Pharmacology Biochemistry & Behavior, Vol. 3, pp. 665--670. Copyright © 1975 by ANKHO International Inc. All rights of reproduction in any form reserved. Printed in the U.S.A.
Thyrotropin Releasing Hormone (TRH): DOPA Potentiation and Biogenic Amine Studies N I C H O L A S P. P L O T N I K O F F
Experimental Therapy Division, Abbott Laboratories, North Chicago, IL 60064 AND G E O R G E R. B R E E S E A N D A R T H U R J. P R A N G E , J R .
Department o f Psyehiatry and Pharmacology, University o f North Carolina, School o f Medicine Chapel Hill, NC
(Received 8 N o v e m b e r 1974) PLOTNIKOFF, N. P., G. R. BREESE AND A. J. PRANGE, JR. Thyrotropin releasing hormone (TRH): DOPA potentiation and biogenic amine studies. PHARMAC. BIOCHEM. BEHAV. 3(4) 665-670, 1975. - The present study in mice demonstrated that TRH when administered over 5 days remained active in the Everett Dopamine Potentiation Test. No evidence of tolerance was observed. In fact, an accumulative effect of TRH appeared to take place. Ablation of the adrenals, ovaries, testes, pineal, spleen, parathyroid, one kidney, or thymus did not disrupt this behavioral potentiation of dopamine by TRH. TRH was found to potentiate the effects of imipramine. T3, T4, and TSH were found to be active in the DOPA potentiation test. No overt toxicity was observed between TRH and pargyline or between TRH and DOPA. Toxicity was seen only when all three agents were used together. TRH was found active in young and old mice. TRH was also found active in potentiating the central effects of serotonin. Biogenic amine brain levels in mice were not altered by TRH when administered for five days. Alpha-methyl-p-tyrosine reduced the activity of TRH in the dopamine potentiation test, suggesting dopaminergic mechanisms are involved by a direct receptor interaction. TRH
DOPA
Biogenic amines
THE finding b y o u r g r o u p t h a t t h y r o t r o p i n releasing h o r m o n e ( T R H ) is active in the p a r g y l i n e - D O P A m o u s e a c t i v a t i o n test f o r m e d in p a r t t h e m o t i v e f o r trials o f t h e h o r m o n e as a r e m e d y for d e p r e s s i o n [ 1 6 ] . Prange a n d Wilson [ 1 9 , 2 0 ] s h o w e d t h a t T R H has a fast o n s e t o f a n t i d e p r e s s a n t activity. This finding was c o n f i r m e d b y Kastin, et al. [10] a n d V a n d e r et al. [ 2 3 ] . H o w e v e r , m o r e r e c e n t r e p o r t s failed t o c o n f i r m t h e earlier studies [4, 13, 221. With these clinical findings at h a n d , it b e c a m e a m a t t e r o f i n t e r e s t t o e x a m i n e t h e activity o f T R H in m o r e detail o n t h e b e h a v i o r a l r e s p o n s e o f D O P A a n d to e x a m i n e t h e activity of related s u b s t a n c e s in this test. In t h e course o f this w o r k , it was r a t i o n a l to i n c l u d e in a d d i t i o n to t h e D O P A test, an a n i m a l test t h a t relates to cerebral indolea m i n e s b e c a u s e these s u b s t a n c e s have also b e e n i m p l i c a t e d in t h e p a t h o g e n e s i s o f d e p r e s s i o n [3, 6, 1 2 ] . T h e final phase o f t h e p r e s e n t w o r k was to e x a m i n e t h e i n f l u e n c e o f T R H o n levels o f cerebral b i o g e n i c a m i n e s in t h e b r a i n .
METHOD
Behavioral Studies DOPA potentiation test. T h e E v e r e t t D O P A / p a r g y l i n e P o t e n t i a t i o n Test was used in t h e p r e s e n t s t u d y [ 5 ] . T h e m e t h o d consists o f p r e t r e a t i n g mice ( 1 7 - 2 2 g) w i t h pargyline HC1 orally ( 4 0 mg/kg), at least 2 h r b e f o r e a d m i n i s t e r i n g T R H . A f t e r pargyline and T R H t r e a t m e n t , d 1 D O P A ( 2 0 0 m g / k g ) is given i n t r a p e r i t o n e a l l y and t h e animals (4 per cage) o b s e r v e d for 1 hr. One t o 3 cages were e m p l o y e d for e a c h dose. P o t e n t i a t i o n o f t h e D O P A - i n d u c e d response was scored as m a r k e d increase in irritability a n d reactivity, j u m p i n g , squeaking, a n d aggressive fighting. T h e Swiss albino mice used in these b e h a v i o r a l studies a n d a b l a t e d a n i m a l s were o b t a i n e d f r o m t h e Charles River L a b o r a t o r i e s a n d t h e A l t e c h L a b o r a t o r i e s , Madison, Wisconsin. Serotonin potentiation test. Just as D O P A is t h e a m i n o acid p r e c u r s o r of t h e c a t e c h o l a m i n e s , so is 5 - h y d r o x y 665
666
P L O T N I K O F F , B R E E S E A N D PRANGF
t r y p t o p h a n (5-HTP) the proximate precursor for serotonin. Because of the findings with DOPA, the effect of T R H on the biological responses to pargyline (40 mg/kg, PO) plus 5-HTP (100 mg/kg, IP) were studied. Characteristically, a syndrome develops which includes tremors, head movements, abduction of the limbs, and irritability. The effects are arbitrarily graded at 1 (for slight behavioral effects, 2 (for moderate effects), and 3 (for marked effects). Neuroehemical studies. Three to 4 groups (4 mice per dose) were used for each dose in the d e t e r m i n a t i o n of brain biogenic amines. ICR strain male mice ( 1 7 - 2 2 g) were used. The levels of dopamine, norepinephrine, and serotonin were determined by techniques previously reported from this laboratory [5].
TABLE 1 EFFECTS OF TRH ON DOPA RESPONSE IN PARGYLINETREATED MICE AFTER SINGLE AND MULTIPLE DOSE ADMINISTRATION Drug
Dose mg/kg (oral route)
Single dose
Chronic (5 days)
Control
I
1
1
TRH
0.1 0.2 0.4 0.8 1.6
i 1 2 3
3 3 3 3 3
RESULTS
Behavioral Studies' No loss of activity or f o r m a t i o n of tolerance to the behavioral p o t e n t i a t i o n of DOPA by T R H (Table 1) was observed. Thus, in a wide dose range (0.1 to 0.6 mg/kg) T R H was f o u n d to be active in the Everett D O P A Potentiation Test when administered daily for five days. Indeed, there appeared to be an accumulative effect resulting in greater p o t e n c y on the fifth day.
TABLE 2 EFFECT OF INTRAPERITONEAL TRH ON INTACT AND OVARIECTOMIZED FEMALE MICE IN THE DOPA POTENTIATION TEST*
TRH (mg/kg)
Behavioral rating
Ablation Studies Table 2 shows that intraperitoneal administration of T R H is effective in female mice. It also shows that activity of T R H is not lost in ovariectomized mice. By comparison with Table 3 it can be seen that there is a t e n d e n c y for T R H to be more p o t e n t in female (0.1 mg/kg) than in male mice (0.4 mg/kg) in the pargyline-DOPA test. By these criteria, it is more p o t e n t in normal female than ovariectomized mice as well. In a series of ablation studies, T R H was found to be active in the D O P A test in adrenalectomized, t h y m e c t o m i z e d , unilaterally n e p h r e c t o m i z e d , splenectomized, p a r a t h y r o i d e c t o m i z e d , castrated, and pinealectomized mice (Table 3). T R H appeared to be more active in p a r a t h y r o i d e c t o m i z e d , castrated, and pinealectomized mice than in normal intact male mice. TRH plus imipramine. In the present b o d y of work, the pargyline-DOPA test was used for a final time to discern whether T R H would amplify the well k n o w n effects of imipramine in this test model. Table 4 confirmed once again the activity of T R H and the activity of imipramine. F u r t h e r m o r e , it shows that the activity of the two substances potentiate or at least add to each other's effects (Table 4). Serotonin potentiation by TRH. Table 5 shows that TRH is active in the serotonin p o t e n t i a t i o n test. However, when T R H and imipramine were given togehter, no potentiation was observed. Imipramine at the highest dose tested 80 mg]kg, showed only m o d e r a t e activity. T3, T4, and TSH in the DOPA potentiation test. Table 6 clearly shows that T3, T4, and TSH are active in the DOPA potentiation test. Toxicity studies in mice. In Table 7 are shown data of interaction effects b e t w e e n T R H , pargyline, and d l DOPA. When all substances are used together in the D O P A potentiation test, a p p r o x i m a t e l y half the mice die overnight in coma (Group 1), however, no t o x i c i t y was observed b e t w e e n pargyline and T R H ( G r o u p 2). Also, no t o x i c i t y
Intact female mice 0.05 0.1 0.2 0.4 0.8 1.6
2 3 3 3 3 3
1 3 2 3 3 3
Ovariectomized mice 0.05 0.1 0.2 0.4 0.8 1.6
1
1
1
1
1
1
l
2
1
2
2 3 3
2 3 3
2 2 3
2 3 3
*Animals were treated and rated as in Table 1. TRH was administered 1 hr before DOPA. Rows of numbers indicate different experiments.
was seen b e t w e e n T R H and DOPA (Group 3) in overnight studies. TRH in older mice. T R H was found active in older mice (24 to 30 g) in the DOPA potentiation test. However, it was necessary to increase threshold by increasing the dose of pargyline from 40 to 45 mg/kg to d e m o n s t r a t e activity (Table 8). Alpha-methyl-p-tyrosine and TRH. Pretreatment of mice with alpha-methyl-p-tyrosine markedly reduced the activity of T R H in the DOPA p o t e n t i a t i o n test but did not abolish it (Table 9).
T R H A N D DOPA P O T E N T I A T I O N
667
TABLE 3 DOPA POTENTIATION BY TRH IN ABLATED MALE MICE
Surgical procedure None (intact)
Adrenalectomy
Thymectomy
Nephrectomy (unilateral)
Splenectomy
Parathyroidectomy
Castration
Dose mg/kg IP route
Behavioral rating at 1 hr
0.05
1
0.1 0.2
TABLE 4 POTENTIATION OF BEHAVIORAL EFFECTS OF DOPA IN MICE BY TRH AND IMIPRAMINE Behavioral rating Imipramine dose mg/kg (oral)
Imipramine alone
2
0.5
1
1
0.4 0.8
3 3
1
1
2
2
l
2
0.05
1
2.5
1
3
0.I 0.2
2 2
5
1
3
10
2
3
0.4
3
20
2
3
0.8
3
Imipramine plus TRH (0.1 mg/kg IP)
1
0.05
1
0.1
2
TABLE 5 EFFECTS OF TRH ON THE MOUSE 5-HTP POTENTIATION TEST
0.2
2
0.4
3
0.8
3
IP Route TRH alone
0.05 0.1
1 1
mg/kg Behavioral rating
0.2
2
0.4 0.8
3 3
0.05
1
0.1 0.2
1
2
0.4
3
0.8
3
0.05
2
0.1
3
0.2 0.4
3 3
0.8
3
0.05
1
0.1
2
0.2 0.4
3 3
0.8
3
0.1
2
0.2
3
0.4 0.8
3 2
Oral Route lmipramine alone mg/kg Behavior
lmipramine plus TRH* mg/kg Behavior
0.1
1
10
1
10
1
0.2
2
20
1
20
1
0.4
2
40
1
40
1
0.8
3
80
2
80
3
The doses of TRH were adminstered 1 hr prior to DL-5-HTP (100 mg/kg IP). All mic received pargyline (40 mg/kg, p.o.) 2 hr before TRH. The behavioral rating was based on a syndrome of tremors, head movements, abduction of limbs, and irritability and graded 1+ slight, 2+ moderate, and 3+ marked. *Dose o f TRH was 0.1 mg/kg which produced a 1+ slight effect.
t r e a t e d w i t h various doses o f IP T R H for 5 days or w i t h c o n t r o l l e d injection. Table 10 s h o w s t h a t n o n e o f the doses e m p l o y e d had a reliable effect on brain d o p a m i n e , nore p i n e p h r i n e , o r s e r o t o n i n . It can be seen that the behaviorally active doses o f T R H were w i t h o u t e f f e c t o n brain amines.
DISCUSSION Pinealectomy
The e f f e c t s o f T R H o n brain b i o g e n i c amines. The above w o r k s t r o n g l y suggested an i n t e r a c t i o n b e t w e e n T R H and brain biogenic amines. F o r this reason it was o f i n t e r e s t to e x a m i n e t h e e f f e c t s o f T R H t r e a t m e n t on the levels o f t h e s e substances. G r o u p s o f male Swiss-Webster mice were
It can be readily seen that the four hormones of the hypothalamic-pituitary-thyroid a x i s are active in the pargyline-DOPA test in mice. So far as has been discerned, TRH is active in both sexes. In female mice, the actions of TRH were not dependent upon the presence of the ovary. F u r t h e r m o r e , t h e a c t i v i t y o f T R H d e p e n d s n e i t h e r u p o n the p i t u i t a r y gland, n o r o n t h e t h y r o i d gland. Nevertheless, T3, T4, and TSH were active in the test. Thus, the possibility remains that increased secretion o f these s u b s t a n c e s a f t e r T R H could sustain the a n t i d e p r e s s a n t e f f e c t o f t h e h y p o t h a l a m i c h o r m o n e . T R H was also active in a test t h a t relates
668
PLOTNIKOFF,
BREESE
AND PRANGE
TABLE 6
TABLE 8
EFFECT OF T3, T4, AND TSH ON THE RESPONSE TO DOPA IN PARGYLINE T R E A T E D MICE
T R H IN THE DOPA POTENTIATION TEST IN O L D E R MICE* t
IP route (mg/kg)
Group
Degree of behavioral potentiation 1 hr
4 hr
Pargyline 40 mg/kg
Treatment
Plus
Behavioral rating at 1 hr
TRH (IP) mg/kg
T3
0.2
1
0.4 0.8
1 2
2.0
2
2
4.0
3
2
1.6
2
8.0
3
3
Control
1
T4 2.0
1
1
4.0
1
2
3
8.0
Pargyline 45 mg/kg
Plus
T R H (IP)
3
1
3
4.0
2
3
8.0
2
3
3
1.6
3
Control
1
*Mice 2 4 - 3 0 g body weight. t T R H plus dl DOPA (200 mg/kg IP).
TAB LE 7 CONTROL
TOXICITY
STUDIES IN MOUSE DOPA POTENTIATION TEST Treatment
Group
1. Pargyline (oral)
Plus
TRH* IP mg/kg
24 hr Mortality
40 mg/kg
0.05 mg/kg
40 mg/kg
0.I
mg/kg
4/8
40 mg/kg
0.2
mg/kg
4/8
40 mg/kg
0.4
mg/kg
4/8
40 mg/kg
0.8
mg/kg
4/8
2. Pargyline (oral)
TABLE 9 EFFECT OF ALPHA-METHYL-P-TYROSINE ON TRH IN THE DOPA POTENTIATION TEST
T R H (IP) plus DOPA (IP)
Control
1 2
0.8 TSH 2.0
0.2 0.4
Behavioral rating at 1 hr
0.4
1
0.8
2
1.6
3
4/8
0/8
*Pretreatment with alpha-methyl-p-tyrosine 300 mg/kg IP 4 hr prior to T R H administration. T A B L E 10 EFFECT OF TRH ON THE BRAIN BIOGENIC AMINES IN MICE* lag/g
TRH (IP)
40 mg/kg
1 mg/kg
0/4
40 mg/kg
10 mg/kg
0/4
40 mg/kg
100 mg/kg
0/4
3. T R H (IP)
DOPA (IP)
100 mg/kg
200 mg/kg
0/4
100 mg/kg
400 mg/kg
100 mg/kg
800 mg/kg
Oral route
Dopamine
Norepinephrine
Serotonin
0.91 ± 0.02
0.59 ± 0.01
0.55 ± 0.02
TRH 0.5 mg/kg
0.96 + 0.02
0.61 ± 0.01
0.56 + 0.05
0/4
1.0 mg/kg
0.94 _+0.03
0.57 -+ 0.02
0.52 -+ 0.01
0/4
2.0 mg/kg
0.91 -+ 0.01
0.55 ± 0.01
0.55 + 0.02
4.0 mg/kg
1.01 -+ 0.02
0.55 ± 0.01
0.51 -+ 0.02
8.0 mg/kg
0.93 -+ 0.01
0.55 ± 0.01
0.54 ± 0.01
Controls
*Animals were treated with various doses of TRH for 5 days and sacrificed 1 hr after the last dose of TRH.
TRH AND DOPA POTENTIATION
669
to the behavioral effects of indoleamines. F u r t h e r m o r e , when T R H was given daily for 5 days, there appeared an accumulative effect as judged by the results in the d o p a m i n e p o t e n t i a t i o n test. The present studies indicate that removal of the adrenals, thymus, one kidney, spleen, parathyroid, testes, ovaries, or pineal gland did n o t prevent the activity of T R H in the d o p a m i n e p o t e n t i a t i o n test. However, the p o t e n c y of T R H was reduced in ovariectomized and increased in castrated, p a r a t h y r o i d e c t o m i z e d , or pinealectomized mice compared to n o r m a l intact mice. Thus, there appear to be interactions b e t w e e n T R H and the secretions of these glands which will require further study. The antidepressant activities of T R H as well as imipramine appear to enhance each o t h e r in the d o p a m i n e p o t e n t i a t i o n test. However, the mechanisms of the two substances may be quite different. R e c e n t l y , Breese. e t al [1] found no effect of T R H on uptake o f H~-nor epinephrine in rat brain. This finding is in sharp contrast to imipramine which has significant effects on synaptic uptake and m e t a b o l i s m of amines. F u r t h e r m o r e , present findings are consistent with those of Breese, e t al, [1] that T R H does n o t elevate brain levels of d o p a m i n e , norepinephrine, and serotonin. Dopaminergic receptors are probably affected since our studies showed alpha-methyl-p-tyrosine reduced the effects of T R H in the D O P A test and d o p a m i n e has been shown to be elevated in T R H treated mice given DOPA after pargyline [ 1 ]. In addition, Keller e t aL [1], Breese, e t al. ( [ 2 ] , as well as Horst and Spirt [ 7 ] ) , have r e p o r t e d that T R H increases the rate of norepinephrine t u r n o v e r and release in brain tissue. This may be consistent with the finding of Reigle e t aL [14] who r e p o r t e d that T R H p r o d u c e d an increase in
brain levels of tritiated n o r m e t a n e p h r i n e in rats. R e c e n t l y , P l o t n i k o f f et al. [14,15] r e p o r t e d that MIF also potentiates the actions o f D O P A and at the same time does not elevate brain levels o f d o p a m i n e , norepinephrine, and serotonin. These findings with T R H and MIF in the DOPA test were confirmed by H u i d o b r o - T o r o et al. [8]. However, by way of contrast, MIF has been reported not to be active in the 5-HTP test (Plotnikoff). In another place we have suggested that b o t h serotonin and catecholamines are involved in the chemical pathogenesis of depression, and that in depression b o t h groups of substances are deficient [17,18]. It is also of interest to n o t e that MIF has been found useful in the t r e a t m e n t of Parkinson's disease [9] while T R H appears to be inactive. These findings are consistent with the view that in Parkinson's disease a disruption of the normal balance of amines is involved [18]. Thus, peptides which p r o m o t e catecholamine activity a n d / o r indoleamine activity might reasonably be expected to have activity in psychiatric a n d / o r neurological disorders. In this regard, no t o x i c interactions were observed between pargyline and T R H or b e t w e e n DOPA and T R H . T o x i c i t y was observed only when all three substances were administered to test animals. In addition we f o u n d that the level of threshold changes of m o n o a m i n o - o x i d a s e inhibition was critical in relationship to age. The older mice required a higher dose of pargyline to produce behavioral potentiation of d o p a m i n e by T R H . It appears that critical levels of biogenic amines are required for behavioral expression of T R H effects. It is possible that T R H may be increasing the sensitivity o f dopaminergic receptor systems.
REFERENCES 1. Breese, G. R., B. R. Cooper, A. J. Prange, Jr., J. M. Cott and M. A. Lipton. In: The Thyroid Axis, Drugs and Behavior, edited by A. J. Prange, Jr. New York: Raven Press, 1974. 2. Breese, G. R., Cott, J. M., Cooper, B. R., Prange, A. J., Lipton, M. A., and Plotnikoff, N. P.: Effects of Thyrotropin Releasing Hormone (TRH) on the Actions of Pentobarbital and other Centrally Acting Drugs. J. Pharmac exp. Therap. In Press. 3. Coppen, A. The biochemistry of affective disorders. Br. J. Psychia. 113: 1237-1264, 1967. 4. Coppen, A., M. Peer, S. Montgomery and J. Bailey. Thyrotropin-releasing hormone in the treatment of depression. Lancet 2: 433-435, 1974. 5. Everett, G. M. The DOPA response potentiation test and its use in screening for antidepressant drugs. Excerpta reed. 122: 164-167, 1966. 6. Glassman, A. Indoleamines and affective disorders. Psychosom. Med. 31: 107-114, 1969. 7. Horst, W. D., and N. Spirt. A possible mechanism for the antidepressant activity of thyrotropin-releasing hormone. Life Sci. 15: 1073-1082, 1974. 8. Huidobro-Toro, J. P., A. Scotti De Carolis and V. G. Longo. Action of two hypothalamic factors (TRH, MIF) and of angiotensin II on the behavioral effects of L-DOPA and 5-hydroxytryptophan in mice. Pharmac. Biochem. Behav. 2: 105-109, 1974. 9. Kastin, A. J. and A. Barbeau. Preliminary clinical studies with L-prolyl-l-leucyl glycine amide in Parkinson's disease. J. can. med. Ass. 107: 1079-1081, 1973. 10. Kastin, A. B., R. H. Ehrensing, D. S. Schalch and M. S. Anderson. Improvement in mental depression with decreased thyrotropin response after administration of thyrotropinreleasing hormone. Lacet 2: 740-742, 1972.
11. Keller, H. H., G. Bartholini and A. Pletscher. Enhancement of cerebral noradrenaline turnover by thyrotropin-releasing hormone. Lancet 248: 528-529, 1974. 12. Lapin, I. P. and G. F. Oxenkrug. Intensification of the central serotoninergic process as a possible determinant of the thymoleptic effect. Lancet 1:132-136, 1969. 13. Mountjoy, C. Q., M. Weller, R. Hall, J. S. Price, P. Hunter and J. H. Dewar. A double-blind crossover sequential trial of oral thyrotropin-releasing hormone in depression. Lancet 2: 958 -960. 14, Plotnikoff, N. P., A. J. Kastin, M. S. Anderson and A. V. Schally. DOPA potentiation by a hypothalamic factor, MSH release-inhibiting hormone (MIF). Life Sci. 10: 1279-1283, 1971. 15, Plotnikoff, N. P., F. N. Minard and A. J. Kastin. DOPA potentiation in ablated animals and brain levels of biogenic amines in intact animals after prolyl-leucylglycinamide. Neuroendocrinology 14: 271-179, 1974. 16. Plotnikoff, N. P:, A. J. Prange, Jr., G. R. Breese, M. S. Anderson and I. C. Wilson. Thyrotropin-releasing hormone: Enhancement of DOPA activity by a hypothalamic hormone. Science 178: 417, 1972. 17. Prange, A. J., Jr. The use of drugs in depression: its theoretical and practical basis. Psychiat. Ann. 13: 56-75, 1973. 18. Prange, A. J., J. L. Sisk, I. C. Wilson, C. E. Morris, C. D. Hall and J. S. Carman. Balance, permission, and discrimination among amines: a theoretical consideration of L-tryptophan in disorders of movement and affect. In: Serotonin and Behavior, edited by J. D. Barchas and E. Usdin, New York: Academic Press, in press.
670 19. Prange, A. J., Jr. and I. C. Wilson. Thyrotropin-releasing hormone (TRH) for the immediate relief of depression: a preliminary report. Psychopharmacologia suppl. 26: 82, 1972. 20. Prange, A. J., Jr., Wilson, I. C., Lara, P. P., Alltop, L. B., and Breese, G. R. (1972): Effects of thyrotropin-releasing hormone in depression. Lancet, 2: 999-1002. 21. Reigle, T. G., J. Avni, P. A. Platz, J. J. Schildkraut, and N. P. Plotnikoff. Norepinephrine metabolism in the rat brain following acute and chronic administration of thyrotropin-releasing hormone. Pwchopharmacologia, 37: 1 - 6 , 1974.
PLOTNIKOFF, BREESE AND PRANGE 22. Takahashi, S., H. Kondo, M. Yoshimura and Y. Ochi. Antidepressant effect of thyrotropin-releasing hormone (TRH) and the plasma thyrotropin levels in depression. Folia Psychiat. neurol, jap. 27: 4 , 3 0 5 - 3 1 4 , 1973. 23. Van der Vis-Melsen, M. J. E. and J. D. Wiener. Improvement in mental depression with decreased thyrotropin response after administration of thyrotropin-releasing hormone. Lancet 2: 1415, 1972.
Thyrotropin Releasing Hormone (TRH): DOPA Potentiation and Biogenic Amine Studies N I C H O L A S P. P L O T N I K O F F
Experimental Therapy Division, Abbott Laboratories, North Chicago, IL 60064 AND G E O R G E R. B R E E S E A N D A R T H U R J. P R A N G E , J R .
Department o f Psyehiatry and Pharmacology, University o f North Carolina, School o f Medicine Chapel Hill, NC
(Received 8 N o v e m b e r 1974) PLOTNIKOFF, N. P., G. R. BREESE AND A. J. PRANGE, JR. Thyrotropin releasing hormone (TRH): DOPA potentiation and biogenic amine studies. PHARMAC. BIOCHEM. BEHAV. 3(4) 665-670, 1975. - The present study in mice demonstrated that TRH when administered over 5 days remained active in the Everett Dopamine Potentiation Test. No evidence of tolerance was observed. In fact, an accumulative effect of TRH appeared to take place. Ablation of the adrenals, ovaries, testes, pineal, spleen, parathyroid, one kidney, or thymus did not disrupt this behavioral potentiation of dopamine by TRH. TRH was found to potentiate the effects of imipramine. T3, T4, and TSH were found to be active in the DOPA potentiation test. No overt toxicity was observed between TRH and pargyline or between TRH and DOPA. Toxicity was seen only when all three agents were used together. TRH was found active in young and old mice. TRH was also found active in potentiating the central effects of serotonin. Biogenic amine brain levels in mice were not altered by TRH when administered for five days. Alpha-methyl-p-tyrosine reduced the activity of TRH in the dopamine potentiation test, suggesting dopaminergic mechanisms are involved by a direct receptor interaction. TRH
DOPA
Biogenic amines
THE finding b y o u r g r o u p t h a t t h y r o t r o p i n releasing h o r m o n e ( T R H ) is active in the p a r g y l i n e - D O P A m o u s e a c t i v a t i o n test f o r m e d in p a r t t h e m o t i v e f o r trials o f t h e h o r m o n e as a r e m e d y for d e p r e s s i o n [ 1 6 ] . Prange a n d Wilson [ 1 9 , 2 0 ] s h o w e d t h a t T R H has a fast o n s e t o f a n t i d e p r e s s a n t activity. This finding was c o n f i r m e d b y Kastin, et al. [10] a n d V a n d e r et al. [ 2 3 ] . H o w e v e r , m o r e r e c e n t r e p o r t s failed t o c o n f i r m t h e earlier studies [4, 13, 221. With these clinical findings at h a n d , it b e c a m e a m a t t e r o f i n t e r e s t t o e x a m i n e t h e activity o f T R H in m o r e detail o n t h e b e h a v i o r a l r e s p o n s e o f D O P A a n d to e x a m i n e t h e activity of related s u b s t a n c e s in this test. In t h e course o f this w o r k , it was r a t i o n a l to i n c l u d e in a d d i t i o n to t h e D O P A test, an a n i m a l test t h a t relates to cerebral indolea m i n e s b e c a u s e these s u b s t a n c e s have also b e e n i m p l i c a t e d in t h e p a t h o g e n e s i s o f d e p r e s s i o n [3, 6, 1 2 ] . T h e final phase o f t h e p r e s e n t w o r k was to e x a m i n e t h e i n f l u e n c e o f T R H o n levels o f cerebral b i o g e n i c a m i n e s in t h e b r a i n .
METHOD
Behavioral Studies DOPA potentiation test. T h e E v e r e t t D O P A / p a r g y l i n e P o t e n t i a t i o n Test was used in t h e p r e s e n t s t u d y [ 5 ] . T h e m e t h o d consists o f p r e t r e a t i n g mice ( 1 7 - 2 2 g) w i t h pargyline HC1 orally ( 4 0 mg/kg), at least 2 h r b e f o r e a d m i n i s t e r i n g T R H . A f t e r pargyline and T R H t r e a t m e n t , d 1 D O P A ( 2 0 0 m g / k g ) is given i n t r a p e r i t o n e a l l y and t h e animals (4 per cage) o b s e r v e d for 1 hr. One t o 3 cages were e m p l o y e d for e a c h dose. P o t e n t i a t i o n o f t h e D O P A - i n d u c e d response was scored as m a r k e d increase in irritability a n d reactivity, j u m p i n g , squeaking, a n d aggressive fighting. T h e Swiss albino mice used in these b e h a v i o r a l studies a n d a b l a t e d a n i m a l s were o b t a i n e d f r o m t h e Charles River L a b o r a t o r i e s a n d t h e A l t e c h L a b o r a t o r i e s , Madison, Wisconsin. Serotonin potentiation test. Just as D O P A is t h e a m i n o acid p r e c u r s o r of t h e c a t e c h o l a m i n e s , so is 5 - h y d r o x y 665
666
P L O T N I K O F F , B R E E S E A N D PRANGF
t r y p t o p h a n (5-HTP) the proximate precursor for serotonin. Because of the findings with DOPA, the effect of T R H on the biological responses to pargyline (40 mg/kg, PO) plus 5-HTP (100 mg/kg, IP) were studied. Characteristically, a syndrome develops which includes tremors, head movements, abduction of the limbs, and irritability. The effects are arbitrarily graded at 1 (for slight behavioral effects, 2 (for moderate effects), and 3 (for marked effects). Neuroehemical studies. Three to 4 groups (4 mice per dose) were used for each dose in the d e t e r m i n a t i o n of brain biogenic amines. ICR strain male mice ( 1 7 - 2 2 g) were used. The levels of dopamine, norepinephrine, and serotonin were determined by techniques previously reported from this laboratory [5].
TABLE 1 EFFECTS OF TRH ON DOPA RESPONSE IN PARGYLINETREATED MICE AFTER SINGLE AND MULTIPLE DOSE ADMINISTRATION Drug
Dose mg/kg (oral route)
Single dose
Chronic (5 days)
Control
I
1
1
TRH
0.1 0.2 0.4 0.8 1.6
i 1 2 3
3 3 3 3 3
RESULTS
Behavioral Studies' No loss of activity or f o r m a t i o n of tolerance to the behavioral p o t e n t i a t i o n of DOPA by T R H (Table 1) was observed. Thus, in a wide dose range (0.1 to 0.6 mg/kg) T R H was f o u n d to be active in the Everett D O P A Potentiation Test when administered daily for five days. Indeed, there appeared to be an accumulative effect resulting in greater p o t e n c y on the fifth day.
TABLE 2 EFFECT OF INTRAPERITONEAL TRH ON INTACT AND OVARIECTOMIZED FEMALE MICE IN THE DOPA POTENTIATION TEST*
TRH (mg/kg)
Behavioral rating
Ablation Studies Table 2 shows that intraperitoneal administration of T R H is effective in female mice. It also shows that activity of T R H is not lost in ovariectomized mice. By comparison with Table 3 it can be seen that there is a t e n d e n c y for T R H to be more p o t e n t in female (0.1 mg/kg) than in male mice (0.4 mg/kg) in the pargyline-DOPA test. By these criteria, it is more p o t e n t in normal female than ovariectomized mice as well. In a series of ablation studies, T R H was found to be active in the D O P A test in adrenalectomized, t h y m e c t o m i z e d , unilaterally n e p h r e c t o m i z e d , splenectomized, p a r a t h y r o i d e c t o m i z e d , castrated, and pinealectomized mice (Table 3). T R H appeared to be more active in p a r a t h y r o i d e c t o m i z e d , castrated, and pinealectomized mice than in normal intact male mice. TRH plus imipramine. In the present b o d y of work, the pargyline-DOPA test was used for a final time to discern whether T R H would amplify the well k n o w n effects of imipramine in this test model. Table 4 confirmed once again the activity of T R H and the activity of imipramine. F u r t h e r m o r e , it shows that the activity of the two substances potentiate or at least add to each other's effects (Table 4). Serotonin potentiation by TRH. Table 5 shows that TRH is active in the serotonin p o t e n t i a t i o n test. However, when T R H and imipramine were given togehter, no potentiation was observed. Imipramine at the highest dose tested 80 mg]kg, showed only m o d e r a t e activity. T3, T4, and TSH in the DOPA potentiation test. Table 6 clearly shows that T3, T4, and TSH are active in the DOPA potentiation test. Toxicity studies in mice. In Table 7 are shown data of interaction effects b e t w e e n T R H , pargyline, and d l DOPA. When all substances are used together in the D O P A potentiation test, a p p r o x i m a t e l y half the mice die overnight in coma (Group 1), however, no t o x i c i t y was observed b e t w e e n pargyline and T R H ( G r o u p 2). Also, no t o x i c i t y
Intact female mice 0.05 0.1 0.2 0.4 0.8 1.6
2 3 3 3 3 3
1 3 2 3 3 3
Ovariectomized mice 0.05 0.1 0.2 0.4 0.8 1.6
1
1
1
1
1
1
l
2
1
2
2 3 3
2 3 3
2 2 3
2 3 3
*Animals were treated and rated as in Table 1. TRH was administered 1 hr before DOPA. Rows of numbers indicate different experiments.
was seen b e t w e e n T R H and DOPA (Group 3) in overnight studies. TRH in older mice. T R H was found active in older mice (24 to 30 g) in the DOPA potentiation test. However, it was necessary to increase threshold by increasing the dose of pargyline from 40 to 45 mg/kg to d e m o n s t r a t e activity (Table 8). Alpha-methyl-p-tyrosine and TRH. Pretreatment of mice with alpha-methyl-p-tyrosine markedly reduced the activity of T R H in the DOPA p o t e n t i a t i o n test but did not abolish it (Table 9).
T R H A N D DOPA P O T E N T I A T I O N
667
TABLE 3 DOPA POTENTIATION BY TRH IN ABLATED MALE MICE
Surgical procedure None (intact)
Adrenalectomy
Thymectomy
Nephrectomy (unilateral)
Splenectomy
Parathyroidectomy
Castration
Dose mg/kg IP route
Behavioral rating at 1 hr
0.05
1
0.1 0.2
TABLE 4 POTENTIATION OF BEHAVIORAL EFFECTS OF DOPA IN MICE BY TRH AND IMIPRAMINE Behavioral rating Imipramine dose mg/kg (oral)
Imipramine alone
2
0.5
1
1
0.4 0.8
3 3
1
1
2
2
l
2
0.05
1
2.5
1
3
0.I 0.2
2 2
5
1
3
10
2
3
0.4
3
20
2
3
0.8
3
Imipramine plus TRH (0.1 mg/kg IP)
1
0.05
1
0.1
2
TABLE 5 EFFECTS OF TRH ON THE MOUSE 5-HTP POTENTIATION TEST
0.2
2
0.4
3
0.8
3
IP Route TRH alone
0.05 0.1
1 1
mg/kg Behavioral rating
0.2
2
0.4 0.8
3 3
0.05
1
0.1 0.2
1
2
0.4
3
0.8
3
0.05
2
0.1
3
0.2 0.4
3 3
0.8
3
0.05
1
0.1
2
0.2 0.4
3 3
0.8
3
0.1
2
0.2
3
0.4 0.8
3 2
Oral Route lmipramine alone mg/kg Behavior
lmipramine plus TRH* mg/kg Behavior
0.1
1
10
1
10
1
0.2
2
20
1
20
1
0.4
2
40
1
40
1
0.8
3
80
2
80
3
The doses of TRH were adminstered 1 hr prior to DL-5-HTP (100 mg/kg IP). All mic received pargyline (40 mg/kg, p.o.) 2 hr before TRH. The behavioral rating was based on a syndrome of tremors, head movements, abduction of limbs, and irritability and graded 1+ slight, 2+ moderate, and 3+ marked. *Dose o f TRH was 0.1 mg/kg which produced a 1+ slight effect.
t r e a t e d w i t h various doses o f IP T R H for 5 days or w i t h c o n t r o l l e d injection. Table 10 s h o w s t h a t n o n e o f the doses e m p l o y e d had a reliable effect on brain d o p a m i n e , nore p i n e p h r i n e , o r s e r o t o n i n . It can be seen that the behaviorally active doses o f T R H were w i t h o u t e f f e c t o n brain amines.
DISCUSSION Pinealectomy
The e f f e c t s o f T R H o n brain b i o g e n i c amines. The above w o r k s t r o n g l y suggested an i n t e r a c t i o n b e t w e e n T R H and brain biogenic amines. F o r this reason it was o f i n t e r e s t to e x a m i n e t h e e f f e c t s o f T R H t r e a t m e n t on the levels o f t h e s e substances. G r o u p s o f male Swiss-Webster mice were
It can be readily seen that the four hormones of the hypothalamic-pituitary-thyroid a x i s are active in the pargyline-DOPA test in mice. So far as has been discerned, TRH is active in both sexes. In female mice, the actions of TRH were not dependent upon the presence of the ovary. F u r t h e r m o r e , t h e a c t i v i t y o f T R H d e p e n d s n e i t h e r u p o n the p i t u i t a r y gland, n o r o n t h e t h y r o i d gland. Nevertheless, T3, T4, and TSH were active in the test. Thus, the possibility remains that increased secretion o f these s u b s t a n c e s a f t e r T R H could sustain the a n t i d e p r e s s a n t e f f e c t o f t h e h y p o t h a l a m i c h o r m o n e . T R H was also active in a test t h a t relates
668
PLOTNIKOFF,
BREESE
AND PRANGE
TABLE 6
TABLE 8
EFFECT OF T3, T4, AND TSH ON THE RESPONSE TO DOPA IN PARGYLINE T R E A T E D MICE
T R H IN THE DOPA POTENTIATION TEST IN O L D E R MICE* t
IP route (mg/kg)
Group
Degree of behavioral potentiation 1 hr
4 hr
Pargyline 40 mg/kg
Treatment
Plus
Behavioral rating at 1 hr
TRH (IP) mg/kg
T3
0.2
1
0.4 0.8
1 2
2.0
2
2
4.0
3
2
1.6
2
8.0
3
3
Control
1
T4 2.0
1
1
4.0
1
2
3
8.0
Pargyline 45 mg/kg
Plus
T R H (IP)
3
1
3
4.0
2
3
8.0
2
3
3
1.6
3
Control
1
*Mice 2 4 - 3 0 g body weight. t T R H plus dl DOPA (200 mg/kg IP).
TAB LE 7 CONTROL
TOXICITY
STUDIES IN MOUSE DOPA POTENTIATION TEST Treatment
Group
1. Pargyline (oral)
Plus
TRH* IP mg/kg
24 hr Mortality
40 mg/kg
0.05 mg/kg
40 mg/kg
0.I
mg/kg
4/8
40 mg/kg
0.2
mg/kg
4/8
40 mg/kg
0.4
mg/kg
4/8
40 mg/kg
0.8
mg/kg
4/8
2. Pargyline (oral)
TABLE 9 EFFECT OF ALPHA-METHYL-P-TYROSINE ON TRH IN THE DOPA POTENTIATION TEST
T R H (IP) plus DOPA (IP)
Control
1 2
0.8 TSH 2.0
0.2 0.4
Behavioral rating at 1 hr
0.4
1
0.8
2
1.6
3
4/8
0/8
*Pretreatment with alpha-methyl-p-tyrosine 300 mg/kg IP 4 hr prior to T R H administration. T A B L E 10 EFFECT OF TRH ON THE BRAIN BIOGENIC AMINES IN MICE* lag/g
TRH (IP)
40 mg/kg
1 mg/kg
0/4
40 mg/kg
10 mg/kg
0/4
40 mg/kg
100 mg/kg
0/4
3. T R H (IP)
DOPA (IP)
100 mg/kg
200 mg/kg
0/4
100 mg/kg
400 mg/kg
100 mg/kg
800 mg/kg
Oral route
Dopamine
Norepinephrine
Serotonin
0.91 ± 0.02
0.59 ± 0.01
0.55 ± 0.02
TRH 0.5 mg/kg
0.96 + 0.02
0.61 ± 0.01
0.56 + 0.05
0/4
1.0 mg/kg
0.94 _+0.03
0.57 -+ 0.02
0.52 -+ 0.01
0/4
2.0 mg/kg
0.91 -+ 0.01
0.55 ± 0.01
0.55 + 0.02
4.0 mg/kg
1.01 -+ 0.02
0.55 ± 0.01
0.51 -+ 0.02
8.0 mg/kg
0.93 -+ 0.01
0.55 ± 0.01
0.54 ± 0.01
Controls
*Animals were treated with various doses of TRH for 5 days and sacrificed 1 hr after the last dose of TRH.
TRH AND DOPA POTENTIATION
669
to the behavioral effects of indoleamines. F u r t h e r m o r e , when T R H was given daily for 5 days, there appeared an accumulative effect as judged by the results in the d o p a m i n e p o t e n t i a t i o n test. The present studies indicate that removal of the adrenals, thymus, one kidney, spleen, parathyroid, testes, ovaries, or pineal gland did n o t prevent the activity of T R H in the d o p a m i n e p o t e n t i a t i o n test. However, the p o t e n c y of T R H was reduced in ovariectomized and increased in castrated, p a r a t h y r o i d e c t o m i z e d , or pinealectomized mice compared to n o r m a l intact mice. Thus, there appear to be interactions b e t w e e n T R H and the secretions of these glands which will require further study. The antidepressant activities of T R H as well as imipramine appear to enhance each o t h e r in the d o p a m i n e p o t e n t i a t i o n test. However, the mechanisms of the two substances may be quite different. R e c e n t l y , Breese. e t al [1] found no effect of T R H on uptake o f H~-nor epinephrine in rat brain. This finding is in sharp contrast to imipramine which has significant effects on synaptic uptake and m e t a b o l i s m of amines. F u r t h e r m o r e , present findings are consistent with those of Breese, e t al, [1] that T R H does n o t elevate brain levels of d o p a m i n e , norepinephrine, and serotonin. Dopaminergic receptors are probably affected since our studies showed alpha-methyl-p-tyrosine reduced the effects of T R H in the D O P A test and d o p a m i n e has been shown to be elevated in T R H treated mice given DOPA after pargyline [ 1 ]. In addition, Keller e t aL [1], Breese, e t al. ( [ 2 ] , as well as Horst and Spirt [ 7 ] ) , have r e p o r t e d that T R H increases the rate of norepinephrine t u r n o v e r and release in brain tissue. This may be consistent with the finding of Reigle e t aL [14] who r e p o r t e d that T R H p r o d u c e d an increase in
brain levels of tritiated n o r m e t a n e p h r i n e in rats. R e c e n t l y , P l o t n i k o f f et al. [14,15] r e p o r t e d that MIF also potentiates the actions o f D O P A and at the same time does not elevate brain levels o f d o p a m i n e , norepinephrine, and serotonin. These findings with T R H and MIF in the DOPA test were confirmed by H u i d o b r o - T o r o et al. [8]. However, by way of contrast, MIF has been reported not to be active in the 5-HTP test (Plotnikoff). In another place we have suggested that b o t h serotonin and catecholamines are involved in the chemical pathogenesis of depression, and that in depression b o t h groups of substances are deficient [17,18]. It is also of interest to n o t e that MIF has been found useful in the t r e a t m e n t of Parkinson's disease [9] while T R H appears to be inactive. These findings are consistent with the view that in Parkinson's disease a disruption of the normal balance of amines is involved [18]. Thus, peptides which p r o m o t e catecholamine activity a n d / o r indoleamine activity might reasonably be expected to have activity in psychiatric a n d / o r neurological disorders. In this regard, no t o x i c interactions were observed between pargyline and T R H or b e t w e e n DOPA and T R H . T o x i c i t y was observed only when all three substances were administered to test animals. In addition we f o u n d that the level of threshold changes of m o n o a m i n o - o x i d a s e inhibition was critical in relationship to age. The older mice required a higher dose of pargyline to produce behavioral potentiation of d o p a m i n e by T R H . It appears that critical levels of biogenic amines are required for behavioral expression of T R H effects. It is possible that T R H may be increasing the sensitivity o f dopaminergic receptor systems.
REFERENCES 1. Breese, G. R., B. R. Cooper, A. J. Prange, Jr., J. M. Cott and M. A. Lipton. In: The Thyroid Axis, Drugs and Behavior, edited by A. J. Prange, Jr. New York: Raven Press, 1974. 2. Breese, G. R., Cott, J. M., Cooper, B. R., Prange, A. J., Lipton, M. A., and Plotnikoff, N. P.: Effects of Thyrotropin Releasing Hormone (TRH) on the Actions of Pentobarbital and other Centrally Acting Drugs. J. Pharmac exp. Therap. In Press. 3. Coppen, A. The biochemistry of affective disorders. Br. J. Psychia. 113: 1237-1264, 1967. 4. Coppen, A., M. Peer, S. Montgomery and J. Bailey. Thyrotropin-releasing hormone in the treatment of depression. Lancet 2: 433-435, 1974. 5. Everett, G. M. The DOPA response potentiation test and its use in screening for antidepressant drugs. Excerpta reed. 122: 164-167, 1966. 6. Glassman, A. Indoleamines and affective disorders. Psychosom. Med. 31: 107-114, 1969. 7. Horst, W. D., and N. Spirt. A possible mechanism for the antidepressant activity of thyrotropin-releasing hormone. Life Sci. 15: 1073-1082, 1974. 8. Huidobro-Toro, J. P., A. Scotti De Carolis and V. G. Longo. Action of two hypothalamic factors (TRH, MIF) and of angiotensin II on the behavioral effects of L-DOPA and 5-hydroxytryptophan in mice. Pharmac. Biochem. Behav. 2: 105-109, 1974. 9. Kastin, A. J. and A. Barbeau. Preliminary clinical studies with L-prolyl-l-leucyl glycine amide in Parkinson's disease. J. can. med. Ass. 107: 1079-1081, 1973. 10. Kastin, A. B., R. H. Ehrensing, D. S. Schalch and M. S. Anderson. Improvement in mental depression with decreased thyrotropin response after administration of thyrotropinreleasing hormone. Lacet 2: 740-742, 1972.
11. Keller, H. H., G. Bartholini and A. Pletscher. Enhancement of cerebral noradrenaline turnover by thyrotropin-releasing hormone. Lancet 248: 528-529, 1974. 12. Lapin, I. P. and G. F. Oxenkrug. Intensification of the central serotoninergic process as a possible determinant of the thymoleptic effect. Lancet 1:132-136, 1969. 13. Mountjoy, C. Q., M. Weller, R. Hall, J. S. Price, P. Hunter and J. H. Dewar. A double-blind crossover sequential trial of oral thyrotropin-releasing hormone in depression. Lancet 2: 958 -960. 14, Plotnikoff, N. P., A. J. Kastin, M. S. Anderson and A. V. Schally. DOPA potentiation by a hypothalamic factor, MSH release-inhibiting hormone (MIF). Life Sci. 10: 1279-1283, 1971. 15, Plotnikoff, N. P., F. N. Minard and A. J. Kastin. DOPA potentiation in ablated animals and brain levels of biogenic amines in intact animals after prolyl-leucylglycinamide. Neuroendocrinology 14: 271-179, 1974. 16. Plotnikoff, N. P:, A. J. Prange, Jr., G. R. Breese, M. S. Anderson and I. C. Wilson. Thyrotropin-releasing hormone: Enhancement of DOPA activity by a hypothalamic hormone. Science 178: 417, 1972. 17. Prange, A. J., Jr. The use of drugs in depression: its theoretical and practical basis. Psychiat. Ann. 13: 56-75, 1973. 18. Prange, A. J., J. L. Sisk, I. C. Wilson, C. E. Morris, C. D. Hall and J. S. Carman. Balance, permission, and discrimination among amines: a theoretical consideration of L-tryptophan in disorders of movement and affect. In: Serotonin and Behavior, edited by J. D. Barchas and E. Usdin, New York: Academic Press, in press.
670 19. Prange, A. J., Jr. and I. C. Wilson. Thyrotropin-releasing hormone (TRH) for the immediate relief of depression: a preliminary report. Psychopharmacologia suppl. 26: 82, 1972. 20. Prange, A. J., Jr., Wilson, I. C., Lara, P. P., Alltop, L. B., and Breese, G. R. (1972): Effects of thyrotropin-releasing hormone in depression. Lancet, 2: 999-1002. 21. Reigle, T. G., J. Avni, P. A. Platz, J. J. Schildkraut, and N. P. Plotnikoff. Norepinephrine metabolism in the rat brain following acute and chronic administration of thyrotropin-releasing hormone. Pwchopharmacologia, 37: 1 - 6 , 1974.
PLOTNIKOFF, BREESE AND PRANGE 22. Takahashi, S., H. Kondo, M. Yoshimura and Y. Ochi. Antidepressant effect of thyrotropin-releasing hormone (TRH) and the plasma thyrotropin levels in depression. Folia Psychiat. neurol, jap. 27: 4 , 3 0 5 - 3 1 4 , 1973. 23. Van der Vis-Melsen, M. J. E. and J. D. Wiener. Improvement in mental depression with decreased thyrotropin response after administration of thyrotropin-releasing hormone. Lancet 2: 1415, 1972.
