a basal value of 1.9 gU/ml to a maximum of 13.1 gU/ml 3 h after ingestion of TRH. The mean maximum is somewhat higher compared to the intravenous stimulation. The mean value after 8 h is still above the basal value. 3 h after the third stimulation mean plasma TSH is again elevated but to a mean maximum of only 5.8 gU/ml (Table 3). T3 in7 creases to a mean maximum of 2.19 ng/ml 3 h after 40 mg of TRH. After 8 h the mean T 3 concentration is only slightly above the basal value. 3 h after the third dose of T R H the mean T3 concentration is very similar to that 3 h after the first dose. The T3 increase of the oral stimulation is significantly more pronounced than after intravenous stimulation (2P 0.001). Thyroxine shows a significant increase 8 h af-' ter a single oral dose of 40 mg TRH. 3 h after the third dose, the mean thyroxine concentration is again significantly elevated above the basal value. 15 of 24 individuals experienced side effects after

1.3 7.I

intravenous injection (Nausea 7 x , feeling of hot flush 4 x , ill-defined " o d d feelings" 3 x ). After oral intake of 40 mg of T R H side effects have never been observed. Discussion The TSH response to intravenous injection of 200 gg of T R H is well documented [11, 32, 23, 31, 20, t2]. The extent of the TSH increase corresponds to previously published data. In the present series the maximum value was observed at 20 or 30 min. It is preferable therefore to take the 30 min rather than the 20 min value. A significant sex difference could not be found although all mean values are slightly higher in females compared to males and the mean maximum in females is observed after 20 min. An effect of estrogen containing oral contraceptives could not be shown. These findings are in agreement with the reports of Wenzel et al. [34] and contradict those of Lemarchand [22] and Faglia [10, 11] using the same dose of TRH. The determination at 60 min is of limited interest. A delayed maximum has been regarded as characteristic for a hypothalamic lesion [3, 9, 17]. The brief increase in plasma TSH is followed by a delayed, significant increase in T3. The findings in the present series correspond to those which have been published previously [25, 21, 29]. Although the mean increase in T 3 is significant a definitive interpretation in individual cases is not always possible, T 3 cannot therefore replace the TSH assay as a parameter of hypothalamo-pituitary thyroid function in response to i.v. TRH. The somewhat lower Ta increase in males reflects again the slight sex difference. As previously reported the thyroxine increase is too small to be useful. The normal response to an intravenous stimulation with 200 gg of T R H can therefore be defined as an increase to a TSH level within 2 standard deviations of the mean maximum reported. Addi-

34 tional information can be obtained by measuring the increase. An age-dependant decrease of T S H response to T R H has been shown in patients older than 60 years. The T S H stimulation in children is similar to that observed in the younger adult group [13, 20, 23]. Oral stimulation with a 200 times higher dose of T R H induced a slower but much more prolonged increase in T S H secretion which reaches its m a x i m u m 3 h after ingestion of 40 mg of T R H . In the present series the extent of T S H stimulation was not significantly different c o m p a r e d to intravenous stimulation. The long duration of T S H stimulation led however to a clear increase in triiodothyronine. A significant increase in thyroxine could be observed 8 h after intake of T R H . A significant sex difference or an influence o f oral contraceptives could again not be shown after this prolonged stimulation of T S H synthesis and secretion, The normal T S H response to oral stimulation with T R H has to our knowledge not yet been defined. Previous studies applied the prolonged stimulation of oral T R H to measure T3, T4 and PBI increase [31]. The few studies published on T S H response to oral T R H are based on different doses and different timing for the blood sampling [t5, 21, 28]. Repeated subsequent stimulation with 3 times 40 mg of T R H was expected to give a m o r e pronounced stimulation and subsequently higher TSH, T 3 and T4 values. The present results show however that the T S H response to the third stimulus is significantly lower compared to the T S H value 3 h after the first dose of T R H . The explanation could be seen in a feedback effect of the stimulated circulating T3 levels. The same effect is however seen in hypothyroid and thyroidectomised patients [28, 30, 35]. It must be assumed therefore that secretion of T S H is more rapidly stimulated than synthesis and that the capacity to synthesize T S H is exhausted in euthyroid patients. A short loop feedback of T S H on its own secretion can be considered. The situation in primary hypothyroidism can only be explained by a different steady state which allows a constant, high synthesis and secretion of TSH. The self-regulation against a continous stimulation with exogenous T R H is not fully explained. Long term treatment with oral T R H in psychotic patients or the treatment of hypothalamic-hypothyroidism with prolonged administration of T R H does not induce an overstimulation of thyroid activity [35]. The prolonged administration of T R H is however limited by its capacity to release prolactin [28, 32]. T R H is thus not destroyed after ingestion and its concentration in plasma measured by radioimmu-

T3

P. Vogt et al. : Oral and Intravenous TRH-Test noassay stays below the peak concentration after intravenous stimulation [27]. For practical purposes the significant increase in circulating T3 allows an estimation of the complete hypothalamo-pituitary thyroid axis after oral stimulation, Furthermore, the timing of blood sampling is not critical. In peadiatrics the oral route combined with a low volume T S H and T 3 assay would avoid venipuncture. Oral T R H does not lead to side effects. The intravenous test is of short duration and well studied. The m o r e prolonged and therefore more powerful stimulation after oral T R H is however useful when studying an insignificant or low T S H response to intravenous stimulation which can be observed in apparently euthyroid young adults and more often in elderly patients.

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35 25. PateI & Burger: Serum T 3 in Health and Disease. Clinical Endocrinology 35, 5 (I972) 26. Piva, F. &Steiner, H. : Bioassayand Toxicology of TRH. Animals Studies. Frontiers of hormone Research, Karger, Basel 1972 27. Powers, R., Wilber, J., Utiger, R , Montoya, E. : Thyrotropin (TSH) and protactin (HPr) inhibition by endogenous thyroxine (T4) and triiodothyronine (T3) after sequential oral thyrotropin releasing hormone (TRH). Clin. Res. 21, 866 (1973) 28. Rabello, M.M., Snyder, P.J., Utiger, R.D. : Effects on the pituitary-thyroid axis and prolactin secretion of single and repetitive oral doses of thyrotropin releasing hormone (TRH). J. clin. Endocr. 39, 571 (1974) 29~ Shenkman, L., Mitsuma, T., Hollander, C.S. : Modulation of pituitary responsiveness to thyrotropin-releasing hormone by triiodothyronin. J. clin. Invest. 52, 205 (1973) 30. Staub, J.J.: Diagnostische Anwendung yon TSH-ReleasingHormon (TRH) Untersuchung der hypophys~iren TSH-Reserve mit radioimmunologischer Bestimmung des TSH vor und nach oraIem und intraven6sem TRH. Habilitationsschrift 31. Staub, J.J., Weiss, S., Kohler, H., Bfirgi, H., K6nig, M.P., Studer, H.: Effekt yon peroral verabreichtem "TSH releasing hormone" (TRH) auf die Thyroxinkonzentration des Blutes. Schweiz. recd. Wschr. 101, 1295 (1971) 32. Torjesen, A., Haug, E., Sand, T.: Effect of TRH on serum tevels of pituitary hormones in men and women. Acta Endocrinologica, 1973 33. Vagenakis, A.G., Rapoport, B., Azizi, F,, Portnay, G.I., Braverman, L.E., Inbar, S.H. : Hyperresponse to thyrotropin-releasing hormone accompanying small decreases in serum thyroid hormone concentrations. J. clin. Invest. 54, 913 (1974) 34. Wenzel, K.W, Meinhold, H., Herpich, M., Adlkofer, F., Schleusener, H.: TRH Stimulationstest mit alters- und geschlechtsabhfingigem TSH-Anstieg bei Normalpersonen. Klin. Wschr. 52, 722 (1974) 35. Zuppinger, K.A., Joss, E.E., K6nig, M.P., Staub, J.J., Girard, J., Ehrengruber, H. : Effect of oral thyrotrophin-reteasing hormone in serum thyroxine in growth hormone-deficient and normal children. Clin. Endocr. 4, 119 (1975)

Prof. Dr. J. Girard Basler Kinderspital R6mergasse 8 CH-4005 Basel Schweiz