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Mesolimbic involvement in the locomotor stimulant action of thyrotropin-releasing hormone (TRH) in rats
European Journal o f Pharmacology, 44 (1977) 143--152
143
© Elsevier/North-Holland Biomedical Press
MESOLIMBIC INVOLVEMENT IN THE LOCOMOTOR STIMULANT ACTION OF T H Y R O T R O P I N - R E L E A S I N G H O R M O N E ( T R H ) IN R A T S MASAOMI MIYAMOTO and YUJI NAGAWA Medicinal Laboratories, Central Research Division, Takeda Chemical Industries, Ltd., Osaka 532, Japan
Received 16 November 1976, revised MS received 18 February 1977, accepted 24 March 1977
M. MIYAMOTO and Y. NAGAWA, Mesolimbic involvement in the locomotor stimulant action o f thyrotropinreleasing hormone (TRH) in rats, European J. Pharmacol, 44 (1977) 143--152. Thyrotropin-releasing hormone (TRH) injected i.p. in doses of 5 mg/kg and higher had a strong locomotor stimulant action with development of frequent rearing, mild sniffing, grooming, preening and other excitatory behaviours. The locomotor stimulation was also produced by bilateral injection of TRH and dopamine (DA) into the nucleus accumbens but not by bilateral injection of these substances into the caudate nucleus. Unilateral intracaudate injection of TRH provoked no behavioural changes in contrast to a distinct circling response to similarly injected DA. Either i.p. or bilateral intra-accumbens injection of haloperidol or pimozide in low doses effectively blocked the locomotor stimulant action of TRH. These results indicate that the DA system in the nucleus accumbens may be of importance in mediation of the locomotor stimulant action of TRH. Differential affinity of TRH to the two DA systems, the mesolimbic and nigrostriatal DA systems is also suggested. Thyrotropin-releasing hormone Locomotor stimulation
Nucleus accumbens Caudate nucleus
1. I n t r o d u c t i o n Thyrotropin-releasing hormone (TRH), one of t h e h y p o t h a l a m i c releasing f a c t o r s , is a p o t e n t releaser o f p r o l a c t i n as well as o f t h y r o t r o p i n . In a d d i t i o n t o t h e s e a c t i o n s , T R H has b e e n r e p o r t e d to p o t e n t i a t e an excit a t o r y a c t i o n o f L - D O P A in h y p o p h y s e c t o m i z e d m i c e as well as in n o r m a l m i c e (Plotnik o f f et al., 1 9 7 2 ) . R a d i o i m m u n o l o g i c a l a n d i m m u n o h i s t o c h e m i c a l studies have d e m o n s t r a t e d t h e d i s t r i b u t i o n o f T R H and T R H - c o n taining n e r v e t e r m i n a l s in t h e n u c l e u s a c c u m bens, lateral septal n u c l e u s a n d several m o t o r nuclei o f the brain s t e m a n d spinal c o r d outside t h e h y p o t h a l a m i c area ( W i n o k u r and Utiger, 1 9 7 4 ; H S k f e l t et al., 1 9 7 5 ) . T h e s e findings suggest t h a t , in the brain, T R H m a y p l a y p h y s i o l o g i c a l roles o t h e r t h a n t h a t o f a releasing h o r m o n e . D o p a m i n e (DA) is localised t o t w o specific a n a t o m i c a l s y s t e m s , t h e striatal p a t h w a y
Dopamlne
Circling
originating in t h e A8 and A9 g r o u p s o f cell b o d i e s in t h e s u b s t a n t i a nigra, and the m e s o limbic p a t h w a y w i t h cell b o d i e s o f its origin in t h e A 1 0 g r o u p l o c a t e d m e d i a l t o the A9 g r o u p in t h e m i d b r a i n and w i t h its nerve terminals in t h e nucleus a c c u m b e n s and t u b e r c u l u m o l f a c t o r i u m ( U n g e r s t e d t , 1971). T h e r e are m a n y r e p o r t s suggesting t h a t s t e r e o t y p y and l o c o m o t o r s t i m u l a t i o n i n d u c e d b y certain drugs are m e d i a t e d via the nigrostriatal D A s y s t e m and t h e m e s o l i m b i c D A s y s t e m , r e s p e c t i v e l y (Ernst and Smelik, 1966; A s h e r and A g h a j a n i a n , 1 9 7 4 ; K e l l y et al., 1 9 7 5 ) . An i m p o r t a n t role o f t h e m e s o l i m b i c D A s y s t e m in l o c o m o t o r a c t i v i t y has b e e n f u r t h e r evidenced by the observation that locomotor s t i m u l a t i o n is p r o d u c e d b y i n j e c t i o n of D A into t h e nucleus a c c u m b e n s in t h e rat pretreated with a monoamine oxidase inhibitor ( P i j n e n b u r g and Van R o s s u m , 1973; Pijnenb u r g et al., 1 9 7 5 ) . We h a v e o b s e r v e d m a r k e d l o c o m o t o r s t i m u l a t i o n also r e p o r t e d b y G o u -
144 jet et al. (1975), following the i.p. injection of T R H in rats. Until now little a t t e nt i on has been paid to these different DA systems with regard to this action of TRH. In the present study, an a t t e m p t to det er m i ne the site of the l o c o m o t o r stimulant action of T R H has been made by comparing the behavioural effect of T R H and DA injected into the nucleus accumbens or caudate nucleus of rats. Modification of the T R H effect by some DA r e c e pt or antagonists was also investigated.
M. MIYAMOTO, Y. NAGAWA counts of less than 50 during a 10-min period, the l o c o m o t o r activity was measured for 60 min following the drug injection. General behaviour was simultaneously observed. In anot her experiment, each rat was placed in a cylindrical cage measuring 60 cm in diameter and 40 cm in height, in which the number of circlings was c o u n t e d continuously by visual observation for 60 min following unilateral injection of the drug into the caudate nucleus.
2.3. Drugs and drug injection 2. Materials and m e t h o d s
2.1. Animals and surgery Male Sprague--Dawley (JCL : SD) rats weighing 2 0 0 - - 3 0 0 g, 6--8 weeks old, were used. In preparing the rats for intracerebral injection of the drug, under sodium pentobarbital anesthesia (40 mg/kg i.p.) stainless steel cannulae with an o u t er and inner diameter of 0.8 and 0.5 mm respectively were implanted bilaterally into th e nucleus accumbens or caudate nucleus. Coordinates for the nucleus accumbens were A 9.4, L 1.6, H 1.0; for the head o f the caudate nucleus A 8.2, L 2.8, H 1.0, according to the De G r o o t atlas (De G r o o t , 1959). Cannulae were fixed ont o the skull with acrylic dental cement. The rats were used for experiments at intervals of at least 5 days after a 7-day postoperative period.
2.2. Measurement o f locomotor activity and circling behaviour Each rat was placed in a perspex cage (26 X 42 X 15 cm) m o u n t e d on t op of a Varimex m e t e r (Columbus I ns t r um ent Co.), which was tuned to a sensitivity 20 pA in order to c o u n t mainly large mo v em e nt s , and l o c o m o t o r counts were recorded on a printer. All experiments were p e r f o r m e d be t w een 8:00 a.m. and 6:00 p.m. at an ambient t e m p e r a t u r e of 22 + 2°C. After the rat was well enough accust o m e d to the activity cage to show l o c o m o t o r
The following drugs were used: T RH (L-pyroglutamyl-L-histidyl-L-prolineamide L-tartrate m o n o h y d r a t e , c r y s t a l B described by Hatanaka et al., 1976: Takeda); dopamine h y d r o c h l o r i d e ( T o k y o Kasei); haloperidol (Serenase ®, Dainippon); pimozide (Orap ®, Fujisawa); p h e n o x y b e n z a m i n e h y d r o c h l o r i d e ( T o k y o Kasei). Pimozide was suspended in a 5% gum arabic solution and all ot her drugs were dissolved in physiological saline. Saline was used as a control. The doses q u o t e d refer to amounts of the salts used. Intracerebral injection of drug was made bilaterally or unilaterally by means of a microsyringe with a needle 0.4 mm out er and 0.2 mm inner diameter which e x t e n d e d into the brain tissues 1.5 mm below the tip of the p e r m a n e n t l y implanted cannula. The injection volume was 1 or 2 pl on each side. The volume for i.p. injection was 0.2 ml/100 g b.wt.
2.4. Statistics Statistical comparison between different treatments was made by means of Student's t-test and the Mann--Whitney's U-test. Twotailed probability values are given.
2.5. Histology At the end of the experiments the rats were perfused first with 30 ml of saline and t hen with 50 ml of a 10% formalin solution through t he left cardiac ventricle under
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH A 9.4
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Fig. 1. Diagrams representing the location of the cannulae tips for injections into the nucleus accumbens (o) or the caudate nucleus (T) according to the De Groot atlas (1959). Only unilateral sites are shown here from histological data obtained from 30 rats, although bilateral sites were examined. Abbreviations: ACB: Nucleus accumbens septi; CC: Corpus callosum; CPU: Nucleus caudatus Putamen. s o d i u m p e n t o b a r b i t a l anesthesia. The brains were r e m o v e d and k e p t in a 10% f o r m a l i n solution for at least 7 days and t h e n s e c t i o n e d t o verify the site o f injection. Position o f the tips of t h e n e e d l e tracks are m a r k e d on a frontal diagram o f the rat brain (fig. 1).
3. Results
3.1. L o c o m o t o r stimulation and other behavioural changes induced by T R H The c o n t r o l rats injected with saline i.p. or i n t r a c e r e b r a l l y o n l y s h o w e d a mild l o c o m o t o r h y p e r a c t i v i t y d u r i n g the initial 10-rain p e r i o d , p r o b a b l y due t o s t i m u l a t i o n b y t h e i n j e c t i o n p r o c e d u r e b u t t h e r e a f t e r s p e n t m o s t o f the
remaining observation period in a resting state. Table I shows a c o m p a r i s o n o f behavioural changes i n d u c e d by a p p r o p r i a t e doses o f T R H and DA injected i.p. o r intracerebrally. The i.p. injection o f T R H in a dose o f 2.5 m g / k g p r o d u c e d n e i t h e r a significant increase in l o c o m o t o r activity n o r o t h e r behavioural changes. T h e rats receiving a 5 m g / k g d o s e s h o w e d significant h y p e r a c t i v i t y consisting o f f r e q u e n t a m b u l a t i o n and rearing. G r o o m i n g and p r e e n i n g were also f r e q u e n t l y observed during pauses b e t w e e n l o c o m o t i o n s . Higher doses (10 and 20 m g / k g ) a l s o p r o d u c e d , in a d d i t i o n to these behaviours, mild sniffing and h e a d - m o v e m e n t s such as head-shaking or h e a d - t w i t c h i n g in all animals, and mild tailelevation and self-biting o f forelegs in s o m e animals. The self-biting was, h o w e v e r , n o t
146
M. M I Y A M O T O , Y. N A G A W A
TABLE 1 C o m p a r i s o n o f behavioural changes i n d u c e d by i.p. and intracerebral injection o f T R H and DA in rats, Abbreviations : bi.N.Acc : bilateral Nucleus A c c u m b e n s : bi.Cd.N : bilateral C a u d a t e Nucleus ; LS : l o c o m o t o r s t i m u l a t i o n ; RE: rearing; SN: sniffing; HM: head m o v e m e n t such as shaking or t w i t c h i n g ; GR: g r o o m i n g ; PR: p r e e n i n g ; T E : tail-elevation; SB: self-biting o f forelegs. Degree o f behavioural changes: --: negative; +: mild; ++: m o d e r a t e or m a r k e d . The d o s e for intracerebral i n j e c t i o n is e x p r e s s e d as the a m o u n t i n j e c t e d o n each side. Drug
Route
Dose
TRH
i.p.
TRH DA TRH DA
bi.N.Acc bi.N.Acc bi.Cd.N. bi.Cd.N.
severe enough Of
the
peripheral locomotor preening
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HM
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PR
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SB
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bens, although
to induce bleeding.
behaviours
observed
administration stimulation, were
tion of TRH
Behavioural changes
following
of TRH, by
bilateral
varied
the degree of sniffing and rear-
depending
administration.
the strong
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reproduced
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the
after
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but
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of
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Fig. 2. The e f f e c t o f i.p. injections o f various doses o f T R H u p o n s p o n t a n e o u s m o t o r activity (SMA). Saline was used as a c o n t r o l . Each value is the m e a n activity o f rats and t h e vertical bars r e p r e s e n t the s t a n d a r d errors o f the means. N u m b e r o f rats u s e d is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, ** p < 0.01, c o m p a r e d w i t h saline c o n t r o l ( S t u d e n t ' s t-test).
M E S O L I M B I C SYSTEM A N D L O C O M O T O R S T I M U L A T I O N BY T R H 500
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Fig. 3. The e f f e c t o f bilateral o f T R H or DA into the nucleus a c c u m b e n s u p o n s p o n t a n e o u s m o t o r activity (SMA). Saline was used as a c o n t r o l . Each value is t h e m e a n activity o f rats and the vertical bars r e p r e s e n t t h e standard errors o f t h e means. N u m b e r o f rats used is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, * * p < 0.01, c o m p a r e d w i t h saline c o n t r o l ( S t u d e n t ' s t-test).
hand, sniffing, grooming and preening were p ro d u ced to a slight e x t e n t by bilateral injection o f 50 pg of T R H and DA into the caudate nucleus b u t h e a d - m o v e m e n t only by TRH. L o c o m o t o r stimulation was never produced by either TRH or DA injected via this route. Neither tail-elevation nor self-biting was observed after the intra-accumbens or intracaudate injection of T R H or DA. The l o c o m o t o r stimulation induced by the i.p. injection of TR H in doses of 5--20 mg/kg and the intra-accumbens injection of T R H (5 and 10 pg) and DA (10 and 20 pg) was dosed e p e n d e n t (figs. 2 and 3). This effect appeared at its m a x i m u m during the initial 20- or 30-min period and t h e r e a f t e r declined gradually. With respect to the total l o c o m o t o r counts for 60 min as measured with a Varimex meter, a dose of 20 mg/kg o f T R H injected i.p. was roughly equivalent to 10 pg of T RH and 20 pg of DA injected into the nucleus accumbens.
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148
M. M I Y A M O T O , Y. N A G A W A A
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Fig. 5. T h e e f f e c t o f i n t r a - a c c u m b e n s p r e t r e a t m e n t w i t h h a l o p e r i d o l ( H P D ) or p h e n o x y b e n z a m i n e (PBZ) (15 m i n p r i o r t o T R H or D A ) u p o n s p o n t a n e o u s m o t o r activity (SMA) s t i m u l a t i o n i n d u c e d b y T R H (20 m g / k g i.p.) (A) or t h e bilateral i n t r a - a c c u m b e n s i n j e c t i o n o f D A (20 pg) (B). E a c h value is t h e m e a n activity o f rats a n d the vertical bars r e p r e s e n t t h e s t a n d a r d errors o f t h e means. N u m b e r o f rats used is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, ** p < 0.01, c o m p a r e d w i t h T R H or D A c o n t r o l ( S t u d e n t ' s t-test). A
TRH
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Fig. 6. T h e e f f e c t of i.p. p r e t r e a t m e n t w i t h h a l o p e r i d o l ( H D P ) (10 m i n p r i o r t o T R H or D A ) o r p i m o z i d e ( P M Z ) (4 h p r i o r to T R H or D A ) u p o n s p o n t a n e o u s m o t o r activity ( S M A ) s t i m u l a t i o n i n d u c e d b y t h e bilateral intraa c c u m b e n s i n j e c t i o n o f T R H (A) or D A (B). E a c h value is t h e m e a n activity of rats a n d t h e vertical bars r e p r e s e n t t h e s t a n d a r d errors o f t h e means. N u m b e r o f rats used is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, • * p < 0.01, c o m p a r e d w i t h T R H or D A c o n t r o l ( S t u d e n t ' s t-test).
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH Unilateral injection of as much as 50 pg of T R H into the caudate nucleus produced no remarkable behavioural change, while similar injection of the same dose of DA caused mild b u t distinct circling behaviour contralateral to the injected side (fig. 4). 3.2. Effect o f DA and noradrenergic (NA) receptor antagonists on TRH-induced locomotor stimulation The l o c o m o t o r hyperactivity induced by the i.p. injection of TRH in a dose of 20 mg/kg was significantly suppressed for the initial 30- or 40-min by pretreatment with bilateral intra-accumbens injection of the DA receptor antagonist, haloperidol (2.5 and 5 pg). Haloperidol (5 pg) also blocked the locom o t o r stimulation induced by bilateral intraaccumbens injection of 20 pg of DA, for the initial 20-min period. However, pretreatment with similar intracerebral injection of a NA receptor antagonist, phenoxybenzamine (10 pg), had no significant effect on the locom o t o r stimulant action of TRH and DA (fig. 5). When the animals were pretreated with haloperidol (0.2 and 1 mg/kg i.p.) or pimozide (0.5 mg/kg i.p.); b o t h drugs markedly suppressed the l o c o m o t o r stimulation induced by the intra-accumbens injection of T R H or DA (fig. 6). 4. Discussion
The most significant sites of the mesolimbic DA system are the nucleus accumbens and tuberculum olfactorium. As mentioned in the Introduction, the DA system of the nucleus accumbens is o f great importance in the mediation of l o c o m o t o r activity. T R H has been already reported by several investigators to produce various types of behaviours. In this work, we noted that T R H injected i.p. caused marked l o c o m o t o r hyperactivity accompanied by rearing, sniffing, grooming, preening and other behaviours in rats. The effect is probably mediated by an action on the mesolimbic DA system. This assumption
149
is supported b y the fact that of the behavioural changes obtained b y the peripheral administration of TRH, l o c o m o t o r hyperactivity, rearing, sniffing, grooming and preening were reproduced by bilateral intra-accumbens injection of TRH and DA. On the other hand, no l o c o m o t o r hyperactivity was caused by bilateral injection of T R H into the caudate nucleus, where the nigrostriatal pathway terminates, although mild sniffing, grooming and preening did occur. Furthermore, the locom o t o r stimulation induced by T R H injected peripherally or into the nucleus accumbens was markedly blocked by the DA receptor antagonists haloperidol and pimozide injected either peripherally or into the nucleus accumbens. These findings strongly suggest that the peripherally administered T R H reaches the nucleus accumbens and exerts its l o c o m o t o r stimulant action, although an action on other levels of brain may be partly involved with the achievement of l o c o m o t o r hyperactivity. On a molar basis, TRH was about 5 times as active as DA in the l o c o m o t o r stimulation induced by intra-accumbens injection, suggesting that TRH may act in some other way on the DA system than as a direct receptor stimulant. In this respect, Cohn et al. (1975), using behavioural studies and Green et al. (1976), using biochemical studies have postulated that T R H acts as an indirect dopamine releaser. A TRH-mediated increase in sensitivity of the DA receptor at postsynaptic sites was also suggested by Plotnikoff et al. (1975). Our present results could be explained by either of these mechanisms. Antagonism of the depressant effects of chlorpromazine in various animals (Kruse, 1976) and o f prochlorperazine-induced catalepsy in rats (Goujet et al., 1975) by TRH appear to be due to its dopaminergic stimulation, as seen in the interaction of amphetamine with neuroleptics. Although the main catecholamine in the nucleus accumbens is DA, the presence of NA in this nucleus has also been described (Koob et al., 1974). However, the weak l o c o m o t o r stimulation induced by intra-accumbens injec-
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH f o u n d an i m p r o v e m e n t o f p s y c h i c s y m p t o m s , especially facial expression, r a p p o r t and p s y c h o m o t o r activity, w h i c h are resistant to t h e r a p y w i t h n e u r o l e p t i c s alone, b y t r e a t m e n t with a c o m b i n a t i o n o f T R H in daily oral doses as l o w as 4 m g and n e u r o l e p t i c s in s c h i z o p h r e n i c patients. Similar m o o d - e l e v a tion or i m p r o v e m e n t in " i n s t i n c t i v e " behavi o u r b y T R H a d m i n i s t r a t i o n in depressive p a t i e n t s has been also described b y Wilson et al. ( 1 9 7 3 ) and Itil et al. (1975). This p s y c h o t r o p i c p r o p e r t y o f T R H appears to be m o r e important p h y s i o l o g i c a l l y t h a n useful in p s y c h o t h e r a p y . I n view o f the d i s t r i b u t i o n o f T R H in brain, o u r p r e s e n t s t u d y suggests t h a t T R H m a y p l a y a role as m o d u l a t o r o f D A a c t i v i t y in the m e s o l i m b i c s y s t e m t o c o n t r o l t h e e m o t i o n a l and m e n t a l f u n c t i o n s .
References Asher, I.M. and G.K. Aghajanian, 1974, 6-Hydroxydopamine lesions of olfactory tubercles and caudate nuclei: effect on amphetamine-induced stereotyped behaviour in rats, Brain Res. 82, 1. Benkert, O., A. Gordon and D. Martschke, 1974, The comparison of thyrotropin-releasing hormone, luteinising hormone-releasing hormone and placebo in depressive patients using a double-blind cross-over technique, Psychopharmacologia {Berlin) 40,191. Cohn, M.L., M. Cohn and F.H. Taylor, 1975, Thyrotropin releasing factor (TRF) regulation of rotation in the non-lesioned rat, Brain Res. 96, 134. Costall, B. and R.J. Naylor, 1973, The role of telencephalic dopaminergic systems in the mediation of apomorphine-stereotyped behaviour, European J. Pharmacol. 24, 8. Davis, K.L., L.E. Hollister and P.A. Berger, 1975, Thyrotropin-releasing hormone in schizophrenia, Amer. J, Psychiatry 132, 751. De Groot, J., 1959, The rat forebrain in stereotaxic coordinates, Verh. K. Acad. Wet. 52, 1. Ernst, A.M. and P.G. Smelik, 1966, Site of action of dopamine and apomorphine on compulsive gnawing behaviour in rats, Experientia 22, 837. Goujet, M.A., P. Simon, R. Chermat and J.R. Boissier, 1975, Profile de la T.R.H. en psychopharmacologie exp~rimentale, Psychopharmacologia (Berlin) 45, 87. Green, A.R., D.J. Heal, D.J. Grahame-Smith and P.H. Kelly, 1976, The contrasting action of TRH and
151
cycloheximide in altering the effects of centrally acting drugs: evidence for the non-involvement of dopamine sensitive adenylate cyclase, Neuropharmacology 15,591. Hatanaka, C., H. Nakamachi, M. Kaneko, H. Yonezawa, N. Toukai and M. Fujino, 1976, Synthesis and polymorphs of TRH L-tartrate (TRH-T), J. Takeda Res. Lab. 35, 16. HSkfelt, T., K. Fuxe, O. Johansson, S. Jeffcoate and N. White, 1975, Distribution of thyrotropin-releasing hormone (TRH) in the central nervous system as revealed with immunohistochemistry, European J. Pharmacol. 34, 389. Hollister, L.E., P. Berger, F.L. Ogle, R.C. Arnold and A. Johnson, 1974, Protirelin (TRH) in depression, Arch. Gen. Psychiatry 31,468. Inagaga, K., 1976, "Behavioral effects of thyrotropinreleasing hormone in schizophrenia" presented at the 10th Congress of Collegium Internationale Neuro-psychopharmacologium, Quebec, Canada, July 4--9. Itil, T.M.~ C.D. Patterson, N. Polvan, A. Bigelow and B. Bergery, 1975, Clinical and CNS effects of oral and i.v. thyrotropin-releasing hormone in depressed patients, Dis. Nerv. Syst. 36,529. Iversen, L.L., 1975, Dopamine receptor in the brain: a dopamine-sensitive adenylcyclase models synaptic receptors, illuminating drug action, Science 188, 1084. Kastin, A.J., R.H. Ehrensing, D.S. Schalch and M.S. Anderson, 1972, Improvement in mental depression with decreased thyrotropin response after administration of thyrotropin-releasing hormone, Lancet II, 740. Keller, H.H., G. Bartholini and A. Pletscher, 1974, Enhancement of cerebral noradrenaline turnover by thyrotropin-releasing hormone, Nature {London) 248, 528. Kelly, P.H., P.W. Seviour and S.D. Iversen, 1975, Amphetamine and apomorphine response in the rat following 6-OHDA lesions of the nucleus accumbens septi and corpus striatum, Brain Res. 94, 507. Koob, G.F., G.J. Balcom and J.L. Meyerhoff, 1974, Changes in dopamine and norepinephrine in the nucleus accumbens septi, olfactory tubercle and corpus striatum following lesions in ventral tegmental area in the rat, Federation Proc. 33, 246. Kruse, H., 1976, Thyrotropin releasing hormone (TRH): restoration of oxotremorine tremor in mice, Naunyn-Schmiedeb. Arch. Pharmacol. 294, 39. Lotti, V.J., 1971, Action of various centrally acting agents in mice with unilateral caudate lesions, Life Sci. 10,781. Pijnenburg, A.J.J. and J.M. Van Rossum, 1973, Stimulation of locomotor activity following injection
152 of dopamine into the nucleus accumbens, J. Pharm. Pharmacol. 25, 1003. Pijnenburg, A.J.J., W.M.M. Honig and J.M. Van Rossum, 1975, Effect of antagonists upon locomotor stimulation induced by injection of dopamine and noradrenaline into the nucleus accumbens of nialamide-pretreated rats, Psychopharmacologia (Berlin) 41, 175. Plotnikoff, N.P., A.J. Prange Jr., G.R. Breese, M.S. Anderson and I.C. Wilson, 1972, Thyrotropin releasing hormone: enhancement of DOPA activity by a hypothalamic hormone, Science 178, 417. Plotnikoff, N.P., G.R. Breese and A.J. Prange Jr., 1975, Thyrotropin-releasing hormone (TRH): DOPA potentiation and biogenic amine studies, Pharmacol. Biochem. Behav. 3, 665. Powell, E.W. and R.B. Leman, 1976, Connections of the nucleus accumbens, Brain Res. 105, 389. Prange Jr., A.J., I.C. Wilson, P.P. Lala, L.H. Alltop and G.R. Breese, 1972, Effect of thyrotropin-releasing hormone in depression, Lancet II, 999.
M. MIYAMOTO, Y. NAGAWA Razzak, A., M. Fujiwara and S. Ueki, 1975, Automutilation induced by clonidine in mice, European J. Pharmacol. 30, 356. Stevens, J., 1973, An anatomy of schizophrenia? Arch. Gen. Psychiatry 29, 177. Ungerstedt, U., 1971, Stereotaxic mapping of the monoamine pathways in the rat brain, Acta Physiol. Scand., SuppL. 367, 1. Van Rossum, J.M., P.A. Janssen, J.R. Boissier, L. Juiou, D.M. Loew, I. M¢ller Nielsen, I. Munkvad, A. Randrup, G. Stille and D.H. Tedeschi, 1970, Pharmacology, in: Modern Problems of Pharmacopsychiatry, The Neuroleptics, eds. D.P. Bobon, P.A.J. Janssen and J. Bobon (Karger, Basel) p. 23. Wilson, I.C., P.P. Lala and A.J. Prange, Jr., 1973, Thyrotropin-releasing hormone in schizophrenia, Lancet II, 43. Winokur, A. and R.D. Utiger, 1974, Thyrotropin-releasing hormone: regional distribution in rat brain, Science 185, 265.
143
© Elsevier/North-Holland Biomedical Press
MESOLIMBIC INVOLVEMENT IN THE LOCOMOTOR STIMULANT ACTION OF T H Y R O T R O P I N - R E L E A S I N G H O R M O N E ( T R H ) IN R A T S MASAOMI MIYAMOTO and YUJI NAGAWA Medicinal Laboratories, Central Research Division, Takeda Chemical Industries, Ltd., Osaka 532, Japan
Received 16 November 1976, revised MS received 18 February 1977, accepted 24 March 1977
M. MIYAMOTO and Y. NAGAWA, Mesolimbic involvement in the locomotor stimulant action o f thyrotropinreleasing hormone (TRH) in rats, European J. Pharmacol, 44 (1977) 143--152. Thyrotropin-releasing hormone (TRH) injected i.p. in doses of 5 mg/kg and higher had a strong locomotor stimulant action with development of frequent rearing, mild sniffing, grooming, preening and other excitatory behaviours. The locomotor stimulation was also produced by bilateral injection of TRH and dopamine (DA) into the nucleus accumbens but not by bilateral injection of these substances into the caudate nucleus. Unilateral intracaudate injection of TRH provoked no behavioural changes in contrast to a distinct circling response to similarly injected DA. Either i.p. or bilateral intra-accumbens injection of haloperidol or pimozide in low doses effectively blocked the locomotor stimulant action of TRH. These results indicate that the DA system in the nucleus accumbens may be of importance in mediation of the locomotor stimulant action of TRH. Differential affinity of TRH to the two DA systems, the mesolimbic and nigrostriatal DA systems is also suggested. Thyrotropin-releasing hormone Locomotor stimulation
Nucleus accumbens Caudate nucleus
1. I n t r o d u c t i o n Thyrotropin-releasing hormone (TRH), one of t h e h y p o t h a l a m i c releasing f a c t o r s , is a p o t e n t releaser o f p r o l a c t i n as well as o f t h y r o t r o p i n . In a d d i t i o n t o t h e s e a c t i o n s , T R H has b e e n r e p o r t e d to p o t e n t i a t e an excit a t o r y a c t i o n o f L - D O P A in h y p o p h y s e c t o m i z e d m i c e as well as in n o r m a l m i c e (Plotnik o f f et al., 1 9 7 2 ) . R a d i o i m m u n o l o g i c a l a n d i m m u n o h i s t o c h e m i c a l studies have d e m o n s t r a t e d t h e d i s t r i b u t i o n o f T R H and T R H - c o n taining n e r v e t e r m i n a l s in t h e n u c l e u s a c c u m bens, lateral septal n u c l e u s a n d several m o t o r nuclei o f the brain s t e m a n d spinal c o r d outside t h e h y p o t h a l a m i c area ( W i n o k u r and Utiger, 1 9 7 4 ; H S k f e l t et al., 1 9 7 5 ) . T h e s e findings suggest t h a t , in the brain, T R H m a y p l a y p h y s i o l o g i c a l roles o t h e r t h a n t h a t o f a releasing h o r m o n e . D o p a m i n e (DA) is localised t o t w o specific a n a t o m i c a l s y s t e m s , t h e striatal p a t h w a y
Dopamlne
Circling
originating in t h e A8 and A9 g r o u p s o f cell b o d i e s in t h e s u b s t a n t i a nigra, and the m e s o limbic p a t h w a y w i t h cell b o d i e s o f its origin in t h e A 1 0 g r o u p l o c a t e d m e d i a l t o the A9 g r o u p in t h e m i d b r a i n and w i t h its nerve terminals in t h e nucleus a c c u m b e n s and t u b e r c u l u m o l f a c t o r i u m ( U n g e r s t e d t , 1971). T h e r e are m a n y r e p o r t s suggesting t h a t s t e r e o t y p y and l o c o m o t o r s t i m u l a t i o n i n d u c e d b y certain drugs are m e d i a t e d via the nigrostriatal D A s y s t e m and t h e m e s o l i m b i c D A s y s t e m , r e s p e c t i v e l y (Ernst and Smelik, 1966; A s h e r and A g h a j a n i a n , 1 9 7 4 ; K e l l y et al., 1 9 7 5 ) . An i m p o r t a n t role o f t h e m e s o l i m b i c D A s y s t e m in l o c o m o t o r a c t i v i t y has b e e n f u r t h e r evidenced by the observation that locomotor s t i m u l a t i o n is p r o d u c e d b y i n j e c t i o n of D A into t h e nucleus a c c u m b e n s in t h e rat pretreated with a monoamine oxidase inhibitor ( P i j n e n b u r g and Van R o s s u m , 1973; Pijnenb u r g et al., 1 9 7 5 ) . We h a v e o b s e r v e d m a r k e d l o c o m o t o r s t i m u l a t i o n also r e p o r t e d b y G o u -
144 jet et al. (1975), following the i.p. injection of T R H in rats. Until now little a t t e nt i on has been paid to these different DA systems with regard to this action of TRH. In the present study, an a t t e m p t to det er m i ne the site of the l o c o m o t o r stimulant action of T R H has been made by comparing the behavioural effect of T R H and DA injected into the nucleus accumbens or caudate nucleus of rats. Modification of the T R H effect by some DA r e c e pt or antagonists was also investigated.
M. MIYAMOTO, Y. NAGAWA counts of less than 50 during a 10-min period, the l o c o m o t o r activity was measured for 60 min following the drug injection. General behaviour was simultaneously observed. In anot her experiment, each rat was placed in a cylindrical cage measuring 60 cm in diameter and 40 cm in height, in which the number of circlings was c o u n t e d continuously by visual observation for 60 min following unilateral injection of the drug into the caudate nucleus.
2.3. Drugs and drug injection 2. Materials and m e t h o d s
2.1. Animals and surgery Male Sprague--Dawley (JCL : SD) rats weighing 2 0 0 - - 3 0 0 g, 6--8 weeks old, were used. In preparing the rats for intracerebral injection of the drug, under sodium pentobarbital anesthesia (40 mg/kg i.p.) stainless steel cannulae with an o u t er and inner diameter of 0.8 and 0.5 mm respectively were implanted bilaterally into th e nucleus accumbens or caudate nucleus. Coordinates for the nucleus accumbens were A 9.4, L 1.6, H 1.0; for the head o f the caudate nucleus A 8.2, L 2.8, H 1.0, according to the De G r o o t atlas (De G r o o t , 1959). Cannulae were fixed ont o the skull with acrylic dental cement. The rats were used for experiments at intervals of at least 5 days after a 7-day postoperative period.
2.2. Measurement o f locomotor activity and circling behaviour Each rat was placed in a perspex cage (26 X 42 X 15 cm) m o u n t e d on t op of a Varimex m e t e r (Columbus I ns t r um ent Co.), which was tuned to a sensitivity 20 pA in order to c o u n t mainly large mo v em e nt s , and l o c o m o t o r counts were recorded on a printer. All experiments were p e r f o r m e d be t w een 8:00 a.m. and 6:00 p.m. at an ambient t e m p e r a t u r e of 22 + 2°C. After the rat was well enough accust o m e d to the activity cage to show l o c o m o t o r
The following drugs were used: T RH (L-pyroglutamyl-L-histidyl-L-prolineamide L-tartrate m o n o h y d r a t e , c r y s t a l B described by Hatanaka et al., 1976: Takeda); dopamine h y d r o c h l o r i d e ( T o k y o Kasei); haloperidol (Serenase ®, Dainippon); pimozide (Orap ®, Fujisawa); p h e n o x y b e n z a m i n e h y d r o c h l o r i d e ( T o k y o Kasei). Pimozide was suspended in a 5% gum arabic solution and all ot her drugs were dissolved in physiological saline. Saline was used as a control. The doses q u o t e d refer to amounts of the salts used. Intracerebral injection of drug was made bilaterally or unilaterally by means of a microsyringe with a needle 0.4 mm out er and 0.2 mm inner diameter which e x t e n d e d into the brain tissues 1.5 mm below the tip of the p e r m a n e n t l y implanted cannula. The injection volume was 1 or 2 pl on each side. The volume for i.p. injection was 0.2 ml/100 g b.wt.
2.4. Statistics Statistical comparison between different treatments was made by means of Student's t-test and the Mann--Whitney's U-test. Twotailed probability values are given.
2.5. Histology At the end of the experiments the rats were perfused first with 30 ml of saline and t hen with 50 ml of a 10% formalin solution through t he left cardiac ventricle under
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH A 9.4
A 9.8
145
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~
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-
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'
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Fig. 1. Diagrams representing the location of the cannulae tips for injections into the nucleus accumbens (o) or the caudate nucleus (T) according to the De Groot atlas (1959). Only unilateral sites are shown here from histological data obtained from 30 rats, although bilateral sites were examined. Abbreviations: ACB: Nucleus accumbens septi; CC: Corpus callosum; CPU: Nucleus caudatus Putamen. s o d i u m p e n t o b a r b i t a l anesthesia. The brains were r e m o v e d and k e p t in a 10% f o r m a l i n solution for at least 7 days and t h e n s e c t i o n e d t o verify the site o f injection. Position o f the tips of t h e n e e d l e tracks are m a r k e d on a frontal diagram o f the rat brain (fig. 1).
3. Results
3.1. L o c o m o t o r stimulation and other behavioural changes induced by T R H The c o n t r o l rats injected with saline i.p. or i n t r a c e r e b r a l l y o n l y s h o w e d a mild l o c o m o t o r h y p e r a c t i v i t y d u r i n g the initial 10-rain p e r i o d , p r o b a b l y due t o s t i m u l a t i o n b y t h e i n j e c t i o n p r o c e d u r e b u t t h e r e a f t e r s p e n t m o s t o f the
remaining observation period in a resting state. Table I shows a c o m p a r i s o n o f behavioural changes i n d u c e d by a p p r o p r i a t e doses o f T R H and DA injected i.p. o r intracerebrally. The i.p. injection o f T R H in a dose o f 2.5 m g / k g p r o d u c e d n e i t h e r a significant increase in l o c o m o t o r activity n o r o t h e r behavioural changes. T h e rats receiving a 5 m g / k g d o s e s h o w e d significant h y p e r a c t i v i t y consisting o f f r e q u e n t a m b u l a t i o n and rearing. G r o o m i n g and p r e e n i n g were also f r e q u e n t l y observed during pauses b e t w e e n l o c o m o t i o n s . Higher doses (10 and 20 m g / k g ) a l s o p r o d u c e d , in a d d i t i o n to these behaviours, mild sniffing and h e a d - m o v e m e n t s such as head-shaking or h e a d - t w i t c h i n g in all animals, and mild tailelevation and self-biting o f forelegs in s o m e animals. The self-biting was, h o w e v e r , n o t
146
M. M I Y A M O T O , Y. N A G A W A
TABLE 1 C o m p a r i s o n o f behavioural changes i n d u c e d by i.p. and intracerebral injection o f T R H and DA in rats, Abbreviations : bi.N.Acc : bilateral Nucleus A c c u m b e n s : bi.Cd.N : bilateral C a u d a t e Nucleus ; LS : l o c o m o t o r s t i m u l a t i o n ; RE: rearing; SN: sniffing; HM: head m o v e m e n t such as shaking or t w i t c h i n g ; GR: g r o o m i n g ; PR: p r e e n i n g ; T E : tail-elevation; SB: self-biting o f forelegs. Degree o f behavioural changes: --: negative; +: mild; ++: m o d e r a t e or m a r k e d . The d o s e for intracerebral i n j e c t i o n is e x p r e s s e d as the a m o u n t i n j e c t e d o n each side. Drug
Route
Dose
TRH
i.p.
TRH DA TRH DA
bi.N.Acc bi.N.Acc bi.Cd.N. bi.Cd.N.
severe enough Of
the
peripheral locomotor preening
5 mg/kg 20 m g / k g 10 pg 20 pg 50 ~g 50//g
LS
RE
SN
HM
GR
PR
TE
SB
+ ++ ++ ++ ---
+ ++ + + ---
-+ + ++ + +
-++ + -+ --
++ ++ ++ ++ + +
++ ++ ++ ++ + +
-+ -----
-+ -----
bens, although
to induce bleeding.
behaviours
observed
administration stimulation, were
tion of TRH
Behavioural changes
following
of TRH, by
bilateral
varied
the degree of sniffing and rear-
depending
administration.
the strong
sniffing, grooming
reproduced
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the
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the
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also
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of
TRH
upon
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The head-movement the
but
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of TRH
intra-accumbens of DA.
of
observed
administration
by
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On
the
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(6) 500
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Fig. 2. The e f f e c t o f i.p. injections o f various doses o f T R H u p o n s p o n t a n e o u s m o t o r activity (SMA). Saline was used as a c o n t r o l . Each value is the m e a n activity o f rats and t h e vertical bars r e p r e s e n t the s t a n d a r d errors o f the means. N u m b e r o f rats u s e d is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, ** p < 0.01, c o m p a r e d w i t h saline c o n t r o l ( S t u d e n t ' s t-test).
M E S O L I M B I C SYSTEM A N D L O C O M O T O R S T I M U L A T I O N BY T R H 500
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Fig. 3. The e f f e c t o f bilateral o f T R H or DA into the nucleus a c c u m b e n s u p o n s p o n t a n e o u s m o t o r activity (SMA). Saline was used as a c o n t r o l . Each value is t h e m e a n activity o f rats and the vertical bars r e p r e s e n t t h e standard errors o f t h e means. N u m b e r o f rats used is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, * * p < 0.01, c o m p a r e d w i t h saline c o n t r o l ( S t u d e n t ' s t-test).
hand, sniffing, grooming and preening were p ro d u ced to a slight e x t e n t by bilateral injection o f 50 pg of T R H and DA into the caudate nucleus b u t h e a d - m o v e m e n t only by TRH. L o c o m o t o r stimulation was never produced by either TRH or DA injected via this route. Neither tail-elevation nor self-biting was observed after the intra-accumbens or intracaudate injection of T R H or DA. The l o c o m o t o r stimulation induced by the i.p. injection of TR H in doses of 5--20 mg/kg and the intra-accumbens injection of T R H (5 and 10 pg) and DA (10 and 20 pg) was dosed e p e n d e n t (figs. 2 and 3). This effect appeared at its m a x i m u m during the initial 20- or 30-min period and t h e r e a f t e r declined gradually. With respect to the total l o c o m o t o r counts for 60 min as measured with a Varimex meter, a dose of 20 mg/kg o f T R H injected i.p. was roughly equivalent to 10 pg of T RH and 20 pg of DA injected into the nucleus accumbens.
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148
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Fig. 6. T h e e f f e c t of i.p. p r e t r e a t m e n t w i t h h a l o p e r i d o l ( H D P ) (10 m i n p r i o r t o T R H or D A ) o r p i m o z i d e ( P M Z ) (4 h p r i o r to T R H or D A ) u p o n s p o n t a n e o u s m o t o r activity ( S M A ) s t i m u l a t i o n i n d u c e d b y t h e bilateral intraa c c u m b e n s i n j e c t i o n o f T R H (A) or D A (B). E a c h value is t h e m e a n activity of rats a n d t h e vertical bars r e p r e s e n t t h e s t a n d a r d errors o f t h e means. N u m b e r o f rats used is s h o w n in p a r e n t h e s e s . Level o f significance: * p < 0.05, • * p < 0.01, c o m p a r e d w i t h T R H or D A c o n t r o l ( S t u d e n t ' s t-test).
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH Unilateral injection of as much as 50 pg of T R H into the caudate nucleus produced no remarkable behavioural change, while similar injection of the same dose of DA caused mild b u t distinct circling behaviour contralateral to the injected side (fig. 4). 3.2. Effect o f DA and noradrenergic (NA) receptor antagonists on TRH-induced locomotor stimulation The l o c o m o t o r hyperactivity induced by the i.p. injection of TRH in a dose of 20 mg/kg was significantly suppressed for the initial 30- or 40-min by pretreatment with bilateral intra-accumbens injection of the DA receptor antagonist, haloperidol (2.5 and 5 pg). Haloperidol (5 pg) also blocked the locom o t o r stimulation induced by bilateral intraaccumbens injection of 20 pg of DA, for the initial 20-min period. However, pretreatment with similar intracerebral injection of a NA receptor antagonist, phenoxybenzamine (10 pg), had no significant effect on the locom o t o r stimulant action of TRH and DA (fig. 5). When the animals were pretreated with haloperidol (0.2 and 1 mg/kg i.p.) or pimozide (0.5 mg/kg i.p.); b o t h drugs markedly suppressed the l o c o m o t o r stimulation induced by the intra-accumbens injection of T R H or DA (fig. 6). 4. Discussion
The most significant sites of the mesolimbic DA system are the nucleus accumbens and tuberculum olfactorium. As mentioned in the Introduction, the DA system of the nucleus accumbens is o f great importance in the mediation of l o c o m o t o r activity. T R H has been already reported by several investigators to produce various types of behaviours. In this work, we noted that T R H injected i.p. caused marked l o c o m o t o r hyperactivity accompanied by rearing, sniffing, grooming, preening and other behaviours in rats. The effect is probably mediated by an action on the mesolimbic DA system. This assumption
149
is supported b y the fact that of the behavioural changes obtained b y the peripheral administration of TRH, l o c o m o t o r hyperactivity, rearing, sniffing, grooming and preening were reproduced by bilateral intra-accumbens injection of TRH and DA. On the other hand, no l o c o m o t o r hyperactivity was caused by bilateral injection of T R H into the caudate nucleus, where the nigrostriatal pathway terminates, although mild sniffing, grooming and preening did occur. Furthermore, the locom o t o r stimulation induced by T R H injected peripherally or into the nucleus accumbens was markedly blocked by the DA receptor antagonists haloperidol and pimozide injected either peripherally or into the nucleus accumbens. These findings strongly suggest that the peripherally administered T R H reaches the nucleus accumbens and exerts its l o c o m o t o r stimulant action, although an action on other levels of brain may be partly involved with the achievement of l o c o m o t o r hyperactivity. On a molar basis, TRH was about 5 times as active as DA in the l o c o m o t o r stimulation induced by intra-accumbens injection, suggesting that TRH may act in some other way on the DA system than as a direct receptor stimulant. In this respect, Cohn et al. (1975), using behavioural studies and Green et al. (1976), using biochemical studies have postulated that T R H acts as an indirect dopamine releaser. A TRH-mediated increase in sensitivity of the DA receptor at postsynaptic sites was also suggested by Plotnikoff et al. (1975). Our present results could be explained by either of these mechanisms. Antagonism of the depressant effects of chlorpromazine in various animals (Kruse, 1976) and o f prochlorperazine-induced catalepsy in rats (Goujet et al., 1975) by TRH appear to be due to its dopaminergic stimulation, as seen in the interaction of amphetamine with neuroleptics. Although the main catecholamine in the nucleus accumbens is DA, the presence of NA in this nucleus has also been described (Koob et al., 1974). However, the weak l o c o m o t o r stimulation induced by intra-accumbens injec-
MESOLIMBIC SYSTEM AND LOCOMOTOR STIMULATION BY TRH f o u n d an i m p r o v e m e n t o f p s y c h i c s y m p t o m s , especially facial expression, r a p p o r t and p s y c h o m o t o r activity, w h i c h are resistant to t h e r a p y w i t h n e u r o l e p t i c s alone, b y t r e a t m e n t with a c o m b i n a t i o n o f T R H in daily oral doses as l o w as 4 m g and n e u r o l e p t i c s in s c h i z o p h r e n i c patients. Similar m o o d - e l e v a tion or i m p r o v e m e n t in " i n s t i n c t i v e " behavi o u r b y T R H a d m i n i s t r a t i o n in depressive p a t i e n t s has been also described b y Wilson et al. ( 1 9 7 3 ) and Itil et al. (1975). This p s y c h o t r o p i c p r o p e r t y o f T R H appears to be m o r e important p h y s i o l o g i c a l l y t h a n useful in p s y c h o t h e r a p y . I n view o f the d i s t r i b u t i o n o f T R H in brain, o u r p r e s e n t s t u d y suggests t h a t T R H m a y p l a y a role as m o d u l a t o r o f D A a c t i v i t y in the m e s o l i m b i c s y s t e m t o c o n t r o l t h e e m o t i o n a l and m e n t a l f u n c t i o n s .
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