Dopamine agonist-induced restoration of drinking in response to hypertonie saline in adipsic dopamine denervated rats

Brain Research Bluefin, Vof. 8,

pp. 3X-379, 1982. Rated in the U.S.A.

Dopamine Agonist-Induced Restoration of Drinking in Response to Hypertonic Saline in Adipsic Dopamine Denervated Rats COLIN T. DOURISH’ Psy~h~at~~c Research ~jvisj#n,

~niversjty

AND ROLAND S. G, JONES2

~osp~t~~, Saskutuon,

Received 5 October

Saskatchewan

S7N OX0 Canada

1981

DOURISH, C. T. AND R. S. G. JONES. Dopamine agonist-induced restoration of drinking in response to hypertonic denervated rats. BRAIN RES. BULL. &4) 375-379, 1982.--Bilateral 6-hydroxydop~~e lesions of the ascending dopamine neurones of the nigro-striatal bundle or biIateraI electrolytic lesions of the Iateral hypothalamus cause a syndrome of apbagia and adipsia. Rats with such lesions do not drink in response to hypertonic saline which causes thirst in intact animals. However, the systemic administration of the direct dopamine receptor agonists apomorphine, piribedil and lergotrile restored drinking in response to hypertonic saline in ~hyd~~ydop~e treated rats but not in rats with electrolytic lesions of the lateral hypothalamus. Amphetamine and &phenylethylamine failed to restore drinking in response to hypertonic saline in either group of lesioned rats which suggests that these drugs may act indirectly on the dopaminergic system. The restoration of drinking model suggests that dopamine may have a permissive function in thirst, and, in addition, appears to represent a usefut test for the screening of new drugs for a direct dop~~e receptor stimulating action.

saline in adipsic dopamine

Adipsia Restoration

6-Hydroxydop~ine Lateral hypothalamic of drinking Dopamine Thirst

BILATERAL electrolytic lesions of the Iateral hypothalamus (LH) in rats and other animals result in the complete

cessation of ingestive behaviour [1,29]. Ungerstedt [30] proposed that LH lesions, as well as most ext~y~th~~ic lesions that produced aphagia and adipsia, would interrupt dopamine (DA)-containing neurones, as they coursed through the ventral diencephalon. In addition, he observed [31] that bilateral injections of the neurotoxin dhydroxydopamine (6-OHDA) which selectively lesions catecholamine neurones 1161 produced the same syndrome of aphagia and adipsia. After LH or 6-OHDA lesions, rats do not drink in response to physiological stimuli such as hypertonic saline which elicits thirst in intact animals 113, 14, 211. Recently it has been demonstrated that the systemic administration of a low dose of the DA receptor agonist apomorphine can restore drinking in response to hypertonic saline or isoprenaline in 6-OHDA lesioned rats but not in LH lesioned rats [23]. It has been proposed 1191 that this type of experiment might represent a useful test for the screening of possible antiparkinsonian drugs. The goal of the present study was to further examine the possible role of the DA system in the control of drinking using a number of compounds which have been proposed to act directly (e.g., ‘To whom reprint requests should be addressed. *Present address: Research Department, Pharmaceuticals

Copyright

8 1982 ANKHO

lesions

Dopamine agonists

a~mo~hine, piribedil, ~-p~nylethyl~e [PEA]) or indirectly (e.g., amphetamine, PEA) on DA receptors. Restoration of regulatory drinking in response to hypertonic saline adm~istration in 6-OHDA and LH lesioned rats was used to examine the importance of DA transmission in the arousal of thirst. In addition this paradigm was used as a criterion to test for direct post-synaptic DA receptor stimulation. METHOD AhlUlS Male Sprague-Dawley rats (250-300 g) were used in the experiments. Prior to surgery animals were maintained in hanging wire cages with food and water ad lib, Lighting operated on a 12 hour dark/light cycle (lights on 6 a.m.) and temperature was maintained at 2O-22°C.

Surgery The rats were randomly allocated to one of 3 surgical treatments: unoperated controls (n= IO), bilateral injection of 6-OHDA into the ascending DA axons (n=20), or bilateral electrolytic lesions of the lateral hypothalamus (n = 11). Prior

Division, CIBA-GEIGY

international

Limited, CH-4002 Basle, Switzerland.

Inc .-036I-9230/82/040375-05$03.00/O

376

DOURISH AND JONES TABLE I DRUGS USED IN THE INVESTIGATION

Route

Dose

6-Hydroxydopamme hydrochloride /3-Phenylethylamine hydrochloride Apomorphine hydrochloride d-Amp~et~e sulphate

IC IP IP IP

g/% 12.5, 50 m&g 0.1 mgkg 0.25,0.5 mgkg

Lergotrik mesylate Piribedil Methane Sulphonate p-Tyramine Monohydrochloride

IP IP IP

3 mgk3 5 mg/kg 50 m&g

to surgery the animals were anaesthetized with pentobarbitone (45 mg/kg IP) and placed in a Kopf stereotaxic instrument. To destroy the ascending DA neurons 8 Fg of 6-OHDA dissolved in 4 ~1 of ascorbic acid (0.2 mg/ml) vehicle was injected on each side into the ventral tegmental area (co-ordinates: 4.4 mm posterior to bregma, 1.1 mm Iateral to the sag&al suture and 7.2 mm ventral to the dura, according to the atlas of Konig and Klippel [18]) at a rate of 1 ~1 per minute. To destroy the LH, radio-frequency lesions (20 MA for 10 seconds) were made bilaterally with a Grass LM4 lesion maker (co-ordinates: 2.5 mm posterior to bregma, 2.0 mm lateral to the sag&al suture and 8.0 mm ventral to the dura). After surgery animals were housed ~di~d~ly in metal cages (30 cmx 20 cm x 12 cm). Pellets on the cage floor and water from a 100 ml calibrated bottle were available at all times. In addition the rats were given a liquid nutrient (8 ml PG daily, Ensure, Abbott Labs Ltd.) to maintain viability. Food and water intake and body weight were monitored carefully during the i~e~~te post-operative period. Since the study was designed to examine restoration of drinking in response to hypertonic saliue any lesioned animal which drank in response to this challenge alone was excluded from the experiments. Using this criterion 1 LH lesioned rat and 10 6-OHDA lesioned rats were excluded. Apparatus and nesting Procedure

The testing procedure was similar to that used by Marshall and Ungerstedt [23]. Starting 2 days after surgery the rats were tested during the following 8 days for their drinking response to hypertonic saline (1 M NaCl at 8 volume of 20 ml/kg) which is a potent thirst stimulus in normal rats 1171. Equal numbers of animals were assigned to 2 major treatment groups designated A and B. Each group eventually consisted of 5 unoperated controls, 5 rats with 6-OHDA lesions and 5 rats with electrolytic lesions. Drinking responses were determined in large individual metal cages (45 cmx30 cmx20 cm) to which animals were habituated for 30 minutes prior to the start of each test. Water was provided in shallow petri dishes in order to minimize any motor problems in drinking which lesioned rats may have [21]. At the start of each test the dish was weighed and then positioned adjacent to the rat. At 5 minute intervals during the test, those rats which had walked away from the dish were picked up and a@n placed adjacent to it. If a rat started to walk into the water dish it was picked up and placed next to the dish. At the end of the test the water dishes were weighed and water intake for each animal was

Source Sigma Sigma Sigma Smith, Kline and French Eli LiIIy Servier Calbiochem

calculated (including a correction tion). The following

for spillage and evapora-

tests were administered

to each rat: (1)

Distilled water. The rats were injected with distilled water (1 m&g) and a dish of water was immediately placed in the cage for a 30 minute drinkii test. (2) Hypertonic saline. The rats were injected with NaCl. Thirty minutes later the rats were injected with distilled water and a dish of water was placed in the cag;efor a 30 minute drinking test. (3) Test drug. The rats were injected with the test drug and a dish of water was immediately placed in the cage for a 30 minute drinking test. (4) Hypertonic saline plus test drug. The rats were given an injection of NaCI. Thirty minutes tater the rats were injected with the test drug and a water dish was placed in the cage for a 30 minute drinking test. Animals in group (A) were tested with PEA, piribedil, and lergotrile. Animals in group (B) were tested with apomorphine, p-tyramine and amphetamine (see Table 1 for doses and suppliers). Drugs were dissolved in distilled water except for 6-GHDA which was dissolved in ascorbic acid. Doses of drugs are expressed in terms of the salt. The order in which the tests were administered was varied between rats and there was a minimum of 24 hours between successive tests. H~st~lugy

Following completion of the experiments the brains of all rats were removed and stored in formaldehyde-saline for 1 week before histological verification of lesion locations.

RESULTS

Unoperated

Controls

Hypertonic saline produced a consistent drinking response (mean intake=7.6 ml) in normal rats during a 30 minute test (see Table 2 and Fig. 1). The animals did not drink in response to any of the test drugs or the distilled water control injection. Piribedil and PEA (50 mgkg) signifcantly decreased drinking in response to hypertonic saline (Table 2). In contrast, animals treated with a low dose (0.25 mg/kg) of d-amphetamine, in combination with hypertonic saline, showed elevated water intakes in comparison to rats treated with hypertonic saline alone (Table 2). A higher dose of d-~phet~e (0.5 mg/kg) or a lower dose of PEA (12.5 mglkg) had no effect on hypertonic saline-induced thirst.

DOPAMINE AGONISTS AND THIRST TABLE 2 EFFECTS OF PUTATIVE DOPAhfINERGIC DRUGS ON DRINKING IN HYPERTONIC SALINE-TREATElD RATS Water Intake (ml)

Drug

Unoperated Controls

7.60 z?z0.55 7.86 + 0.84 2.72 + I.Olt 6.18 z!z1.3 11.82 rl; l.J6* 6.02 +- 1.74 6.96 4 0.91 4.36 f 0.51” 7.72 z+z1.40

Distilled Water ~-Phenyle~yl~~ne Apomorphine d-Amphe~ine Lergotrile Piribedil p-Tyramine

Electrolytic Lesion

~Hydroxydopam~ne Lesion

Distilled Water ~-Phenyiethyl~i~e

12.5

A~rno~~e d-Amphetamine

50 0.i 0.25 0.5

Lergotrile Piribedil p-Tyramine

3 5 50

Distilled Water ~-Phenyle~yl~ine Apomorphine d-Amphetamine Lergotrile

~~bedil p-Tyramine

12.5 so 0.1 0.25 0.5 3 5 50

0 0 0 0 0 0 0 0 0 0 0 0 6.16 rfr 1.42t 0 0 6.25 r 0.61f 6.86 Lt:0.73t 0

All values are mean 2 SE of 5 subjects (except unoperated control ~st~l~ water, n- 10). *p
Lesioned Animah Both electrolytic and 6-OHDA lesioned animals were aphagic and adipsic during the entire test period and had to be fed iutragastricaRy to maintain viabiity. The animals were hy~~s~nsive to stimuli, cataleptic and akinetic but not somnolent, as has been previously reported [22]. In contrast to normal rats, lesioned animals did not drink in response to hypertonic saline (Table 2). However when 6-OHDA lesioned rats were tested with apomorphine, piribedil or Iergotrile, in combination with hypertonic saline, they became activated and drank a sin&u volume of water to that consumed by intact controks (see Fig. 1). Administration of a~rno~~e, piribedil or lergotrile alone produced snifRng, walking and exploration but no drinking (Fig. 1). PEA, p-tymmine and d-amphetamine failed to induce drinking when ad~iste~ aione or with hypertonic saline to GOHDA lesioned animals (Table 2). The high doses of PEA and amphetamine produced occasional gnawing and biting at the water dish but no prolonged drinking, whereas low doses of the same drugs had little visible effect. None of the drugs tested either aIone, or in ~ornb~a~on with hypertonic saline, elicited drinking in rats with electrolytic lesions of the LH,

although all drugs (with the exception of p-tyramine) caused some behavioural activation. This activation took the form of stereotyped head movements, sniffing and gnawing, in contrast to the walking and exploration which characterized 6-OHDA lesioned animals.

oxidation of the brains of ah rats which failed to drink after electrolytic lesions revealed that the lateral hypothalamic area received substantial damage. In the majority of animals the lesion was centered on the lateral h~th~~us but occasiona;Hy encroached dorsally into Forels fields H, and H,, medially on the columna fomicis, do~orne~~ and ventromedial hypothalamic nuclei and laterally into the internal capsule. Only one animal was not adipsic tier electrolytic Iesioning and subsequent ~sto~ogy revealed that the lesions were misplaced dorsally and did not destroy the iateral hypothalamus. DISCUSSION

The putative DA receptor agonists a~rno~h~e,

phibedil

378

12

DOURISH

1

a

1

b

6-OHDA

d

1

b- OHDA

1

f

6-OWDA

l2

Control

APO

-1 12

12 7”

c

Control

12

Control

‘2

FIG. 1. Water intake in uno~rated controls and rats with bilatera1 6-hvdroxvdonamine lesions (COHDA) during a 30 minute drinking tes; after’ thi administration ‘of distilled water (DW), apomorphine (APO), hypertonic saline (HS), piribedil (PIR), lergotile (LER), apomorphine plus hypertonic saline (HS+AF’G), piribedil plus hypertonic saline (HS+PIR) or lergotrile plus hypertonic saline (HS+LER). All values are mean-t-SE of at least 5 determinations *p
and lergotrile, all restored drinking in response to hypertonic saline in rats with bilateral 6-OHDA lesions of the ascending DA-~on~~g pathways (Fig. 1). The results with apomorphine confirm a previous observation 1231 and extend it to other DA receptor agonists. In contrast amphetamine, PEA and p-tyramine failed to restore regulatory drinking in 6-OHDA lesioned animals. It is well established that amphetamine acts indirectly on the DA system, probably by releasing DA and inhibiting its reuptake [3, 4, 61 and it is possible that PEA and p-tyramine could have similar actions [27]. In addition Antleman and co-workers [2] have proposed, on the basis of a rat rotational model, that PEA may directly stimulate post synaptic DA receptors. The latter proposal is not supported by the present results since in our model DA receptor agonists restored regulatory drinking but PEA did not. However we cannot draw any definite conclusion from the present experiments with regard to the possible DA agonist properties of p-tyramine. While a previous report

AND JONES

indicates that p-tyramine does not readily cross the bloodbrain barrier [25], recent evidence from this laboratory suggests that at least a small proportion of the compound might reach the brain after peripheral administration 1241. The failure of p-tyramine to cause any behavioural stimulation in our experiments suggests that little or none of the systemically administered compound reaches the central nervous system. Therefore elucidation of the possible DA receptor agonist properties of p-tyramine must await experiments employing direct intracerebral injections of the amine. A previous study [23] has indicated that, in contrast to 6-OHDA lesioned rats, regulatory drinking in LH lesioned animals is not restored by administration of apomorphine. Again, the present results confirm this finding and extend it to other direct DA receptor agonists (piribedil, lergotrile) and also to indirectly acting agonists (amphetamine, PEA). Marshall and Ungerstedt 1231 have proposed that the difference between 6-OHDA lesions and LH lesions is the destruction by the latter of non-dopaminergic pathways (either ascending or descending) which may be essential for the maintenance of normal ingestive function. Although our results do not provide any further insight into the workings of such a mechanism they are in general agreement with its existence since the only apparent effect of dopaminergic drugs in LH lesioned animals was the production of stereotyped head movements, gnawing and sniffing. Therefore it would seem likely that dopaminergic neuronal damage may not be the only important consequence of electrolytic LH lesions. Presumably damage to tibres or cells of other neurotransmitter systems (which could be cholinergic or noradrenergic) may be involved in the lateral hypothalamic syndrome. This interpretation is consistent with the relatively weak enhancement of drinking which is seen in intact rats after central or peripheral administration of a DA agonist, in comparison to the potent dipsogenic properties of the cholinergic agonist carbachol, or the @drenergic agonist, isoprenaline [ 10, 11, 26, 281. Dopaminergic transmission in the nigro-striatal pathway appears to be necessary for the arousal of drinking since DA agonists can stimulate water consumption in adipsic 6-OHDA lesioned animals. However, the failure of DA receptor stimulation to cause thirst in LH lesioned animals suggests that the nigro-sttiatal DA system may play a permissive role in regulatory drinking behaviour (which could be related to its importance in motor control), but is probably not involved in the arousal of thirst. Interestingly a number of the putative dopaminergic drugs used in our experiments had significant effects on the hypertonic saline drinking challenge in neurologically intact animals. A low dose of piribedil depressed drinking and appeared to induce sedation, which is consistent with previous reports of the effects of treatment with a low dose of a DA agonist [5, 12, 201. Similarly, PEA at a dose of 50 mgkg significantly reduced water intake. This effect may have been the result of response competition since the compound is known to induce a stereotyped behavioural syndrome, which is characteristic of stimulation of central SHT activity, at this dose level [7]. However, a.behaviourally specific suppression of drinking cannot be discounted since PEA is known to be a potent anorectic agent and significantly decreases 24-hour water intake after chronic administration for a 7 day period [8,9]. In contrast, a low dose of amphetamine signi~c~tly elevated drinking in response to hypertonic saline, which is in agreement with the report of enhancement

~PA~INE

of prandial drinking after food deprivation by a low dose of amphetamine [15]. In conclusion, the present results confirm and extend the observation that direct DA agonists reverse adipsia caused by &OHDA lesions, thus supping the hy~~esis that an intact nigro-striatal DA system is necessary for the arousal of thirst. However, the failure of DA agonists to restore drinking in LH lesioned animals suggests that the nigro-striatal DA pathway may play a permissive role in thirst, but is not specifically involved in the control of drinking. Restoration of drinking in response to physiological stimuli in 6-OHDA lesioned rats appears to be a useful test for the demonstration of direct Dk- receptor activation. In 1. Anand, B. K. and J. R. Brobeck. Hypothalamic control of food 2. 3. 4.

5. 6.

7.

8.

9.

10.

11. 12.

13.

14.

15.

379

AGONISTS AND THIRST

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model a~mo~h~e, piribedil and lergotrile behave as direct DA receptor agonists whereas PEA and amphetamine do not. The model may prove useful in the future as a tool for screening possible dopaminergic drugs to be used in the treatment of extrapy~mid~ motor disorders or affective illness.

this

The following companies generously donated drugs used in the study: d-amphetamine sulphate (Smith, Kline and French, Canada), lergotrile mesylate (Eli Lilly and Co. USA), piribedil methane sulphonate (Servier U. K. Ltd.) The authors thank Dr. A. A. Boulton for enco~~ement and provision;of facilities, and S~kat~hew~ Health and the M. R. C. of Canada for continuing financial support.

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