The effect of serotonergic agents on haloperidol-induced catalepsy

Life Sciences, Vol. 47, pp. 1609-1615 Printed in the U.S.A.

Pergamon Press

THE EFFECT OF SEROTONERGIC AGENTS ON HALOPERIDOL-INDUCED CATALEPSY Paul B. Hicks Scott and White Clinic, Department of Psychiatry Scott and White Memorial Hospital; Scott, Sherwood and Brindley Foundation Texas A&M University College of Medicine Temple, Texas 76508 (Received in final form August 21, 1990)

$.ammar~ The effect of various classes of serotonergic agents on haloperidol-induced catalepsy was evaluated in male Sprague-Dawley rats. The 5-HT-1A agonists buspirone, ipsapirone and 8-OH-DPAT all potently reversed catalepsy. The mixed 5HT-1A and 5-HT-1B agonist RU 24969 reversed catalepsy only at the highest dose tested. The non-selective 5-HT- 1 antagonist (1)-propranolol did not affect catalepsy. The 5-HT-2 agonist DOI and 5-HT-2 antagonist mesulergine both reversed catalepsy. ICS 205-930 (5-HT-3 antagonist) reversed catalepsy at low doses only. Another 5HT-3 antagonist, GR 38032F, had no effect on catalepsy. These studies suggest that 5-HT-1A and 5-HT-2 receptor sites are important in the serotonergic modulation of haloperidol-induced catalepsy. The dopamine hypothesis of schizophrenia suggests that the positive symptoms of schizophrenia are the result of hyperactive mesolimbic and mesocortical dopaminergic systems. The drugs that have been developed to control the symptoms of schizophrenia oppose brain dopamine systems, but neuroleptics are not selective to the mesolimbic and mesocortical dopaminergic systems. Consequently, the antipsychotic medications currently available in the United States all have the major problem of causing extrapyramidal syndromes (akathisia, dystonia, psuedoparkinsonism and tardive dyskinesia). There is substantial evidence documenting that medial and dorsal raphe serotonergic neurons regulate striatal, mesolimbic and mesocortical dopamine systems. This regulatory effect is thought to be predominantly inhibitory, based upon the observations that lesions of the medial raphe nuclei produce a marked hyperactivity that is suppressed by serotonin injections into the nucleus accumbens (1) and that full expression of stereotypic behaviors is dependent upon suppression of low intensity stereotypy by serotonergic systems (2,3). In rats with lesions of the dorsal and medial raphe nuclei, butyrophenone and phenothiazine-induced catalepsy is reduced (4,5) and there is a simultaneous reduction in the 5-HT content of striatum, mesolimbic and cortical brain regions (6). Also, the serotonin antagonist methysergide opposes haloperidol-induced catalepsy (HIC), but the serotonin agonist quipazine or serotonin uptake inhibitor clomipramine potentiate HIC (7). Finally, the serotonin synthesis inhibitor para-chlorophenylalanine opposes the activity of haloperidol to increase striatal, mesolimbic or frontal cortex dopamine turnover (5,6) and opposes HIC (5,7). Despite the knowledge of this physiological and pharmacological relationship between the serotonergic and dopaminergic systems there have been no treatment paradigms developed for psychosis or neuroleptic-induced extrapyramidal syndromes which utilize this relationship. 0024-3205/90 $3.00 +.00 Copyright (c) 1990 Pergamon Press plc

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In recent years several serotonin receptor subtypes have been identified in brain and various peripheral tissues (8-11). Of these, the 5-HT-1A, -1B, -1C, -1D, -2 and -3 receptor subtypes have been identified in rat brain. With the identification of these specific brain serotonin receptor sites, it becomes important to carefully evaluate the role of serotonergic systems in modulation of dopaminemediated rat behaviors. It may be that only specific receptor subtypes are associated with modulation of the dopaminergic system. Along with the identification of serotonin receptor subtypes, specific agonists and antagonists have been developed. The level of specificity of the agonists and antagonists is significantly better than was available when prior studies were performed on characterization of the interaction of serotonergic drugs on dopamine-mediated behaviors. The following studies were conducted to evaluate the effect of various classes of serotonergic agents on haloperidol-induced catalepsy. Materials and Methods

Drues. 1-(2,5-dimethoxy-4-iodo)-2-aminopropane (DOI), 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) and (1)-propranolol were obtained from Research Biochemicals (Waltham, Mass.). All other drugs were obtained from their respective manufacturers: buspirone (Bristol-Myers); GR 38032F (Glaxo); haloperidol (McNeil); ICS 205-930 and mesulergine (Sandoz); ipsapirone (Miles); RU 24969 (Roussel-Uclaf). Haloperidol was dissolved in a small quantity of lactic acid and diluted in 0.9% saline. Buspirone, ipsapirone, GR 38032F, (1)-propranolol and RU 24969 were dissolved in 0.9% saline. Mesulergine and ICS 205-930 were dissolved in 10% ethanol. Animals. Male Sprague-Dawley rats weighing 200-250g were obtained from Sasco King, Inc. (Omaha, Nebraska). Rats were housed for a minimum of three days prior to behavioral studies with free access to food and water and maintained on a 12 hr on/off (0700-1900 hours) light cycle at constant temperature. Catalepsy Studie~. All catalepsy experiments were performed between 1200 and 1700 hours using the horizontal bar method (4). Haloperidol (0.75 mg/kg) was administered subcutaneously (s.c.) at time zero. This dose of haloperidol was chosen to produce only a moderate degree of catalepsy so that reversal or potentiation of catalepsy could be measured. At 90 minutes the test drugs were administered s.c. Catalepsy was assessed by placing the front limbs of the rats over a horizontal bar with the hind limbs extended and abducted. The time that the rat maintained this posture was recorded. Catalepsy was scored from 0 to 5 as follows: score 0 (< 15 see), score 1 (16-115 sec), score 2 (116-215 sec), score 3 (216-315 sec), score 4 (316-415 sec) and score 5 (>415 sec). Following haloperidol administration, rats were given three trials on the horizontal bar every 30 minutes for 3 hours for the presence of catalepsy. Catalepsy was measured with the observer blinded to the treatments given for each rat. The maximum catalepsy scores for each time interval were used for statistical comparisons. Dose-response relationships for the effect on haloperidol-induced catalepsy were determined for the drugs listed in Table I. Each substance was tested at 3 doses with a minimum of 10 rats tested for each dose. Doses of each drug were chosen that had been shown to be behaviorally active in other paradigms. For the 5-HT-1A agonists, the maximum effect was seen at the lowest dose studied. For each drug a control group was used that received the highest dose of the test drug without haloperidol. Catalepsy was not observed in any of these control groups; and they were, therefore, excluded from the statistical analyses. Also, for each drug a second control group received haloperidol and the test drug vehicle only at the highest dose. For each test drug the mean maximum

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catalepsy scores for 90 to 180 minutes were compared among the three dose groups and the haloperidol control group by analysis of variance with repeated measures (12) and Duncan's Multiple Range Test. TABLE I Serotonergic Drugs Studied Dm~ Buspirone Ipsapirone 8-OH-DPAT RU-24969 (L)-Propranolol 1301 Mesulergine GR38032F ICS 205-930

Serotongr~c Receptor 5-HT- 1A 5-HT- 1A 5-HT- 1A 5-HT-1A and 5-HT-1B 5-HT- 1A and 5-HT- 1B 5-HT-2 5-HT- 1C and 5-HT-2 5-HT-3 5-HT-3

Agonist/An~gonist Agonist Agonist Agonist Agonist Antagonist Agonist Antagonist Antagonist Antagonist

Results

The results are shown in Fig. 1 and Table II. None of the serotonergic drugs tested caused sedation in the doses tested. The 5-HT-1A receptor agonists buspirone, ipsapirone and 8-OH-DPAT all potently reversed HIC. The mixed 5-HT-1A and 5-HT-1B agonist RU 24969 reversed catalepsy at the highest dose only. The non-selective 5-HT-1 receptor antagonist (1)-propranolol had no effect on HIC in the dose range tested. The 5-HT-2 agonist DOI and the 5-HT-2 antagonist mesulergine both reversed HIC. ICS 205-930, a 5-HT-3 antagonist, reversed catalepsy at the two lowest doses, but not at the highest dose. Another 5-HT-3 antagonist, GR 38032F, did not affect HIC. Table II gives the significance levels for differences between each control and treatment groups for each drug tested.

TABLE II Statistical Comparisons Probability of No Dru~ Treatment Effect Buspirone <0.001 Ipsapirone <0.001 8-OH-DPA T <0.001 RU 24969 <0.05 (l)-Propranolol NS DOI <0.001 Mesulergine <0.001 ICS 205-930 <0.05 GR 38032F NS NS: Effect of treatment not significant (P>0.05). Statistical comparisons by analysis of variance with repeated measures and Duncan's Multiple Range Test.

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FIG. 1. Mean maximum catalepsy scores for each 30-minute interval up to 180 minutes for the nine serotonergic drugs tested. N= 10-14 rats for each drug dose. Arrows represent administration of test drug 90 minutes after s.c. injection of 0.75 mg/kg Haloperidol.

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Haloperidol-induced catalepsy is mediated primarily through inhibition of the DA D2 receptor (13). This study has substantiated previous reports that serotonergic agents modify HIC. Previous studies have not addressed the specificity of serotonergic receptor types in modifying HIC. The present data suggests that both the 5-HT-1A and 5-HT-2 receptors are important receptor types associated with modifying HIC. The 5-HT- 1A receptor is clearly implicated as having a role in affecting HIC. All 5-HT-1A receptor agonists evaluated (buspirone, ipsapirone and 8-OH-DPAT) potently and abruptly induced a persistent reversal of catalepsy. Other investigators have also reported reversal of neurolepticinduced catalepsy by 5-HT-1A agonists (14,16). However, the non-selective 5-HT-1 receptor antagonist (l)-propranolol did not affect catalepsy in the doses tested. In similar doses in mice racemic propranolol potentiated catalepsy (17). This effect may be related to the beta-adrenergic potency of (d)-propranolol. The 5-HT-1 agonist RU 24969 which is relatively non-selective for 5HT-1A and 5-HT-1B sites produced reversal of HIC only at the highest dose tested suggesting this is an effect related to its 5-HT-1A agonist properties. In fact, other 5-HT-1B selective drugs also fail to reverse catalepsy (14). The site of action of 5-HT-1A agonists may be the autoreceptors on raphe nuclei cell bodies (18). This is supported by two lines of evidence. Intraventricular injection of the 5-HT neurotoxin 5,7 dihydroxytryptamine destroys 5-HT pathways and blocks the anticataleptic effect of 8-OHDPAT. Also, 8-OH-DPAT applied locally to either the dorsal or median raphe nuclei potently reverses neuroleptic-induced catalepsy in a dose dependent manner. The lowest doses of the 5-HT-1A agonists evaluated were sufficient to produce maximum reversal of catalepsy. These doses were chosen because they produced the serotonergic behavioral syndrome (19). Recently, it has been shown that reversal of catalepsy by these agents is a dosedependent process (18). The 5-HT-2 receptor site also has an influence on HIC. However, interpretation of the present data is difficult because both 5-HT-2 agonists and antagonists reversed catalepsy. The 5-HT2 antagonists methysergide, cyproheptadine and mesulergine all reverse HIC (7,20). Unfortunately, all of these drugs have potent activities at other receptor sites including non-serotonergic sites (8,21): methysergide (5-HT-1A, 5-HT-1B and 5-HT-1C); cyproheptadine (antihistamine-HI) and mesulergine (5-HT-1C). In contrast, drugs that act as non-selective serotonin agonists (e.g. clomipramine and quipazine) potentiate HIC (11,17). Based upon this, a specific 5-HT-2 agonist (e.g. DOI) would have been predicted to also potentiate HIC. To the contrary, DOI produced a potent reversal of HIC. Further clarification of the effect of 5-HT-2 agonists on HIC will require use of other specific 5-HT-2 agonists (e.g. (+)-S- alpha-methyl-5-HT). The 5-HT-3 receptor site has only recently been identified in brain (22,23). The physiological significance of this serotonergic binding site may be very important in the regulation of the mesolimbic DA system (24), but there is no known function in relation to the nigrostriatal DA system (25). As discussed above, the relatively non-selective serotonin agonist quipazine potentiates catalepsy (7). A recent report demonstrates that quipazine is a potent agonist at the 5-HT-3 receptor site (22). Some degradation products of quipazine may also function as 5-HT uptake inhibitors (26). Also, the anorectic effect of quipazine is related to the 5-HT-2 receptor (27). It is unclear whether the HIC potentiating effect is secondary to interaction at the 5-HT-3 site or to its interactions at these other 5-HT sites. Two 5-HT-3 receptor antagonists were evaluated for their effect on HIC, GR 38032F and ICS 205-930. The ICS 205-930 reversed HIC at low doses but had no effect at the

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highest dose. HIC was not affected by GR 38032F which is consistent with the lack of effect on another nigrostriatal DA pathway-mediated behavior, amphetamine-induced stereotypy (25). The discrepancy between the activities of the two 5-HT-3 antagonists could be explained by the presence of 5-HT-3 receptor subtypes. Interestingly, there are proposed 5-HT-3 receptor subtypes in peripheral tissues (10,28). It may be that with further characterization of 5-HT-3 receptors in the central nervous system receptor subtypes for the 5-HT-3 receptor will be identified. Subtypes of 5HT-3 receptors would explain the differences in effects of these relatively specific 5-HT-3 antagonists on HIC. There is clearly a role of serotonin agents in modulation of HIC. This predicts that serotonergic agents may be of benefit in treatment of extrapyramidal symptoms induced by neuroleptics or Parkinson's disease. The 5-HT-1A agonist buspirone and the 5-HT-2 antagonist ritanserin have been tried in Parkinson's disease without benefit (29,30). There was, however, an antidyskinetic activity noted for ritanserin. An antidyskinetic effect has also been found with high dose buspirone (31). There have been no trials for either drug in psuedo-parkinsonism, however. Drug-induced psuedo-parkinsonism is usually seen in the presence of an intact neurological substrate. Because of the presence of an intact neurological substrate it is possible that there may be benefit of serotonergic agents in psuedo-parkinsonism not appreciated in Parkinson's disease. This hypothesis is supported by a primate study where the non-selective serotonin antagonists (mianserin and cyproheptadine) decreased while serotonin uptake inhibitors (paroxetine and CGP 6085A) increased haloperidol-induced dystonia and psuedo-parkinsonism (32). The present data would suggest that 5-HT-1A agonists and 5-HT-2 antagonists should be tested as novel agents for the treatment of acute extrapyramidal syndromes induced by neuroleptics. Acknowledeements

I would like to thank Ms. Rosario Perez-Guerra for her excellent technical assistance and Ms. Dana Evans and Keith Young, Ph.D. for editorial assistance. I would also like to thank Mark Riggs, Ph.D. and the Biostatistics Department of Scott and White Hospital who provided the statistical analysis. This work was funded by Scott and White Clinic Institutional Research Fund grant 250095-0. References

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