The effect of antiparkinson drugs on the linguomandibular reflex in cats

EUROPEAN JOURNAL OF PHARMACOLOGY 21 (1973) 178-182. NORTH-HOLLAND PUBLISHING COMPANY

THE EFFECT OF ANTIPARKINSON LINGUOMANDIBULAR

DRUGS ON THE

REFLEX IN CATS

A. BARNETT and J.W. FIORE Department o f Pharmacology, Biological Research, Schering Corporation, Bloomfield, N.J. 07003, U.S.A.

Received 3 March 1972

Accepted 2 November 1972

A. BARNETT and J.W. FIORE, The effect o f antiparkinson drugs on the linguomandibular reflex in cats, European J. Pharmacol. 21 (1973) 178-182. The linguomandibular reflex (LMR) is a polysynaptic reflex involving interneurons within the brain stem. Electrical stimulation of areas in the basal ganglia have been shown to influence this reflex. The effect of known antiparkinson drugs on the linguomandibular reflex in cats was investigated, l-Dopa and apomorphine produced dose-related inhibition of the LMR whereas amantadine and benztropine were ineffective. Apomorphine-induced inhibition of the LMR was antagonized surmountably by haloperidol, a relatively specific dopamine antagonist but not by phenoxybenzamine, an a-adrenergic blocking agent. 1-Dopa-induced inhibition of the LMR was not antagonized by either haloperidol or phenoxybenzamine. Known metabolites of l-dopa, dopamine and norepinephrine were tested for their effects on the LMR and none was more potent than l-dopa. These results suggest that l-dopa might inhibit the LMR via a direct effect of its own. Antiparkinson drugs Dopamine receptors

l-Dopa

1. Introduction l-Dopa, apomorphine and amantadine reduce the symptoms of rigidity and akinesia in Parkinson patients whereas benztropine has little or no effect on these symptoms and is used for the relief of tremors (Barbeau et al., 1962; Birkmayer and Hornykiewicz, 1961, 1962; Cotzias et al., 1967; Cotzias and Papavasiliou, 1967; Barbeau, 1969; Schwab et al., 1969; Cotzias et al., 1970). Parkinsonian rigidity and akinesia are thought to be due to a deficiency of dopamine in the basal ganglia (Ehringer and Hornykiewicz, 1960; Barbeau et al., 1962; Birkmayer and Hornykiewicz, 1962; Cotzias and Papavasiliou, 1967; Barbeau, 1969). l-Dopa is thought to act via its conversion to dopamine (Birkmayer and Hornykiewicz, 1962; Cotzias and Papavasiliou, 1967; Barbeau, 1969; Davidson et al., 1971) and apomorphine is thought to stimulate dopamine receptors directly (Ernst, 1965; Cotzias et al., 1970). It has been hypothesized that

Apomorphine Amantadine

amantadine releases dopamine (Grelak et a1.,1970; Scatton et al., 1970; Stromberg et al., 1970; Voigtlander and Moore, 1971), and benztropine is thought to act via a central anticholinergic effect. 1-Dopa has previously been reported to antagonize polysynaptic reflexes in the spinal cord but its mechanism of action is unclear (Baker and Anderson, 1970a,b). The linguomandibular reflex (LMR) is a polysynaptic reflex which involves interneurons in the brain stem. There are connections between areas of the basal ganglia containing dopaminergic neurons e.g. caudate nucleus, globus pallidus, and the brain stem reticular formation (King and Unna, 1952; King et al., 1955). Moreover, electrical stimulation of the caudate nucleus and globus pallidus influence this reflex (King and Unna, 1952; King et al., 1955). The present studies were devoted to investigating the effects of known antiparkinson drugs on the LMR and their possible modes of action.

179

A. Barnett, J. W. Fiore, Effect o f antiparkinson drugs on the LMR

2. Materials and methods Cats of either sex, 2 - 4 kg in body weight, were anesthetized with a mixture of diallylbarbituric acid, 60 mg/kg, i.p., and urethane, 240 mg/kg, i.p. Cannulae were placed in the femoral vein for i.v. administration of drugs and in the trachea for artificial respiration, if necessary. Blood pressure was measured using a Statham model P23AC pressure transducer connected to a Grass Model 7 polygraph. The head o f each cat was placed in a holding device to facilitate measurement of the LMR. A piece of 00 surgical thread was tied to the skin at the tip o f the mandible and connected to a Grass FT-IO force-displacement transducer which in turn was connected to the polygraph for measurement of the force of contraction of the LMR. 2 needle electrodes were placed on the underside of the tongue and these were connected to a Grass S-4 stimulator for electrical stimulation o f the tongue. Stimulus parameters were: frequency, 6/min; duration, 500 msec; and sufficient voltage to produce a force of contraction of 4 0 0 - 5 0 0 nag. Force of contraction was kept relatively constant to insure that all drugs were compared against the same baseline of measurement. Increasing i.v. doses of test drugs were administered at 45-min intervals until complete inhibition o f the LMR resulted. In cases wherein no LMR inhibition was obtained, dosing was continued until there was lethality. If the LMR did not return to baseline at the end of each 45-min interval, the voltage was raised by not more than 20%. Changes in the force o f

contraction of the LMR are expressed in terms of the control measurement before each dose. Inhibition of the LMR is expressed as the maximum inhibition occurring within 10 rain after each i.v. dose. For experiments involving drug antagonism, the potential antagonist was administered 10rain before obtaining the agonist d o s e - r e s p o n s e curve. Potential antagonists were used only in doses that did not significantly affect the LMR. Drugs were administered i.v. in a volume of 0.1 ml/kg. All drugs were suspended in a dilute aqueous vehicle of 0.5% carboxymethylcellulose, with the exception of apomorphine'HCl and anaantadine.HCl, which were dissolved in distilled water.

3. Results l-Dopa and apomorphine produced dose-related inhibition of the I_MR whereas amantadine was ineffective, up to lethal doses (table 1). Benztropine inhibited the LMR only at a nearlethal dose (3 mg/kg) which produced marked hypotension (table 1). Apomorphine was a very potent antagonist of the LMR, being effective in a dose range of 0 . 0 3 - 0 . 1 mg/kg (table 1). 1-Dopa was only about 1/300th as potent as apomorphine. In order to determine whether apomorphineinduced inhibition of the LMR was due to an effect on dopamine receptors, haloperidol, a relatively specific dopanfine antagonist (Janssen, 1967; Van Rossum, 1967; Yeh et al., 1969) was administered 10

Table 1 Effect of antiparkinson drugs on the linguomandibular reflex (LMR) in cats. Treatment

ni

% Reduction of the LMR (mean +- S.E.M.) Dose (mg/kg, i.v.) 0.03

~-Dopa Apomorphine Amantadine Benztropine

13 7 6 4

0.l

0.3

1.0

3.0

. . . . 5.5 -+ 5.4 37.7+11.3 77.8+12.2 . . . . . . . . 0.8 -+ 0.8 0 + 0 0 -+ 0 0-+0 28.2+13.9 66.73-+33.3

10.0

30.0

38.3_+10.5 83.5-+7.9 . 9.7-+ 9.7 -2 4 _

1 Number of cats used. 2 3 of 6 cats receiving amantadine, 30 mg/kg, died almost immediately after dosing; the other 3 cats had marked hypotension. 3 This dose of benztropine produced marked hypotension (an average decrease in mean blood pressure of 54 ram). 4 3 of 4 cats died after this dose.

180

A. Barnett, J. W. Ftore, Effect o f antiparkinson drugs on the LMR

Table 2 Effect of haloperidol and phenoxybenzamine on ~-dopa-induced inhibition of the LMR. Treatment

Reduction of the LMR (% of control) (Mean -+S.E.M.) Dose (mg/kg, i.v.)

n 1

l-Dopa Haloperidol, 0.3 mg/kg, + Q-dopa Phenoxybenzamine, 3.0 mg/kg, + ~-dopa

13 8 4

3

10

30

5.5 ± 5.4 10.8 -+4.5 4.2 ± 4.2

38.3 _+10.3 45.0 -+ 19.0 47.7 ± 17.8

81.7 -+ ll.1 88.5 -* 9.3 100.0± 0.0

I Number of cats used. Table 3 Inhibition of the LMR by metabolites of l-dopa, dopamine and norepinephrine. Treatment (Number of cats used)

Reduction of the LMR (% of control) (Mean ± S.E.M.) Dose (mg/kg, i.v.) 1

1-Dopa (13) 3-Methox~dopa (3) Homovanillic acid (5) Vanillylmandelic acid (4) Dihydroxyphenylacetic acid (4) 3-Methoxytyramine (4)

0-+0 0±0

3

10

30

60

100

5.5-+ 5.4 12.1± 12.1 5.0 +- 5.0

38.3-+10.5 0 -+ 0 12.0+_ 8.0 0 -+ 0 12.5 -+ 12.5 58.0-+ 1 6 . 2

81.7_+11.1 0 -+ 0 33.3± 2 1 . 1 32.7 _+1 8 . 0 13.2 -+ 13.2 100.0-+ 0

o± 0 31.7-+20.4 15.9-+15.9 33.3 -+ 33.3

0 -+ 0 55.9-+21.4 57.6 -+ 18.9 -

min before obtaining a dose-response curve with apomorphine. Haloperidol, 0.1 and 0.3 mg/kg, shifted the dose-response curve for apomorphine-induced inhibition of the LMR to the right in a parallel manner (fig. 1). Phenoxybenzamine, an c~-adrenergic blocking agent, did not affect the dose-response curve for apomorphine (fig. 1). Haloperidol, 0.3 mg/kg, which markedly antagonized apomorphine, had no effect on 1-dopa-induced inhibition on the LMR (table 2). As indicated in table 2, phenoxybenzamine did not antagonize l-dopa. Thus the effect of 1-dopa on the LMR was not affected by either haloperidol, a dopamine antagonist or by phenoxybenzamine, an aadrenergic blocking agent. These results suggested that 1-dopa may have an effect of its own or may act via the formation of a different metabolite in the brain. For this reason, the major metabolites of 1dopa, dopamine and norepinephrine which have been reported, 3-methoxydopa, homovanillic acid, vanillylmandelic acid, dihydroxyphenylacetic acid and 3-

methoxytyramine (Kaser, 1970; Goodall and Alton, 1968, 1969; Curzon et al., 1970) were tested for their effects on the LMR. These metabolites were tested i.v. so that they could be compared to our 1-dopa results. None of the metabolites tested were more potent than 1-dopa and the only one that even approximated the potency of l-dopa was 3-methoxytyramine, which was equipotent to 1-dopa (table 3). However, 3-methoxytyramine produced moderate pressor effects at doses which inhibited the LMR, whereas l-dopa did not. The least active of the metabolites tested was 3-methoxydopa which did not affect the LMR in doses as high as 100 mg/kg. Since 3-methoxydopa is a major metabolite of 1-dopa (Kuruma et al., 1970) formed via a reaction which is catalyzed by the enzyme, catechol-O-methyltransferase (COMT), a COMT inhibitor, tropolone, was used in combination with 1-dopa. Tropolone, 5 and 10 mg/kg, administered 10 rain before rising doses of l-dopa, potentiated the inhibitory effects of 1-dopa on the LMR

A. Barnett, J. W. Fiore, Effect o f antiparkinson drugs on the LMR

1oo --

~

4. Discussion

I I I CONTROL APOMORPHfNE 0 . ~ m m 0 HALOPERIDOL b l m~lkq 4- APOMORPHINE A ~ & HALOPERID~JL O 3mq/kg + ~OMORPHINE •~oiI PHENOXYBENZAMINE 1 0 mg/kg 4- APOMORPHINE

UO

~,~o~.l~

.//',.4 .,./

CO

g

. ..-.-"

l///-

g 4O

~3

/ O 03

/

Oil

0.3

I IO

DOSE OF APOMO~PHINE (mglkg, iIv.)

Fig. 1. Effect of haloperidol and phenoxybenzamine on apomorphine-induced inhibition of the LMR. Each point represents the mean -+ S.E.M. of 7 cats for apomorphine alone, 4 cats for haloperidol (0.l mg/kg) + apomorphine, 5 cats for haloperidol (0.3 mg/kg) + apomorphine and 4 cats for the phenoxybenzamine + apomorphine experiments. • • O~le~O ~ &

L'DOPA CONTROL TROPOLONE 5 rng/kg + L -DOPA TROPOLONE I0 mglkg + L'DOPA

/'/t

~00'

-J o

Bc

~ 6c

I

g 4o

,/

I/

I

I

r., ~

20

o

--///

I 3

6 10 DOSE OF L'DOPA (mglkg, i.v.)

181

I

I

30

60

Fig. 2. Effects of tropolone on l-dopa-induced inhibition of the LMR. Each point represents the mean -+S.E.M. of 13 cats for 1-dopa alone, and 6 cats for each dose of tropolone in combination with 1-dopa.

(fig. 2). These same doses of tropolone did not significantly ( p > 0.05, Probit Maximum likelihood method) potentiate apomorphine-induced inhibition of the LMR.

L-Dopa and apomorphine produced dose-related inhibition of the LMR but because of the ineffectiveness of amantadine, the relevance of this model for detecting potential antiparkinson activity is uncertain. Apomorphine-induced inhibition of the LMR was blocked by low doses of haloperidol, but not by phenoxybenzamine. From these results it can be hypothesized that stimulation of dopamine receptors in the CNS produces LMR inhibition. Apomorphine has been reported to depress polysynaptic reflexes in the spinal cord (Schlosser et al., 1970), but the doses used were at least 100 times greater than in the present study. 1-Dopa also produced LMR inhibition, but its effects were not blocked by haloperidol or phenoxybenzamine. This dose of phenoxybenzamine completely blocked the pressor effects of norepinephfine but there is no evidence specifically in cats which proves that central nervous system norepinephrine receptors are blocked by this dose. However, phenoxybenzamine was used in the highest dose which did not lower blood pressure. From these results it is unlikely that 1-dopa-induced inhibition of the LMR in cats is due to the formation of dopamine or norepinephrine in the CNS. 1-Dopa may be converted to an active metabolite or may have a direct action of its own. None of the known catecholamine metabolites were more potent than 1-dopa making the former possibility unlikely. The ability of many of the metabolites to cross the blood-brain barrier has been determined, including 3-O-methyldopa (Bartholini et al., 1971) dihydroxyphenylacetic acid (Ericsson and Wertman, 1971) and presumably 3-methoxytyramine, since it inhibited the LMR. Thus the fact that the metabolites were administered i.v. may not be a problem. The least active of the metabolites tested was 3-methoxydopa. It has been suggested that 3-O-methyldopa gives rise to dopamine in the CNS (Bartholini et al., 1971) but in the present work, we have found no evidence to support the hypothesis. Since tropolone, which presumably interferes with 3-methoxydopa formation, potentiated the effects of l-dopa, and haloperidol or phenoxybenzamine did not antagonize l-dopa, it is likely that l-dopa inhibits the LMR at least in part via a direct action of its own. Work is in progress to substantiate this point by examining the

182

A. Barnett, J. W. Fiore, Effect o f antiparkinson drugs on the LMR

effects o f d o p a d e c a r b o x y l a s e L M R - d e p r e s s a n t e f f e c t o f 1-dopa.

inhibitors

on

the

Acknowledgments The authors are grateful to Roche Laboratories for providing a generous supply of 3-methoxydopa.

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