Antagonism of AP-5- and amphetamine-induced behaviour by timelotem as compared with clozapine and haloperidol

Bilateral intrastriatal injection of DL-2-amino-5-phosphonovaleric acid (AP-5), that blocks glutamatergic transmission at the N-methyl-d-aspartate preferring receptor, induces sniffing and body turns and reduces grooming in rats. Timelotem, a representative of the newly developed chemical class of anellated benzodiazepines antagonized specifically AP-5-induced sniffing and body turns. Classical (haloperidol) as well as atypical (clozapine) neuroleptics had recently been shown to antagonize AP-5-induced sniffing; clozapine, like timelotem, but not haloperidol, additionally antagonized AP-5-induced body turns. Further, timelotem antagonized amphetamine-induced stereotyped behaviour in rats, but was found less active than haloperidol in this test. Comparing the activity of drugs in both paradigms revealed that haloperidol inhibited AP-5-induced sniffing and amphetamine-induced stereotypies within the same dose range, but timelotem and clozapine were found more potent in the AP-5 test than in the amphetamine test. Thus, detailed drug profiles discriminate timelotem and clozapine from haloperidol, linking timelotem again to atypical antipsychotic compounds.     

Decreased antistereotypic effect of neuroliptics after additional treatment with a benzodiazepine, a GABA agonist or an anticholinergic compound.

Neuroleptics are very potent antagonists against stereotypies induced by DA-stimulants including methylphenidate. This effect of neuroleptics is usually related to the antipsychotic effect of these compounds. In contrast we found that GABA agonists potentiate stereotyped gnawing induced by methylphenidate. The GABA agonist muscimol in combination with neuroleptics will attenuate the antagonistic effect of these compounds on stereotyped gnawing induced by methylphenidate. However a differentiation between the neuroleptic drugs was found: Haloperidol, spiroperidol and pimozide were profoundly antagonized by muscimol whereas cis(Z) - flupenthixol and fluphenazine were less antagonized. Baclofen shows no significant effect. Diazepam and scopolamine also strongly antagonized the antistereotypic effect of the butyrophenone-like compounds whereas only scopolamine could antagonize fluphenazine and cis(Z) - flupenthixol. Therefore we conclude that if the antistereotypic effect of neuroleptics correlates to the antipsychotic effect, a GABA agonist would probably not potentiate the antipsychotic effect of neuroleptics but rather antagonize it.     

NE-100, a novel sigma receptor ligand: In vivo tests

It has been suggested that sigma receptor antagonists may be useful as antipsychotic drugs. N, N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride (NE-100) is a novel compound with high affinity for the sigma receptor (IC50 = 4.16 nM), but low affinity (IC50 > 1000 nM) for D1, D2, 5-HT1A, 5-HT2 and phencyclidine (PCP) receptors. The head-weaving behavior induced by either (+)SKF10047 or PCP was dose-dependently antagonized by NE-100 with oral ED50 at 0.27 and 0.12 mg/kg, respectively. NE-100 did not affect dopamine agonists-induced stereotyped behavior and/or hyperactivity. NE-100 failed to induce catalepsy in rats. These findings indicate that NE-100 may have antipsychotic activity without the liability of motor side effects typical of neuroleptics.     

Presynaptic control of dopamine metabolism in the nucleus accumbens. Lack of effect of buspirone as demonstrated using in vivo voltammetry

Buspirone is a non-benzodiazepine drug with anxiolytic properties. It has been reported to induce a marked increase in the metabolism of dopamine in the striatum and the nucleus accumbens which is similar to that induced by neuroleptics. It has been suggested that the effect observed in the striatum reflects an action of buspirone on dopaminergic autoreceptors in both terminals and cell bodies. In the present study, presynaptic effects of buspirone on dopaminergic metabolism in the nucleus accumbens were investigated, and they were compared to the effects of the classical neuroleptic, haloperidol. Dopaminergic terminals were isolated by infusion of tetrodotoxin into the median forebrain bundle in order to evaluate the effects of buspirone and haloperidol on presynaptic receptors. Changes in dopamine metabolism were determined by in vivo voltammetry. Buspirone administered after interruption of the impulse flow did not affect dopamine metabolism. In contrast haloperidol treatment led to an increase in metabolism of dopamine. It is concluded that buspirone did not act at the presynaptic level and furthermore on dopaminergic autoreceptors.     

Differential effects of central serotonin manipulation on hyperactive and stereotyped behaviour

The locomotor response following injection of dopamine into the nucleus accumbens was attenuated by the injection of 5HT and potentiated by the injection of methysergide into the same site. D-amphetamine-induced locomotor activity was also reduced by the intra-accumbens injection of 5HT. In contrast, apomorphine- induced stereotyped behaviour (sniffing, licking, biting, gnawing) was reduced by systematic administration of the putative 5HT receptor antagonists, cyproheptadine and metergoline. In addition the low intensity sniffing responses produced by a low dose of apomorphine were converted to high intensity biting, gnawing or licking by the putative 5HT receptor agonist, quipazine or the putative 5HT uptake blocker, ORG 6582. The selective induction of either hyperactive or stereotyped behaviour may therefore be influenced by the functional state of central serotonergic systems.     

In vitro and in vivo pharmacological profile of 5-[2-[4-(6-fluoro-1H-indole-3-yl)piperidin-1-yl]ethyl]-4-(4-fluorophenyl)thiazole-2-carboxylic acid amide (NRA0562), a novel and putative atypical antipsychotic

In vitro and in vivo pharmacological properties of 5-[2-[4-(6-fluoro-1H-indole-3-yl)piperidin-1-yl]ethyl]-4-(4-fluorophenyl)thiazole-2-carboxylic acid amide (NRA0562), a novel atypical antipsychotic, were investigated. NRA0562 showed high affinities for human cloned dopamine D(1), D(2), D(3) and D(4) receptors with Ki values of 7.09, 2.49, 3.48 and 1.79 nM. In addition, NRA0562 had high affinities for the 5-HT(2A) receptor and the alpha(1) adrenoceptor with Ki values of 1.5 and 0.56 nM, and moderate affinity for the histamine H(1) receptor. Using in vivo and ex vivo receptor binding studies in rats, we showed NRA0562 occupied frontal cortical 5-HT(2A) receptors and alpha(1) adrenoceptor potently, while occupancy of striatal dopamine D(2) receptor was moderate as were other atypical antipsychotics. NRA0562 dose-dependently inhibited methamphetamine (MAP)-induced locomotor hyperactivity in rats. At higher dosage, NRA0562 dose-dependently antagonized MAP-induced stereotyped behavior and induced catalepsy dose-dependently and significantly in rats. But, the ED(50) value in inhibiting MAP-induced locomotion hyperactivity was 10 times lower than that in inhibiting MAP-induced stereotyped behavior, and 30 times lower than that in inducing catalepsy. In addition, the potency of NRA0562 in antagonizing MAP-induced hyperactivity in rats was higher than that of other antipsychotics, clozapine, risperidone and olanzapine. NRA0562 had favorable properties in view of prediction of extrapyramidal side effects. As this antipsychotic has a unique profile with affinity and occupancy for receptors, we propose that NRA0652 may have unique atypical antipsychotic activities, and a moderate liability of extrapyramidal motor side effects seen in the treatment with classical antipsychotics.     

Ligands of the NMDA receptor-associated glycine recognition site and motor behavior

Summary Motor behavior critically depends on glutamatergic functions in the basal ganglia (BG). The dorsal and ventral striatum — the main input structures of the BG - are involved in modulation of stereotyped sniffing behavior, locomotion, catalepsy and prepulse inhibition. The effects of the NMDA receptor have been well characterized in respect to motor behavior in the past. The function of the allosteric glycine site was however disregarded until now, because brain penetrating ligands were missing. The present study summarized the motor behavioral profile of several glycine site ligands (7chlorokynurenate, ACEA 1021, MRZ-2/576, (+) HA-966, D-cycloserine and felbamate). It is shown that through blockade of the glycine site of the NMDA receptor a distinct behavioral profile can be obtained.     

Stereotypies,aggression and the feeding schedules of tethered sows

Thirty tethered sows were observed for 5 min every half-hour for 9 h spanning the two feeding periods. Activity, consisting largely of food searching behaviour and drinking, was largely restricted to two 2-h periods following each feed. Three categories of stereotyped behaviour were observed and these were closely tied to the feeding periods. Short-duration bouts of rubbing, head-waving and bar-biting occurred during food delivery, while long-duration bouts of highly stereotyped and idiosyncratic sequences of rubbing and drinking were shown by older sows immediately after feeding. Vaccuum chewing tended to occur slightly later. I suggest that frustration of feeding motivation rather than under-stimulation underlies stereotypies in pigs, and that the different forms may represent stereotype of the appetitive and consummatory phases. Aggression was rare and was not closely related to the feeding periods or to stereotypies.     

Tolerance in the anxiolytic profile following repeated administration of diazepam but not buspirone is associated with a decrease in the responsiveness of postsynaptic 5-HT-1A receptors

To understand the role of serotonin (5-hydroxytryptamine; 5-HT)-1A receptors in the treatment of anxiety and the development of tolerance to benzodiazepines the present study was designed to monitor the responsiveness of postsynaptic 5-HT-1A receptors following repeated administration of diazepam and buspirone. Results show that tolerance in the anxiolytic profile is produced following repeated administration (2 weeks) of diazepam (2 mg/kg) but not buspirone (0.5 mg/kg). The behavioral effects of 8-OH-DPAT at a dose of 0.25 mg/kg were monitored 3 days after repeated administration of saline or buspirone or diazepam. The results show that 8-OH-DPAT elicited forepaw treading was smaller in repeated diazepam but not repeated buspirone injected rats, while hyperlocomotive effects of 8-OH-DPAT were smaller in both repeated buspirone and repeated diazepam injected rats. The results suggest that postsynaptic 5-HT-1A receptor-dependent responses were attenuated following long-term administration of diazepam but not buspirone. Role of 5-HT-1A receptors in the development of tolerance to the anxiolytic effects of diazepam but not buspirone is discussed.     

Effects of dopaminergic agonists and antagonists on cerebellar guanosine-3',5'-monophosphate (cGMP).

Apomorphine was found to cause an increase in cerebellar cGMP content. Bromocriptine, at a dose that caused stereotypies, neither elevated cGMP, nor blocked the apomorphine- induced rise in cGMP. The apomorphine-induced rise in cGMP was effectively blocked by haloperidol and some other neuroleptics, but not by sulpiride. These actions of the neuroleptics correlated with their ability to displace ³H-spiroperidol from striatal membranes, suggesting that dopamine receptor interactions were important in the cGMP changes noted. Based on the observation that haloperidol antagonized the increase induced by restraint, it is suggested that dopaminergic systems are involved in the reaction to stress.     

Anxiolytic profile of girisopam and GYKI 52,322 (EGIS 6775). Comparison with chlordiazepoxide and buspirone.

The anxiolytic action of two 2,3-benzodiazepines: girisopam: GYKI 51,189 (EGIS 5810): (1-(3-chlorophenyl)-4-methyl-7,8-dimethoxy-5H-2,3-benzodiazepine), and GYKI 52,322 (EGIS 6775): (1-(4-aminophenyl)-4-methyl-7,8-dimethoxy-5H-2,3-benzodiazepine) was investigated in comparison to chlordiazepoxide and buspirone using three different animal models of anxiety: the lick conflict, the elevated plus maze and the open field methods in rats. Both 2,3-benzodiazepines exerted anxiolytic effect in all three tests used, however their pharmacological profile differs considerably from that of either chlordiazepoxide or buspirone. Using the animal models mentioned above the order of potency was GYKI 52,322 (EGIS 6775) > chlordiazepoxide > girisopam > buspirone.     

A selective dopamine D4 receptor antagonist, NRA0160: a preclinical neuropharmacological profile

NRA0160, 5 - [2- ( 4- ( 3 - fluorobenzylidene) piperidin-1-yl) ethyl] - 4 -(4-fluorophenyl) thiazole-2-carboxamide, has a high affinity for human cloned dopamine D4.2, D4.4 and D4.7 receptors, with Ki values of 0.5, 0.9 and 2.7 nM, respectively. NRA0160 is over 20,000fold more potent at the dopamine D4.2 receptor compared with the human cloned dopamine D2L receptor. NRA0160 has negligible affinity for the human cloned dopamine D3 receptor (Ki=39 nM), rat serotonin (5-HT)2A receptors (Ki=180 nM) and rat alpha1 adrenoceptor (Ki=237 nM). NRA0160 and clozapine antagonized locomotor hyperactivity induced by methamphetamine (MAP) in mice. NRA0160 and clozapine antagonized MAP-induced stereotyped behavior in mice, although their effects did not exceed 50% inhibition, even at the highest dose given. NRA0160 and clozapine significantly induced catalepsy in rats, although their effects did not exceed 50% induction even at the highest dose given. NRA0160 and clozapine significantly reversed the disruption of prepulse inhibition (PPI) in rats produced by apomorphine. NRA0160 and clozapine significantly shortened the phencyclidine (PCP)-induced prolonged swimming latency in rats in a water maze task. These findings suggest that NRA0160 may have unique antipsychotic activities without the liability of motor side effects typical of classical antipsychotics.     

Effects of chronic buspirone treatment on female mice exposed to the long-lasting psychoemotional influence

The effects of chronic treatment (30 days) with the 5-HT1A receptor partial agonist buspirone (0.05, 1 and 10 mg/kg i.p.) on the behaviour of C57BL/6J female mice exposed to long-lasting psychoemotional influence were studied. The influence involved forced living of each female with an aggressive male separated with a perforated transparent partition in the same cage and daily female's presence during 10-min intermale confrontations behind a partition caused by introducing of another male to the aggressive male. Chronic buspirone injection (in all used doses) did not affect the behaviour of females estimated in the "partitions" and "open field" tests at the end of the drug treatment. The anxiolytic effect of buspirone only at the dose of 1 mg/kg on the female's plus-maze behaviour was revealed. In the Porsolt, test buspirone in the dose of 1 mg/kg caused a slight increase in the duration of immobility indicating a slight pro-depressive effect. Thus, chronic buspirone treatment of females exposed to the long-lasting psychoemotional impact has a different effect on their behaviour depending on the dose and test conditions.     

Behavioural effects of N-methyl-D-aspartate in the anterodorsal striatum of the rat

N-methyl-D-aspartate (NMDA) (0.5 and 1 microgram/0.5 microliter) bilaterally injected into the anterodorsal striatum of rats reduced locomotion, sniffing, rearing and feeding upon presentation of palatable food. Consequently, the number of all behavioural bouts exhibited was reduced and the duration of akinetic phases was prolonged. These results are discussed in connection with previous findings showing that the NMDA receptor blocker DL-2-amino-5-phosphonovaleric acid (AP-5) injected at the same site - produced opposite effects: AP-5 enhanced locomotion, rearing, sniffing as well as the total number of behavioural bouts exhibited.     

Concomitant D1 dopamine receptor activation (SKF 38393) unmasks D2 dopaminergic actions of B-HT 920 in mice.

The present study attempts to demonstrate D1/D2 dopamine (DA) receptor interactions during stereotyped behaviour in mice. B-HT 920 [2-amino-6-allyl-5, 6, 7, 8-tetrahydro-4H-thiazolo-(4, 5-d)-azepine] (0.05-1.0 mg/kg), a selective D2-DA agonist, induced mild per se stereotypy consisting mainly of sniffing and rearing responses. Apomorphine, a mixed D1/D2 agonist, also produced typical stereotypic response in mice. The stereotypic response of B-HT 920 was blocked by D2-DA antagonist, sulpiride (50 mg/kg). The effect of apomorphine was not influenced by co-treatment with SKF 38393. Simultaneous administration of B-HT 920 (0.1-0.5 mg/kg) with SKF 38393 (5 mg/kg), a selective D1-DA agonist, elicited dramatic increase in stereotyped behaviours in naive as well as in 24 hr reserpinised (2 mg/kg) mice. Co-treatment of apomorphine (0.5 mg/kg) with B-HT 920 (0.1, 0.25 mg/kg) also resulted in a clearly synergistic effect on stereotyped behaviour. These potentiated responses were reduced or blocked by haloperidol, a D2-DA antagonist. The data suggest that in presence of concomitant stimulation of D1-DA receptors. B-HT 920 exhibits full expression of postsynaptic D2-DA receptor mediated behavioural effects.     

SCH 23390 dissociated from conventional neuroleptics in apomorphine climbing and primate acute dyskinesia models

SCH 23390 induced only a negligible incidence of the acute dyskinetic syndrome, a predictor of neuroleptic-induced extrapyramidal liability, in squirrel monkeys. However, haloperidol-induced dyskinesias were potentiated by SCH 23390 and were blocked by the D-1 agonist, SKF 38393. When administered orally or intraperitoneally to mice, SCH 23390 showed a considerably wider dose separation than did conventional neuroleptics between antagonism of apomorphine climbing and antagonism of stereotyped sniffing. Clinically relevant distinctions may exist between D-1 and D-2 antagonists, with D-1 antagonists (exemplified by SCH 23390) showing lower, although possibly not negligible, potential to cause extrapyramidal side effects.     

N-methyl-D-aspartate (NMDA)-mediated corticotropin-releasing factor (CRF) release in cultured rat amygdala neurons

Corticotropin-releasing factor (CRF) plays an important role in the activation of centrally mediated responses to stress. The amygdala, a limbic structure involved in the stress response, has a significant number of CRF cell bodies and CRF receptors. Activation of glutamatergic projections to the amygdala has been implicated in the stress response. Few studies have evaluated neurotransmitter-stimulated CRF release in the amygdala. We measured the effects of glutamate (0.1-1000 microM) and N-methyl-D-aspartate (NMDA, 0.1-1000 microM) on CRF release from the amygdala using primary neuronal cultures from embryonic rat brains (E18-19). Experiments were performed after the cultures grew for 17-20 days. CRF was measured using radioimmunoassay. The excitatory amino acid neurotransmitters, glutamate and NMDA, stimulated CRF release in a concentration-dependent manner. The apparent EC50 values for glutamate and NMDA were 17.5 microM and 12 microM, respectively. Consistent with a NMDA receptor-driven event, glutamate-stimulated CRF release was blocked by the NMDA antagonist, 2-amino-5-phosphonovaleric acid (AP-5, 1-100 microM) and antagonized by the addition of 1.2 mM MgCl2 to the incubation medium. These results implicate an inhibition of CRF release in the amygdala as a possible mechanism for the reported anxiolytic effects of NMDA antagonists.     

In vitro and in vivo pharmacological profile of 4-(4-fluorobenzylidene)-1-[2-[5-(4-fluorophenyl)-1H-pyrazol-4-yl] ethyl] piperidine (NRA0161)

Atypical antipsychotic properties of 4-(4-fluorobenzylidene)-1-[2-[5-(4-fluorophenyl)-1H-pyrazol-4-yl]ethyl] piperidine (NRA0161) were investigated by in vitro receptor affinities, in vivo receptor occupancies and findings were compared with those of risperidone and haloperidol in rodent behavioral studies. In in vitro receptor binding studies, NRA0161 has a high affinity for human cloned dopamine D(4) and 5-HT(2A) receptor with Ki values of 1.00 and 2.52 nM, respectively. NRA0161 had a relatively high affinity for the alpha(1) adrenoceptor (Ki; 10.44 nM) and a low affinity for the dopamine D(2) receptor (Ki; 95.80 nM). In in vivo receptor binding studies, NRA0161 highly occupied the 5-HT(2A) receptor in rat frontal cortex. In contrast, NRA0161 did not occupy the striatal D(2) receptor. In behavioral studies, NRA0161, risperidone and haloperidol antagonized the locomotor hyperactivity in mice, as induced by methamphetamine (MAP). At a higher dosage, NRA0161, risperidone and haloperidol dose-dependently antagonized the MAP-induced stereotyped behavior in mice and NRA0161 dose-dependently and significantly induced catalepsy in rats. The ED(50) value in inhibiting the MAP-induced locomotor hyperactivity was 30 times lower than that inhibiting the MAP-induced stereotyped behavior and 50 times lower than that which induced catalepsy.These findings suggest that NRA0161 may have atypical antipsychotic activities yet without producing extrapyramidal side effects.     

The Paw test: an animal model for neuroleptic drugs which fulfils the criteria for pharmacological isomorphism

Recently we have developed an animal model which could discriminate between classical and atypical neuroleptic drugs: the PAW TEST. This test measures the ability of rats to spontaneously withdraw its force- and hindlimbs. It was found that the ability of drugs to affect the rat's forelimb retraction time was associated with the liability of the drug to induce so-called extra-pyramidal side-effects in man. Likewise the ability of drugs to affect the rat's hindlimb retraction time was associated with the antipsychotic efficacy of the drug. These data open the perspective that the forelimb retraction time (FRT) is an animal analogue of parkinsonian side-effects, and that the hindlimb retraction time (HRT) is an animal analogue of antipsychotic effects In the present series of experiments we further evaluated the validity of these notions by applying the criteria of "pharmacological isomorphism" as proposed by Matthysse (1). Thus HRT had to fulfil the criteria of pharmacological isomorphism for the therapeutic effects of neuroleptics, whereas FRT had to fulfil these criteria for the parkinsonian side-effects. The results of the present study show that both FRT and HRT met these criteria. Thus both classical and atypical neuroleptics were effective in prolonging HRT, whereas only the classical neuroleptics prolonged FRT (criterion 1); the nonneuroleptic phenothiazine promethazine (as well as the narcotic morphine, the muscle relaxant diazepam and the antidepressant desipramine) were ineffective in this respect (criterion 2); the acetylcholinergic antagonist scopolamine blocked the FRT, but not the HRT (criterion 3); chronic neuroleptic treatment reduced the FRT, but not the HRT (criterion 4). In conclusion the paw test, an animal model for testing antipsychotic drugs, was found to fulfil the criteria for "pharmacological isomorphism". Although the exact mechanism underlying the paw test is as yet unknown, the present data improve its validity as an animal model.     

Two new phenylpiperazines with atypical antipsychotic potential

Two new series of substituted arylpiperazines with heterocyclic 3-propoxy-benzimidazole or 3-propoxy-benzimidazole-2-thione groups were synthesized and their in vitro binding affinities for the D(2), 5-HT(1A), 5-HT(2A), and alpha(1)-adrenergic receptors determined. Among them, only two compounds with phenyl aryl-constituent (8a and 9a) showed 5-HT(2A)/D(2) pK(i) binding ratios proposed for atypical neuroleptics. As to their behavioral screening on rodents, both compounds exhibited a non-cataleptic action in rats and antagonized D-amphetamine-induced hyperlocomotion in mice, suggesting their possible atypical antipsychotic potency.