Evidence that striatal efferents relate to different dopamine receptors

Brain Research, 323 (I t#84)26t)-278 Elsevier

269

BRE 10419

Evidence that Striatal Efferents Relate to Different Dopamine Receptors MARIO HERRERA-MARSCHITZ and URBAN UNGERSTEDT Dept. c{f Pharrnacology, Karolinska institutet. S-104 Ol Stockholm ¢Sweden)

(Accepted April 3rd. 1984) Key words': striatal efferents - - doparnine receptors - - rotation - - apornorphine - - pergolide - 6-hydroxy-dopamine (6-OHDA) --kainic acid

11 was proposed I~ that apomorpbine and the ergot derivative pergolide induce rotation in 6-hydroxy-doparninc (6-OHI)A)-dcncrwired rats by diffcrcnt receptor mechanisms, since these dopamine agonists induce different patterns of rotational bchaviour, have different dose-response curves and are differently inhibited by neuroleptics acting on D I or D2 type receptors. The synaptic continuation of the striatonigral pathway was interrupted in unilaterally 6-OHDA-lesioned rats by adding a kainic acid lesion to the 6-O[-tDAlesioned nigra. This lesion affected apomorphine and pcrgolide rotation differently. After an initial short peak of contralateral rotation, apomorphine induced ipsilatcral rotation that increased with dose. although a final contralateral peak was always maintained. The only change in pcrgolide rotation was a shift of the dose-response curve to the right. In another group ol animals, the continuation of the striatonigral pathway was lesioned by unilateral kainic acid injections without previous 6-OHDA-lesions of the doparnine system. In these animals apornorphin¢ but not pergolide induced rotational behaviour. In contrast, both drugs induced rotation in animals where all striatal efferents were lesioned unilaterally by kainic acid injections into one striaturn, The results suggest that the apomorphine rotation is mainly dependent upon striatonigral pathways while the pergolide rotation is dependent upon other striatal efferent pathways.

INTRODU('TION

lide and bromocriptine induce dose-dependent contralateral rotation in 6 - O H D A - t r e a t e d rats. large dif-

The importance of the striatonigral efferents for the expression of rotational behaviour in rats has

ferences are found when comparing their dose-res-

been well documented a,~",~s. The rotation induced by low doses of apomorphine (0.25 mg/kg) or L-DOPA (50 mg/kg) in unilaterally 6 - O H D A - d e n e r v a t e d animals is markedly reduced by ipsilateral electrocoagulation of the internal capsule or ipsilateral knife-cuts immediately rostral to the substantia nigra ~s. When the electrolytic lesions are positioned at the level of the internal capsule where the fibres of the nigrostriatal system have not entered, the contralateral apomorphine rotation is converted into ipsilateral turning after doses higher than 0.5 mg/kg s.c. >. The same results are produced by kainic acid (KA) lesions of the ipsilateral substantia nigra reticulata of previously 6-OH D A - d e n e r v a t e d rats 4. In recent studiesl-~,e~L we have shown that although both apomorphinc and the ergot derivatives pergo('orrespondence:

ponse curves as well as the patterns of rotation which they elicit. A p o m o r p h i n e induces a two-peak pattern of rotation which never occurs after pergolide nor bromocriptine treatment. We have suggested that the differences between apomorphine and the ergot derivatives may be explained in terms of actions on different dopamine (DA) receptors, since they are differently inhibited by neuroleptics acting on D1 or D2 type of receptors i~,>. In this study we present evidence which suggests that the DA receptors preferentially stimulated by apomorphine or the ergot derivative pergolide may be associated with different striatal efferents, since lesions of the synaptic continuation of striato-nigral pathways qualitatively modify the apomorphine, but not the pergolide response.

M. Herrera-Murschitz. Dept. of Pharmacology, Karolinska Instituter, S-104 0l Stockholm, Sweden.

00n(~-~c193,~4/$(t3.i)0@ 1994 Elscvicr Science Publishers B.V,

270 MATERIALS AND METHODS

Additional KA lesion oJthe ipsilateral substantia nigra reticutata of 6-OHDA-denervated rats. 6 - O H D A -

Male S p r a g u e - D a w l e y rats iweighing 150-170 g at the time of the 6 - O H D A stereotaxic injectionsl were used in all experiments. The animals were housed 4 in each cage with free access to food and water. They were maintained in a t e m p e r a t u r e con-

d e n e r v a t e d rats showing a two-peak patern of rotation when challenged with a p o m o r p h i n e 0,05 mg;k~

trolled environment on a 12 h light/dark cycle when not in experimental sessions.

Surgical p rocedures Unilateral 6-OHDA-denervation.

Rats were anaesthetized using a mixture of air and halothane. and placed in a David Kopf stereotaxic frame. The anaesthesia was m a i n t a i n e d by free breathing into a mask fitted over the nose of the rat, The rat skull was o r i e n t a t e d according to the K6nig and klippel stereotaxic atlas ~7 and 8/~g of 6 - O H D A - H C I (calculated as the base, S I G M A ) in 4 ul of saline (with 0.2% ascorbic acid) was injected into the area ventralis tegmenti which contains the bundle of axons leaving the mesencephalic D A cell bodies. The coordinates were 4.4 mm posterior and 1.2 mm lateral to B r e g m a and 7.8 mm vertical m e a s u r e d from the brain surface. This lesion extensively denervates the D A innervated forebrain areas on the side of the lesion 2s--'s

Unilateral KA lesion of the substantia mgra reticulata of normal rats. Rats weighing 150-170 g received 0.5/~g of K A (Sigmal in 0.5 I~1 into the left substantia nigra (coordinates: 5.0 mm posterior, 2.0 mm lateral to B r e g m a and 7.7 m m verticalL A prophylactic dose of diazepam 10 mg/kg s.c. was given at the time of the stereotaxic intervention in order to reduce seizure damages induced acutely b~ the K A treatment. This K A t r e a t m e n t aimed to destroy the nerve cell bodies within the substantia nigra reticulata that receive the major part of the striatonigral

s.c.:~ were tested with pergolide II.01 mg/kg s.c. and then subjected to an additional stereotaxic procedure. This involved injection ol 1 ug of K A in 1 u ot physiological ringer into the ipsilateral substantia mgra reticu[ata Icoordinates: 5.0 m m posterior, 2.[1 mm lateral to Bregma and 7 7 mm verticali. A prophylactic dose of diazepam ~ ltl mg/kg s,c. ~ was given at the time of the stereolaxic intervention. This intervenuon was aimed to extensively impair the elferent pathway mediating the a p o m o r p h i n e rotation in 6 - O H D A - d e n e r v a t e d rats 1"-1~ All stereotaxic injecnons were made with 5 or 10 ul Hamilton syringes using conically shaped needies with a maximal outer d i a m e t e r of 0.4 ram. Injections were p e r f o r m e d at the rate o t l /A/rain and withdrawn 2 rain following the completion of the respective infusions.

Rotational behaviour Rotational behaviour was recorded using a modified verskm of the original rotometer27. Each 18[V right or left turn was r e c o r d e d with a detector using infrared photocell barriers. The pulses from the detector electronics were fed into a m i c r o c o m p u t e r for intermediate storage and display. A f t e r completion of the experiments, the data were transferred to a host c o m p u t e r for p e r m a n e n t storage on diskettes. The rotational b e h a v i o u r of each individual animal was plotted as the n u m b e r of 360 ° turns/min for the entire duration of the rotational b e h a v i o u r response. Detailed plotting of the rotational behaviour formed the basis for selecting successfully d e n e r v a t e d animals.

projection 7,s.

Unilateral striatal KA lesion. Rats weighing 200-250 g received two stereotaxic injections of 1 ~g of K A in 1 ]~1 into the left striatum (coordinates: 1.0 m m anterior from Bregma, 2.3 m m lateral and 4.5 mm vertical, and 3.8 m m lateral at the Bregma level and 4.5 m m verticall, a prophylactic dose of diazepam was given after the second injection. This K A t r e a t m e n t induces an almost c o m p l e t e loss of nerve cell bodies within the striatum while sparing axons of passage and afferent terminals ~.

Behavioural evaluation off the 6-OHDA lesion." the apomorphine rotation In o r d e r to select the successfully d e n e r v a t e d animals. rats were challenged with a low dose of apomorphine (0.05 mg/kg s.c.~ two weeks after the 6O H D A injection. The a p o m o r p h i n e test was repeated at least two rimes with a o n e - w e e k interval between treatments. Only rats showing a two-peak pattern of rotation were used in the experiments, since in previous studies 2s we have found that this is a high-

271 ly reproducible sign of successful denervation and is only observed in rats with more than 95% striatal DA depletion, Immunohistochemical analysis of the substantia nigra of 6-OHDA-treated rats showing twopeak rotation after incubation with antiserum to tyrosine hydroxylase (TH) reveals an almost total disappearance of TH immunoreactive cell bodies in the

phine (0.5 mg/kg), were selected to be further tested with apomorphine 1.0 mg/kg s.c. After a washing out period of 2 weeks, the same rats were tested with pergolide 0.1 and 0.2 mg/kg s.c. At least one week elapsed between the respective doses.

zona compacta (Herrera-Marschitz et al,, in preparation), The selected rats were further tested with pergolide 0.01 mg/kg s.c.

the left striatum showing ipsilateral rotation after apomorphine 0,5 mg/kg s.c. were selected to be further tested with apomorphine 5.0 mg/kg s.c. After a washing out period of 2 weeks, the same rats were tested with pergolide 0.1 and 0.2 mg/kg s.c. At least one week elapsed between the respective doses.

Behavioural evaluation of the KA lesions Rats with unilateral KA lesions of the substantia nigra reticulam. Since the KA lesion was aimed to impair the synaptic continuation of the striatonigral efferents, successfully lesioned animals were expected to rotate ipsilaterally when stimulated with apomorphine in dose ranges active on the normosensitive postsynaptic DA receptors of the intact side ~. Two weeks after the KA intervention, the rats were challenged with apomorphine 0.5 mg/kg s.c. and their rotational behaviour was continuously recorded in the rotometers. Successfully denervated rats showed ipsilateral rotation. Rats with unilateral KA lesion of the striatum. In order to select the successfully lesioned animals, the rats were challenged with apomorphine 0.5 mg/kg s.c. two weeks after the KA intervention, and their rotational behaviour was continuously recorded. Successfully denervated rats showed ipsilateral rotation after apomorphine in doses active on DA receptors located in the intact side 2~.

Dose-response experiments Dose-response experiments in 6-OHDA-denervated rats with an additive KA lesion of the ipsilateral substantia nigra reticulata. Rats recovered from the acute effects induced by the additive KA lesion in approximately two weeks. Then, the rats were tested with apomorphine 0.02, 0.05, 0.2, 0.5, and 1.0 mg/kg s.c. After a washing out period (l month), the rats were tested with pergolide 0.005, 0.0l, 0.05.0.1 and 0.2 mg/kg s.c. At least 1 week elapsed between every dose.

Dose-response experiments in rats with unilateral KA lesions of the substantia nigra reticulata. Rats with KA lesions of the left substantia nigra which showed ipsilateral rotation after administration of apomor-

Dose-response experiments in rats with unilateral KA lesions of the striatum. Rats with KA lesions of

Histology After the completion of the rotational experiments, the rats were killed by decapitation. The brains were rapidly dissected out and stored in 10% formaldehyde before being embedded in paraffin and cut into serial 10 um sections. Every tenth section was stained with cresyl violet, and the localization and extent of the lesions determined.

Drugs Apomorphine-HCI (Apoteksbolaget, Sweden) and pergolide mesylate (E. Lilly) were dissolved in warm physiological saline. Diazepam (Roche) was injected directly from 5 mg/ml ampoules. All doses were calculated as the free base and injected s.c. in a volume of 1 ml/kg body weight into the left flank.

Statistics Means and S.E.M. were calculated. Dose effects were analysed using F-ANOVA with replications j2. The level of P < 0.05 for the one tail test was considered as statistically significant. RESULTS

Behavioural evaluation of the 6-OHDA lesion From a group of 10 lesioned animals, 8 rats showed the two-peak pattern of rotation when tested with apomorphine 0.05 mg/kg s.c. and 4 were selected for further experiments (total rotations: 61)2 ± 23 turns, duration: 64 ± 3 rain, n = 4) (Fig, 2). The same rats, when tested with pergolide 0.01 mg/kg s.c., showed a strong rotation but without the two-peak pattern (total rotation: 626 ± 143 turns, duration: 167 ± 13 rain, n = 4) (Fig. 3).

[ LJrr~, I(H300

OHDA-denervated rats. T h e

ammals

showed

a

s t r o n g d i a z e p a m - r e l a t e d s e d a t i o n after the i n j e c t i o n

I~OHDA I

of K A and a c o n t r a l a t e r a l postural d e v i a t i o n with =

i

:~

75O0 :

very few seizures. This c o n t r a l a t c r a l postural devia-

i

lion was m a i n t a i n e d for at least one m o n t h after the

I i

Z,

K A m i e c t i o n and a c c o m p a n i e d hv e l e v a t e d m u s c u l a r t o n e especially in the neck region. W h e n stressed,

~= 5 0 0 ( .i

the animals invariably r o t a t e d tov, ards the side contralateral to the d e n e r v a t e d side. { 6OHDA, KA/SNR

2soo

}

Acute behavioural efJ?cts ofthe KA injections. T h e animals r e c e i v i n g a single i n j e c n o n of K A into the

O!

substantia m g r a r o t a t e d c o n t r a l a t e r a t l y just after the OOO1

001

01

-°~'~'APON 4 t0 f l u kq~,, 4~ 16OHDA . K A S N R :

s t e r e o t a x i c injection. O n e m o n t h after the K A treatme|at, the rats s h o w e d a slight postural d e v i a t i o n to-

2 500

wards the l e s i o n e d side. which was e n h a n c e d w h e n they w e r e stressed. T h e rats r e c e i v i n g two K A inlec-

Fig. 1. Modification of apomorphine and pergolide dose-dependent contralateral rotation in 6-OHDA-rats by an added KA lesion of the ipsilateral substantia nigra reticulata. The modification of the dose-dependent contralateral rotation induced by apomorphine (n = 12, triangles) and pergolide (n - 4, circles) in 6-OHDA-rats (data from Herrera-Marschitz and Ungerstedt 13) after an added KA lesion of the ipsilateral substantia nigra reticulata (n = 4, dotted lines). The data is expressed as the total contralateral minus ipsilateral rotation alter the respective doses. It is notable that, following KA lesion, the pergolide curve is shifted to the right while the apomorphine curve is inverted. Abscissa, doses in mg/kg s.c.; Ordinate, total contralatera[-ipsilatera[ rotation.

tions into the [eft s t r i a t u m did not s h o w r o t a t i o n a l activity i m m e d i a t e l y f o l l o w i n g the s t e r e o t a x i c inter~,en-

Turns mid

6OHDA

N=4

20 i .........

~= tpsllateral ° contratateral

10 ¸

30 AOO005

Behavioural evaluation of the KA-lesioned rats KA injected into the substantia nigra reticulata,

Turns rain

F r o m a g r o u p of 8 rats r e c e i v i n g K A into the left sub-

20-

60

6OHDA- KA/SNR

rmr

stantia nigra, 5 s h o w e d a d o m i n a n t ipsilateral rotation w h e n c h a l l e n g e d with a p o m o r p h i n e 0.5 m g / k g s.c. (total ipsilateral r o t a t i o n : 282 +_ 61 turns, total

10-

c o n t r a l a t e r a l r o t a t i o n : 35 + 12 turns, d u r a t i o n : 128 +_ 16 rain, n = 5). All 5 rats w e r e s e l e c t e d for d o s e - r e s p o n s e studies.

KA injected into the striatum. F r o m a g r o u p of 12 rats r e c e i v i n g 2 K A i n j e c t i o n s into the left s t r i a t u m , 8 rats s h o w e d a d o m i n a n t ipsilateral t u r n i n g w h e n chall e n g e d with a p o m o r p h i n e 0.5 m g / k g s.c. (total ipsilateral r o t a t i o n : 216 +_ 38 turns, total c o n t r a l a t e r a l r o t a t i o n : 33 + 9 turns, d u r a t i o n : 140 + 22 rain, n = 8). All 8 rats w e r e s e l e c t e d for the d o s e - r e s p o n s e experiments.

Acute behavioural effects of the additive KA injection into the substantia nigra retieulata of previously 6-

0

120

240min

Fig. 2. Modification of the apomorphine-induced rotation elicited in 6-OHDA-rats by added KA lesion of the=ipsilateral substantia nigra reticulata, 6-OHDA-denervated rats (n = 4) showing a two-peak pattern of contr~[aterat rotation when stimulated with apomorphine 0.05 mg/kg s.c. (upper left of the figure) were subjected to an additional lesion of the ipsilateral substantia nigra reticulata using 1 ug of KA in 1 #l: Onemonth after the lesion, the rats were tested with successively increasing doses of apomorphine up to a dose of I mg/kg s.C. At least one week elapsed between every dose. Abscissa~mindntervals from the time of the apomorphine injection; Ordinate, turns/min.

273

Telr n

6OHDA N=4

ITGID

?C)

10 i

0

=contralateral

T

O 60 PergO01

120 mln

6OHDA

+KA/SNR

Tums rain Perg02 0

400

800

1200

2400

20D

Perg 01

240

300

800

860

10 Perg 0 05

0

~

I%rg 001' 0

"~'~"~ 120

~

~

~

' 240

~

AA

ze, 360 min

Fig. 3. Modification of pergolidc-induced rotation elicited in 6-OHDA-rats by added KA lesion of the ipsilatcral substaniia nigra reliculata. 6-OHDA-denervated rats (n = 4) showing two-peak contralateral rotation when stirnulatcd with apomorphinc 0.05 (tipper left of Fig. 2) were further tested with pergolide 0.01 mg/kg s,c. (upper left of the figure) and then sub ected to an addilional lesion ot lhc ipsilateral substantia nigra reticulata using 1 !zg of KA in 1 !d. Rats were tested with successivelyincreasing doses of {~crgolideup to a dose of 0.2 mgikg s.c. with at least one week elapsing between every dose. Abscissa. rain-intervals from the time of the pcrgolidc mection; Ordinate, Turnsimin.

n o n , but they showed slight epileptic seizures. One month later, the rats showed p e r m a n e n t activated muscular tonus and hyperesthesia.

Changes in rotational behaviour induced by KA lesion oJ" the ipsilateral substantia nigra reticulata oJ" previously 6-OHDA-denervated rats Fig, 1 shows the changes in apomorphine and pergolide d o s e - r e s p o n s e curves induced by the added KA lesion. The comparisons have been carried out using data obtained from previous experiments in animals with 6-OHDA-lesion alone 13. D o s e - r e s p o n s e curves are expressed as the total contralateral minus ipsilateral rotation elicited by the respective doses, The apomorphine curve was profoundly changed. In doses higher than 0.2 mg/kg s.c., apomorphine induced ipsilateral rotation, which was reflected in the

dose-response curve as a shifting to negative values. The quality of the changes induced by the added KA lesion to the apomorphine responses are also indicated in Fig. 2. After the KA lesion, apomorphine 0.02 mg/kg s.c. induced a short peak of contralateral rotation (total rotation: 178 _+ 65, lasting 40 rain, n = 4). After 0.05 mg/kg s.c., apomorphine induced a long lasting contralateral rotation (total rotation: 315 _+ 57 turns, lasting 150 rain. n = 4) without the typical two-peak pattern. At the dose of 0.2 mg/kg s.c. of apomorphine there was a short peak of contralateral rotation lasting 5 rain followed by an intense ipsilateral rotation lasting 30 rain, and then a final peak of contralatera[ rotation. At the dose of I mg/kg of apomorphine, rats immediately started to rotate ipsilaterally (total rotation: 458 _+ 37, n = 4), although a final contralateral peak (total rotation: .6_~ ~ ~+ _ 68, n =

"y'v ../4

4) occurred after 100 rain. At this dose, a p o m o r p h i n e induced strong, mutilating, gnawing and biting on the tail and the legs on the side of the lesion. The intensity, as well as the duration, of this ipsilateral turning increased in a d o s e - d e p e n d e n t m a n n e r up to the dose of 1 mg/kg s.c. of a p o m o r p h i n e ( F - A N O V A = 15, P < 0.001, N , M = 4,5), while the contralateral turning did not show any statistically significant dose-dependent increase. The pergolide d o s e - r e s p o n s e curve was shifted to the right after the K A lesion. The shift in potency was approximately 10 times. A t the dose of 0.01 mg/kg s.c., pergolide induced a low intensity contralateral rotation (total turns: 132 +_ 44, n = 4), followed by a long period of short bursts of turning (Fig. 3). A t the dose of 0.05 mg/kg s.c., pergolide induced stronger contralateral rotation (total turns: 338 _+ 148, n = 4). A t the doses of 0.1 and 0.2 mg/kg s.c., pergolide induced very long lasting contralateral turning (total turns: 1673 +_ 569 and 3998 +_ 1083, lasting 1086 and 2524 rain, respectively, n = 4). T h e r e was no ipsilateral activity nor any strong gnawing/biting b e h a v i o u r even at the dose of 0.2 mg/kg s.c. when the rats turned for a p p r o x i m a t e l y two days.

Turns ~

---

f3

500KA/SNR N=5

± 0

~: 250-

A p o E5

Apo 1.0

Perg 0.1

Perg o.2

KA/Striatum N-8

Turn~

1OO(

,. * * - - 7

L C O

500 "5 B

Dose-response experiments in KA-lesioned rats Unilateral KA lesions of the substantia nigra reticulata. The total rotation induced by a p o m o r p h i n e and pergolide in rats with unilateral lesion of the substantia nigra reticulata are shown in the u p p e r part of the Fig. 4 (A). The ipsilateral rotation induced by apom o r p h i n e increased in a d o s e - d e p e n d e n t m a n n e r (FA N O V A = 34, P < 0.01, N , M = 5,2). In contrast. pergolide did not induce rotational activity at the tested doses, although the animals were clearly activated as revealed by the increase in both the ipsilateral and the contralateral counts. A f t e r pergolide 0.2 mg/kg s.c., the rats tried to leave the rotational bowls, and showed m o v e m e n t s of the head accompanied by sniffing to both sides, but no rotational activity was recorded. Unilateral KA lesions of the striatum. The total rotation induced by two doses of a p o m o r p h i n e and pergolide in rats with lesions of the left striatum are shown in the lower part of Fig. 4 (B). A p o m o r p h i n e induced d o s e - d e p e n d e n t ipsilateral rotation (FA N O V A = 18, P < 0.01, N , M = 8.2). Pergolide also induced d o s e - d e p e n d e n t ipsilateral rotation

ADO 0.5

ADO ,5.8

Perg 0.1

Perg 0.2

Fig. 4, Rotational behaviour induced by apomorphine and pergolide in KA-lesioned rats. Rats with KA lesions into the left substantia nigra reticulata (SNR) (A) or into the left striatum (B) which showed ipsilateral rotation when stimulated with apomorphine 0.5 mg/kg s.c. were selected to be tested with successively increasing doses of apomorphine and pergolide. At least two weeks elapsed between the respective doses, Right and left 360~ turns/rain were recorded in rotometers for the entire duration of the rotational behavlour, and are expressed as total (means and S,E.M ) ipsi{ateral (white columnsl or contralateral (hatched columns) rotation. Dose effects were analysed using F-ANOVA with replications. P<0.05=*.P
( F - A N O V A = 13, P < 0.05. N . M = 8.2), while the contralateral activity did not change significantly.

Histological evaluation of the kainic acid lesions KA-induced changes in the substantia nigra. Light microscopical analysis s h o w e d extensive histological changes affecting the substantia nigra and a d j a c e n t regions. T h e most m a r k e d effect occurred in the sub-

275 slantia nigra reticulata that showed extensive disappearance of nerve cells as well as gliosis. Traces of

KA-induced changes @er injections into the striaturn. Using the light microscope, the K A treatment

the injection cannula were clearly visible in slices corresponding to the levels A2790-2180/~ according to the K6nig and Klippel atlas 17 (Fig. 5). The n u m b e r of nerve cells in thc zona compacta (A9 cell group according to DahlstrOm and Fuxe -~) was diminished. while the Alfl group in the midline seemed to be tinaffected. Trace of damage was observed in the ForeI's field tt2, and some rats showed damage reaching the caudal part of the nucleus subthalamicus, The damage induced by the added 6 - O H D A - and KA treatment was extensive in rostro-caudal as well as in dorso-ventral directions, and damage was also observcd in the z~ma incerta.

was seen to have induced a massive loss of nerve ceils in the striatum on the o p e r a t e d side, intensive gliosis and a m a r k e d reduction of the size of the striatum.

E

i~~ ~' (

I

Fig. 5. Reconstruction ol Icsion produced by injecting KA into the lcfi subslantia nigra reticulata. Successive frontal planes show the cffect of 0.5 ug of KA in 0.5 ul injected into the lelt

substantia nigra 1 year bclore. Damage of the brain tissue, including marked disappearance of nerve cells, intensive gliosis and prolifcration ol non-neuronal elements, were observed approximatcly 1 mm rostral and caudal to the injection place, ltowe~ci, thc spreading of damage in a dorso-ventral extension was minimal since, in almost all rats. nerve cells were ohserved in areas corresponding to the zona compacta. The arrows in F-I indicate the traces of the injection cannula. Coordihales rclcr to the Kimig and Klippel atlase?.

DISCUSSION

A unilateral 6 - O H D A injection into the ascending D A axon bundle in the area ventralis tegmenti induces a nearly complete degeneration of the D A nerve terminals in ipsilateral forebrain areaseS. Animals with such a lesion show a postural deviation towards the side of the lesion. T r e a t m e n t with the D A agonist a p o m o r p h i n e reverses this deviation, inducing rotational behaviour towards the opposite side, due to stimulation of supersensitive D A receptors in the D A - d e n e r v a t e d areas 2,>j,~5.>. In a recent study, comparing the D A agonists apomorphine and pergolide, considerable differences were found 13,>. A p o m o r p h i n e induced a two-peak, head to tail, tight rotation accompanied by intense gnawing and biting while pergolide induced a onepeak, less tight rotation with less oral stereotypies. When pretreating the rats with different D A antagonists, we found that the D2 antagonist sulpiride ll,14,i<'.~'a inhibited pergolide rotation in doses 1000 × lower than those required to inhibit the apomorphine response I:;,>. The D1/D2 antagonist cisflupenthixo115, on the other hand, inhibited both of the agonists in a similar dose range I:~.>. We suggested that the results may be a functional expression of two different populations of D A receptors. The importance of the striatonigral effercnt pathway for the a p o m o r p h i n e rotation in unilaterally 6O H D A - d e n e r w l t e d rats has heen well documented 4,i<>. K A , locally injected into the substantia nigra, destroys the perikarya of cells receiving striatonigral efferents a,5 and blocks rotation in a similar way to electrolytic lesions of nigral neuronse:. An injection of K A into the ipsihiteral substantia nigra reticulata of previously 6 - O H D A - d e n e r v a t e d rats induced, acutely, a contralateral deviation that remained at least one month after the lesion. A remaining exacerbated muscular tone was evident in the neck region. When challenged with low doses of apomorphine, the 6-OHDA/KA-lesioned rats showed a marked inhibition of the contralateral rota-

27~ tion,

which

is in

accordance

portsS,6,m,lL A p o m o r p h i n e

with

previous

re-

al p e a k , s t r o n g i p s i l a t e r a l r o t a t i o n t h a t i n c r e a s e d in

in d o s e s a b o v e 0.5 m g / k g

i n t e n s i t y w i t h t h e dose. H o w e v e r , a final p e a k of c o n -

s.c. i n d u c e d , a f t e r a n initial s h o r t l a s t i n g c o n t r a l a t e r -

tralateral

a) 6 0 H D A - model

rotation

was

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Fig. 6. Rotational models. A: 6-OHDA model, a stereotaxic unilateral injection of 6-OHDA into the area ventrali tegmenti, which contains the bundle of axons leaving the mesencephalic DA cell bodies, induces an unilateral lesion that extensivel~ denervates thc DA-innervated forebrain areas unilaterally4z,46. In this model, both apomorphine and pergolide induce contralateral rotation acting on postsynaptic DA receptors that become supersensitive after they lost the DA innervation. B: 6-OHDA + KA/SNR model, a KA injection into the ipsilateral substantia nigra reticulata of previously 6-OHDA-denervated animals impairs the continuation of the striato-nigral efferent pathway, which is revealed in the rotational model as a shift to ipsilateral rotation when apomorphine is given in postsynaptic dose-range, acting on normosensitive DA receptors of the intact side m. Moreover, although inhibited, the contralateral dose-dependent response induced by pergolide remains unchanged. C: KA/striatum model. Animals with massive KA destruction of one striatum rotate ipsilaterally when stimulated with apomorphine 3s or pergolide in range doses active on normosensitive receptors of the intact side. D: KA/SNR model, an unilateral KA lesion of the substantia nigra reticutata impairs the continuation of the striatonigral efferent pathway alone. In this model, apomorphine induces ipsilateral rotation, probably, by stimulating asymmetricall} the DA receptors on the intact side in line with the striato-nigral efferent pathway that becomes in imbalance after the KA treatment. In this model, pergolide stimulates DA receptors that do not become in imbalance after the KA impairment of the substantia nigra, and then, the animals are activated symmetrically.

277 Fig. 2). In contrast, pergolide did not induce any ipsilateral rotation even at the highest doses ((I.2 mg/kg) when the contralateral rotation lasted for more than 42 h (Fig. 3). The main difference being a shift to the right of the pergolide dose-response curve (Fig. 1), The dissociation of the apomorphine response into two components, contralateral and ipsilateral rotation in 6 - O H D A / K A animals suggests that one of these components revealed as ipsilateral rotation, requires striato-nigral efferents. The ipsiJateral rotation induced bv apomorphine is probably due to a stimulation of DA receptors in the intact side which dominates over the lesioned side. The pergolide-induced contralateral rotation, although inhibited after the KA treatment, was still present and no ipsilateral rotation was observed. KA injected into one substantia nigra reticulata of normal rats mainly destroys the cell bodies of neurons receiving striato-nigral efferents. This lesion is revealed as an imbalance syndrome that may be stimulated with doses of apomorphine able to act on normosensitive postsynaptic DA receptors. Rats rotated towards the lesioned side as the result of a selective action on the intact side (Fig. 4A). In this model, pergolide was not able to induce any significant rotational activity (Fig. 4A), which may be interpreted as due to the inability of pergolide to stimulate normosensitive DA receptors, while apomorphine has this ability. However, this seems not to be the case, since pergolide is at least li) × more potent than apomorphine in eliciting ipsilateral rotation in animals with unilateral KA lesions of the striatum (Fig, 4B)I-L The rat with unilateral KA lesions into the striatum has been considered as a model for studies on intact DA receptors since KA causes a lesion of all postsynaptic striatal efferent pathways-'l. Furthermore, pergolide induced marked activation in animals with KA lesion of the substantia nigra, as revealed by an increase in both ipsilateral and contralateral counts, indicating a stimulation of normosensitive DA receptors that are not in imbalance. A schematic illustration of the rotational models is shown in Fig. 6a-d. The main effects induced by pergolide may be due to selective actions on receptors located in nucleus aceumbens -~'~. However, major accumbens efferents

seem to descend to the substantia nigra 7,-'0, therefore a unilateral lesion of the substantia nigra should impair pergolide as well as apomorphine responses. In addition, we have found that both apomorphine and pergolide induce strong contralateral rotation when injected into the denervated striatum of 6-OHDAtreated rats, in volumes (0.5 ul) which, probably, do not diffuse significantly into the accumbens ~. According to our interpretation, the apomorphine effects are mainly mediated by a striato-nigral pathway passing through the globus pallidus. This interpretation is in agreement with the findings of SlateN -~ showing that electrocoagulations of the globus pallidus abolished the apomorphine rotation in previously 6-OHDA-denervated rats, while KA injected into the external segment of the globus pallidus had little effect on the apomorphine rotation. These findings suggest that the apomorphine response is revealed by fibres passing through the globus pallidus s which arrive in the substantia nigra. The impairment of the striatonigral efferents abolishes the contralateral rotation induced by apomorphine in 6-OHDA-denervated rats. Instead, apomorphine induces ipsilateral rotation whenever apomorphine is administered in doses high enough to be active on normosensitive postsynaptic DA receptor located in the intact side. The pergolide rotation, on the other hand, may mainly depend upon other striatal efferent pathways. Since in previous studies 1-~.> we have found that apomorphine- and pergolide-induced rotational behaviour may be differently inhibited by DA antagonists acting on D I or D2 receptors, we suggest that different populations of DA receptors may be associated with different striatal efferents. ACKNOWLEDGEMENTS We greatfully acknowledge the skilled technical assistance of Mr, Dan Olsson, Mr. Magnus Hedberg and Ms. Kjerstin Cason for expert secretarial work. The study was supported by grants from the Swedish Medical Research Council (03574, 81/2136 A15/253), Bergwdls Stiftelse, the Foundations of the Karolinska Institute, and the Department of Psychology, University of Stockholm.

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