Apomorphine-induced Locomotor Research Articles

An ethanolic extract of Desmodium adscendens exhibits antipsychotic-like activity in mice.

Desmodium adscendens extract (DAE) is used traditionally in Ghana for the management of psychosis. The present study aimed at providing pharmacological evidence for its ethnomedical use by testing the hypothesis that an ethanolic extract of Desmodium adscendens may possess antipsychotic properties. The primary behavioral effects of DAE on the central nervous system of mice were investigated using Irwin's test paradigm. Novelty-induced and apomorphine-induced locomotor and rearing behaviors in mice were explored in an open-field observational test system. Apomorphine-induced cage climbing test in mice was used as the antipsychotic animal model. The ability of DAE to induce catalepsy and enhance haloperidol-induced catalepsy was also investigated in mice. The DAE produced sedation, cholinergic-, and serotonergic-like effects in mice when evaluated using the Irwin's test. No lethality was observed after 24 h post-treatment. The LD50 in mice was estimated to be greater than 3000 mg/kg. The DAE significantly decreased the frequency of novelty- and apomorphine-induced rearing and locomotor activities in mice. It also significantly lowered the frequency and duration of apomorphine-induced climbing activities in mice. It did not induce any cataleptic event in naïve mice but only significantly enhanced haloperidol-induced catalepsy at a dose of 1000 mg/kg. The ethanolic extract of Desmodium adscendens exhibited antipsychotic-like activities in mice. Motor side effects are only likely to develop at higher doses of the extract.

The effects of early-life stress on dopamine system function in adolescent female rats

During adolescence, many neural systems, including the dopamine system, undergo essential remodeling and maturation. It is well known that early-life stress (ELS) increases the risk for many psychopathologies during adolescence and adulthood. It is hypothesized that ELS interferes with the maturation of the dopamine system. There is a sex bias in the prevalence of stress-related mental disorders. Information regarding the effects of ELS on brain functioning in females is very limited. In the current study, maternal separation (MS) procedures were carried out to study the effects of ELS on dopamine system functioning in adolescent female rats. Our study showed that MS increased the density of tyrosine hydroxylase immunoreactive fibers in the prelimbic cortex (PLC) and nucleus accumbens (Acb). These changes were accompanied by a decrease in the level of D5 receptor mRNA and an increase in D2 receptor mRNA expression in the PLC of MS females. Conversely, D1 and D5 receptor mRNA levels were augmented in the caudate putamen (CPu), while the expression of the D3 dopamine receptor transcript was reduced in MS females. Additionally, in the Acb, MS elicited a decrease in D2 receptor mRNA expression. At the behavioral level, MS increased apomorphine-induced locomotion; however, it did not change locomotor responses to selective D1/D5 receptor agonist and attenuated D2/D3 receptor agonist-triggered locomotion. Moreover, MS decreased D1/D5 receptor agonist-induced grooming behavior. These results indicate that ELS disrupts dopamine receptor function in the PLC and basal ganglia during adolescence in females and may predispose them to psychopathologies during adolescence and adulthood.

Psychotropic Effects of an Alcoholic Extract from the Leaves of Albizia zygia (Leguminosae-Mimosoideae).

Background Albizia zygia is used in Ghanaian traditional medicine for the management of mental disorders. The present study tested the hypothesis that an extract of the leaves of Albizia zygia (AZE) may possess antipsychotic and antidepressant properties. Method The novelty- and apomorphine-induced locomotor and rearing behaviours of AZE in mice were explored in an open-field observational test system. The effects of AZE in apomorphine-induced cage climbing test, extract-induced catalepsy, and haloperidol-induced catalepsy on mice were also investigated. Lastly, the forced swimming and tail suspension tests in mice were employed to screen the possible antidepressant effects of AZE. Results AZE (100–3000 mg/kg) showed signs of central nervous system (CNS) depression under observation, with no lethality, 24 h after treatment in mice. AZE (100–1000 mg/kg) produced a significant decrease in the frequency of novelty- and apomorphine-induced locomotor activities in mice. The extract also significantly decreased the frequency and duration of apomorphine-induced climbing activities in mice. AZE, while failing to produce any cataleptic event in naïve mice, significantly enhanced haloperidol-induced catalepsy at a dose of 1000 mg/kg. However, AZE did not produce any significant antidepressant effects in the test models employed. Conclusion The extract of Albizia zygia exhibited an antipsychotic-like activity in mice.

TAAR1-dependent effects of apomorphine in mice.

G protein-coupled trace amine-associated receptor 1 (TAAR1) is expressed in several brain regions and modulates dopaminergic activity partially by affecting D2 dopamine receptor function. In vitro, the nonselective dopamine agonist apomorphine can activate mouse and rat TAAR1. The aim of the present study was to evaluate whether apomorphine activity at the rodent TAAR1 observed in in vitro studies contributes to its behavioral manifestation in mice. For this purpose, we compared the behavioral effects of a wide range of apomorphine doses in wild type (WT) and TAAR1 knockout (TAAR1-KO) mice. Apomorphine-induced locomotor responses (0.01-4.0mg/kg) were tested in locomotor activity boxes, and stereotypic behavior at 5mg/kg was tested by ethological methods. A gnawing test was used to analyze the effects of the highest dose of apomorphine (10mg/kg). No statistically significant differences were observed between TAAR1-KO and WT mice following inhibitory pre-synaptic low doses of apomorphine. At higher doses (2.0-5.0mg/kg), apomorphine-induced climbing behavior was significantly reduced in TAAR1 mutants relative to WT controls. Moreover, the lack of TAAR1 receptors decreased certain types of stereotypies (as reflected in by measures of the global stereotypy score, licking but not sniffing or gnawing) that were induced by high doses of apomorphine. These data indicate that apomorphine activity at TAAR1 contributes to some behavioral manifestations, particularly climbing, in rodents following high doses of this drug. The contribution of TAAR1 to apomorphine-induced climbing in rodents should be considered when apomorphine is used as a screening tool in the search for potential antipsychotics.

Chronic hyperdopaminergic activity of schizophrenia is associated with increased ΔFosB levels and cdk-5 signaling in the nucleus accumbens

Chronic subcortical hyperdopaminergic activity is associated with the positive symptoms of schizophrenia and is a hallmark feature of a number of animal models of the disorder. However, the molecular changes induced by increased dopaminergic activity associated with schizophrenia are not clear. Increased levels of ΔFosB have been found in association with chronic subcortical hyperdopaminergic activity following repeated cocaine or amphetamine administration. Therefore, we investigated ΔFosB signaling in a putative neurodevelopmental animal model of schizophrenia showing chronic subcortical hyperdopaminergic activity. Increased protein levels of the transcription factor ΔFosB as well as cyclin-dependent kinase-5 (cdk-5), p35, p25 and the GluR2 subunit of the AMPA glutamate receptor were observed in the nucleus accumbens (NA). Cdk-5, p35 and GlurR2 are all proteins regulated by ΔFosB, while p25 is a degradation product of p35. Increased total protein levels of cdk-5, p35 and p25 resulted in increased cdk-5 kinase activity as determined by increased phosphorylation of dopamine and adenosine regulated phosphoprotein-32 (DARPP32) at Thr75 in the NA. DARPP32 Thr75 is selectively phosphorylated by cdk-5 and phosphorylation of DARPP32 at Thr75 suppresses DARPP32 activity, a critical step in the regulation of both glutamatergic and dopaminergic activity in neurons. We also found that apomorphine-induced locomotor activity was further increased following intra-accumbens infusions of roscovitine, a cdk-5 blocker, in a dose-dependent manner. Our results indicate that chronic hyperdopaminergic activity, as seen in schizophrenia, may affect glutamate and dopamine function in the NA via ΔFosB-mediated transcriptional modulation.

Maternal separation fails to render animals more susceptible to methamphetamine-induced conditioned place preference

The maternal separation (MS) paradigm is an animal model that has been successfully used to study the long term effects of child abuse and neglect. Experiments showed that animals subjected to trauma and stress early in life display behavioural, endocrinological and growth factor abnormalities at a later stage in life, results that mirrored clinical conditions. It is apparent that adverse events early in life may affect the development and maturation of the brain negatively. The purpose of the present study was to investigate whether the abnormal brain development occurring in separated animals would also enhance the development of a preference for psychostimulant drug usage. Rats were subjected to maternal deprivation and further exposed to methamphetamine-induced conditioned place preference (CPP) which primarily measures drug reward (ventral striatum) learning and memory. Apomorphine-induced locomotor activity was also assessed to investigate the effects of methamphetamine on the dorsal (primarily locomotor activity) striatal dopaminergic system. We found that four consecutive injections of methamphetamine resulted in CPP behaviour 24 h after the 4th injection. A further four injections yielded similar CPP results and this effect lasted for at least 7 days until the third CPP assessment. These animals also had decreased ACTH and corticosterone secretions, but the prolactin levels were increased. Prior exposure to maternal separation did not have any effect on the CPP test. The ACTH and corticosterone secretions were also similarly reduced. However maternal separation decreased the release of prolactin and this reduction was not evident in the separated group that received methamphetamine. There was no significant difference in the apomorphine-induced locomotor activity of normally reared animals whether they received methamphetamine or saline. Interestingly there was a significant difference in locomotor activity between the two groups of animals that were subjected to maternal deprivation. The separated animals that received methamphetamine displayed markedly reduced locomotor activity upon apomorphine administration when compared to those that were treated with saline. Taken together, we conclude that maternal deprivation differentially influences dorsal and ventral striatal regions implicating dopaminergic mechanisms.

Influence of Morphine and Dopamine Receptor Sensitization on Locomotor Activity in Mice

In the present study, the influence of morphine- and dopamine receptor antagonists-induced sensitization on morphine-induced locomotion in mice was investigated. Morphine (30, 40 and 50 mg/kg) increased, while lower doses of the opioid (10 and 20 mg/kg) decreased locomotor activity of mice. Subchronic repeated pretreatment of animals with morphine showed an increase in locomotion induced by the opioid. Clozapine reduced locomotor activity induced by morphine in both the naïve and subchronic morphine- treated animals. Subchronic pretreatment of clozapine also caused an increase in the locomotion induced by morphine. Sulpiride also decreased locomotion induced by morphine and its subchronic administration of the drug caused an increase in morphine- or apomorphine-induced locomotion. Co-administration of clozapine with sulpiride did not elicit potentiation in inhibiting the morphine effect. The D2 receptor mRNA expression was also increased by repeated morphine administration. It may be concluded that morphine-induced sensitization may be due to increase in D2 receptor mRNA expression. Sulpiride and clozapine may induce sensitization and also inhibit morphine-induced locomotion through their dopamine receptor blocking properties.

Conditioned locomotion induced by unilateral intrastriatal administration of Apomorphine: D 2 receptor activation is critical but not the expression of the unconditioned response

The present study examined the role of D 1 and D 2 receptors in the conditioning of Apomorphine-induced locomotor behavior. A Pavlovian conditioning protocol was used in which rats received 5 daily intrastriatal Apomorphine treatments paired or unpaired to an open-field environment followed, 2 days later, by a saline test for conditioning. In the conditioning induction phase, the intrastriatal Apomorphine treatment increased locomotor activity expressed as an increased number of sectional crossings and rearings. In the conditioning test, the Apomorphine-paired group had significantly higher locomotor activity than the unpaired and vehicle groups, consistent with the development of a conditioned locomotor response. The concomitant blockade of D 1 and D 2 receptors with D 1 (SCH23390) and D 2 (Sulpiride) antagonists prevented the Apomorphine-induced behavioral response during the induction phase and in the conditioning test. Pretreatment with the D 1 antagonist SCH 23390 also blocked the Apomorphine-induced behavioral response during the induction phase but did not block the Apomorphine conditioned response. Pretreatment with the selective D 2 antagonist Sulpiride blocked the Apomorphine behavioral response during the induction phase and in the conditioning test. Altogether, these results indicate that antagonism of either the D 1 or D 2 receptors in the dorsal striatum can block Apomorphine-induced locomotor activation but that D 2 but not D 1 antagonism can prevent the development of the Apomorphine conditioned response. Altogether, these findings indicate a key role for the D 2 receptor site in the mediation of Apomorphine conditioned behavior; and, in addition, that Apomorphine conditioned locomotor response can develop without the expression of the locomotor stimulant response during the induction phase of conditioning.

Postnatal iron-induced motor behaviour alterations following chronic neuroleptic administration in mice

C57/BL6 mice were administered either 7.5 mg Fe(2+)/kg or vehicle (saline) postnatally on days 10-12 after birth. From 61 days of age onwards for 21 days, groups of mice were administered either clozapine (1 or 5 mg/kg, s.c.) or haloperidol (1 mg/kg, s.c.) or vehicle (Tween-80). Twenty-four hours after the final injection of either neuroleptic compound or vehicle, spontaneous motor activity was measured over a 60-min interval. Following this, each animal was removed, injected apomorphine (1 mg/kg, s.c.) and replaced in the same test chamber. It was found that postnatal administration of Fe(2+) at the 7.5 mg/kg dose level reduced activity during the initial 20-min periods (0-20 and 20-40 min) and then induced hyperactivity during the final 20-min period over all three parameters of activity. Subchronic treatment with the higher, 5 mg/kg, dose of clozapine abolished or attenuated the hypoactivity in by postnatal Fe(2+) during the 1(st) two 20-min periods over all three parameters of activity. Subchronic treatment with the higher, 5 mg/kg, dose of clozapine abolished or attenuated the hyperactivity in by postnatal Fe(2+) during the 3(rd) and final 20-min period. Subchronic administration of haloperidol, without postnatal iron, increased the level of both locomotion (1(st) 20 min) and rearing (2(nd) 20 min) activity. Postnatal administration of Fe(2+) at the 7.5 mg/kg dose increased the levels of both locomotion and rearing, but not total activity, following administration of apomorphine (1 mg/kg). Subchronic administration of clozapine, at both the 1 and 5 mg/kg doses, reduced the increased locomotor activity caused by postnatal Fe(2+), whereas clozapine, 5 mg/kg, elevated further the postnatal Fe(2+)-induced increased in rearing. Subchronic administration of clozapine, at both the 1 and 5 mg/kg doses, and haloperidol, 1 mg/kg, increased the level of locomotor following administration of apomorphine (1 mg/kg) in mice treated postnatally with vehicle, whereas only clozapine increased the level of rearing. Correlational analyses indicated that both apomorphine-induced locomotion and rearing were highly correlated with the total iron content in the basal ganglia, thereby offering direct evidence of the linear relationship between iron content in the basal ganglia and the behavioural expression of DA D(2)-receptor supersensitivity in mice.

Brain kinetics of methylphenidate (Ritalin) enantiomers after oral administration.

Methylphenidate (MP) (Ritalin) is widely used for the treatment of attention deficit hyperactivity disorder (ADHD). It is a chiral drug, marketed as the racemic mixture of d- and l-threo enantiomers. Our previous studies (PET and microdialysis) in humans, baboons, and rats confirm the notion that pharmacological specificity of MP resides predominantly in the d-isomer. A recent report that intraperitoneally (i.p.) administered l-threo-MP displayed potent, dose-dependent inhibition of cocaine- or apomorphine-induced locomotion in rats, raises the question of whether l-threo-MP has a similar effect when given orally. It has been speculated that l-threo-MP is poorly absorbed in humans when it is given orally because of rapid presystemic metabolism. To investigate whether l-threo-MP or its metabolites can be delivered to the brain when it is given orally, and whether l-threo-MP is pharmacologically active. PET and MicroPET studies were carried out in baboons and rats using orally delivered C-11-labeled d- and l-threo-MP ([methyl-(11)C]d-threo-MP and [methyl-(11)C]l-threo-MP). In addition, we assessed the effects of i.p. l-threo-MP on spontaneous and cocaine-stimulated locomotor activity in mice. There was a higher global uptake of carbon-11 in both baboon and rat brain for oral [(11)C]l-threo-MP than for oral [(11)C]d-threo-MP. Analysis of the chemical form of radioactivity in rat brain after [(11)C]d-threo-MP indicated mainly unchanged tracer, whereas with [(11)C]l-threo-MP, it was mainly a labeled metabolite. The possibility that this labeled metabolite might be [(11)C]methanol or [(11)C]CO(2), derived from demethylation, was excluded by ex vivo studies in rats. When l-threo-MP was given i.p. to mice at a dose of 3 mg/kg, it neither stimulated locomotor activity nor inhibited the increased locomotor activity due to cocaine administration. These results suggest that, in animal models, l-threo-MP or its metabolite(s) is (are) absorbed from the gastrointestinal tract and enters the brain after oral administration, but that l-threo-MP may not be pharmacologically active. These results are pertinent to the question of whether l-threo-MP contributes to the behavioral and side effect profile of MP during treatment of ADHD.

Increased dopamine D2 receptor binding and enhanced apomorphine-induced locomotor activity in μ-opioid receptor knockout mice

Previous studies from our laboratory have indicated possible interactions between opioidergic and dopaminergic neurons in the central nervous system. In this study, apomorphine-induced locomotor activity and the D1 and D2 subtype dopamine receptor binding were examined in mice lacking the μ-opioid receptor genes. The ambulatory time, vertical time and total motor distance of locomotor activity were measured after administration of apomorphine (2 mg/kg, i.p.) for a period of 90 min. The autoradiographic studies of D1 and D2 dopamine receptors were conducted using [ 3 H ] SCH23390 and [ 3 H ] raclopride as ligand, respectively. In wild type mice that received apomorphine, 2 mg/kg, i.p., the locomotor activity such as ambulatory time, vertical time and total motor distance were not significantly altered as compared with that of the saline control group. However, the locomotor activity measured was significantly increased in the same dose of apomorphine treated μ-opioid receptor knockout mice between 5 and 40 min after administration. The results obtained also show that the binding of D2 dopamine receptor in μ-opioid receptor knockout mice was significantly higher than that of the wild type in the caudate putamen. However, the binding of the D1 dopamine receptor in μ-opioid receptor knockout mice was not significantly different from that of the wild type. It appears that the apomorphine treated μ-opioid receptor knockout mice showed enhancement in locomotor activity. The enhanced locomotor activity may be related to the compensatory up-regulation of D2 dopamine receptors in mice lacking μ-opioid receptor genes.

Test Conditions Influence the Response to a Drug Challenge in Rodents

These studies were conducted to examine the differential response to a drug challenge under varied experimental test conditions routinely employed to study drug-induced behavioral and neurophysiological responses in rodents. Apomorphine, a nonselective dopamine agonist, was selected due to its biphasic behavioral effects, its ability to induce hypothermia, and to produce distinct changes to dopamine turnover in the rodent brain. From such experiments there is evidence that characterization and detection of apomorphine-induced activity in rodents critically depends upon the test conditions employed. In rats, detection of apomorphine-induced hyperactivity was facilitated by a period of acclimatization to the test conditions. Moreover, test conditions can impact upon other physiological responses to apomorphine such as drug-induced hypothermia. In mice, apomorphine produced qualitatively different responses under novel conditions when compared to those behaviors elicited in the home test cage. Drug-induced gross activity counts were increased in the novel exploratory box only, while measures of stereotypic behavior were similar in both. By contrast, apomorphine-induced locomotion was more prominent in the novel exploratory box. Dopamine turnover ratios (DOPAC:DA and HVA:DA) were found to be lower in those animals exposed to the exploratory box when compared to their home cage counterparts. However, apomorphine-induced reductions in striatal dopamine turnover were detected in both novel and home cage environments. The implications of these findings are discussed with particular emphasis upon conducting psychopharmacological challenge tests in rodents.

Progressive degeneration of dopamine system functions after transient cerebral oligemia in rats

A reduction in cerebral blood flow to oligemic levels was achieved in pentobarbital-anesthetized adult rats by clamping both carotid arteries (BCCA) for 60 min. To assess the extent to which the animals' dopaminergic system was affected over an increasing time span, their spontaneous locomotor activity in an unfamiliar environment and in response to the subcutaneous administration of apomorphine was tested at various times after either BCCA or sham operation. Eight to 14 days after the operation, it was possible to observe a diminished locomotor activity in response to apomorphine injection in BCCA as compared with sham-operated animals, while oral stereotypical behavior such as licking was increased. At 3 months, there was only a subtle decrease in apomorphine-induced locomotor activity, and stereotypical behavior was similar in both groups. At 7 months, the BCCA rats covered shorter distances than sham-operated controls during the habituation phase; after apomorphine injection, more stereotypic movements, such as, e.g., sniffing, were observed, and less running. Twelve months after surgery, no further differences could be observed between the two groups during the habituation phase, but the injection of apomorphine led to increased stereotypic sniffing movements, rearing and locomotor activity in BCCA animals to a greater extent than in the controls. At 12 months, sensorimotor disturbances elicited by the rota rod test, which were only transiently observed at 11 weeks and 7 months, did not appear any different from the normal age-related motor decline of the sham-operated controls. The animals' motor co-ordination in the chimney test was not significantly disturbed during the time between 7 and 12 months after surgery. At 15 months, nocturnal locomotor activities in BCCA rats were significantly decreased. In situ hybridization (ISH) histochemistry revealed decreased D1 receptor mRNA (D1RmRNA) in striatal neurons 19 months after surgery, while D2 receptor mRNA (D2RmRNA) and the neuronal number remained the same. The present results show that just as is already known for the immature rat brain, the adult rat brain, too, reacts to a transient decrease in its blood supply by appearance of long-lasting alterations in function, and that even a single oligemic episode is capable of inducing progressive dopaminergic dysfunctions and ultimately the partial loss of striatal D1RmRNA.

Changes in adult brain and behavior caused by neonatal limbic damage: implications for the etiology of schizophrenia

We tested the hypothesis that limbic damage in early development can cause aberrant maturation of brain structures known to be abnormal in adult schizophrenics: the hippocampus, prefrontal cortex, ventricles, and forebrain dopamine systems. We measured brain morphology, locomotor response to apomorphine, and cognitive processes in adult rats which received electrolytic damage to amygdala or hippocampus 48 h after birth. The behavioral measurements involved tasks which depend upon the integrity of the hippocampus or prefrontal cortex, and a task sensitive to forebrain dopamine system activation. The tasks included place navigation, egocentric spatial ability, and apomorphine-induced locomotion. The rats with lesions showed poor performance on the place navigation and egocentric spatial tasks and more apomorphine-induced locomotion after puberty than the sham lesion group. Regardless of lesion location, the adult rats showed smaller amygdalae and hippocampi, and larger lateral ventricles. Analyzing the lesion and sham rats together, adult amygdala volume was found to be positively correlated with cerebral cortex, prefrontal cortex, and hippocampal volumes and place navigation performance, and was negatively correlated with lateral ventricle volume. This study contributes to our understanding of the pathogenesis of schizophrenia by showing that early damage to limbic structures produced behavioral, morphological, and neuropharmacological abnormalities related to pathology in adult schizophrenics.

Behavioral interactions produced by co-administration of 7-OH-DPAT with cocaine or apomorphine in the rat.

Previous research from our laboratory suggests that low doses (<0.1 mg/kg) of the dopamine (DA) D3-preferring agonist 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT) attenuate conditioned place preference (CPP) produced by the indirect DA agonist d-amphetamine, but enhance d-amphetamine-induced stereotypic behaviors. This study further examined the effects of 7-OH-DPAT on behaviors produced by the indirect DA agonist, cocaine, and the non-selective direct DA agonist, apomorphine. To examine whether 7-OH-DPAT would alter cocaine and apomorphine dose-response curves for motor behaviors and CPP, 0.1 mg/kg 7-OH-DPAT was co-administered with 0-30 mg/kg cocaine and 0-3 mg/kg apomorphine. To establish place conditioning, drug injections were paired with one of two distinctly different compartments, whereas saline injections were paired with the other compartment. Locomotion, sniffing, oral stereotypy, and headbobbing were measured following acute and repeated drug administration during conditioning, and place conditioning was assessed 24 h following the last conditioning day. 7-OH-DPAT enhanced cocaine- and apomorphine-induced stereotypies following repeated administration. 7-OH-DPAT also attenuated cocaine-CPP, but potentiated apomorphine-CPP. Furthermore, 7-OH-DPAT attenuated locomotion produced by high doses of apomorphine. The attenuation of cocaine-CPP by 7-OH-DPAT likely involves stimulation of D2/D3 autoreceptors in the mesolimbic pathway, whereas the potentiation of apomorphine-CPP likely involves stimulation of D2/D3 postsynaptic receptors. Furthermore, it is suggested that attenuation of apomorphine-induced locomotion by 7-OH-DPAT likely involves stimulation of postsynaptic D3 receptors in the mesolimbic pathway. Thus, if postsynaptic D3 receptors are involved in mediating CPP and locomotion, then stimulation of D3 receptors may facilitate CPP but inhibit locomotion.

New antipsychotic agents with serotonin and dopamine antagonist properties based on a pyrrolo[2,1-b][1,3]benzothiazepine structure.

The development of a synthetic approach to the novel pyrrolo[2, 1-b][1,3]benzothiazepine and its derivatives and their biological evaluation as potential antipsychotic drugs are described. In binding studies these compounds proved to be potent 5-HT2, D2, and D3 receptor ligands. The more potent benzothiazepine (+/-)-3b was resolved into its enantiomers by using HPLC techniques. In vitro testing confirmed that (-)-3b is a more potent D2 receptor ligand, maintaining high affinity for 5-HT2 receptors. In contrast, the (+)-3b enantiomer presents a 35 times higher affinity for 5-HT2 than for dopamine D2 receptors with a similar dopamine D1 receptor affinity to that of (-)-3b. Overall, (+)-3b shows an "atypical" neuroleptic binding profile, while (-)-3b has a more "classical" profile. Furthermore pharmacological and biochemical testing displayed that the novel benzothiazepine (+/-)-3b is able to increase the extracellular levels of dopamine in the rat striatum and causes a dose-related suppression of apomorphine-induced locomotor activity. At low doses (+/-)-3b does not induce catalepsy, showing atypical antipsychotic properties similar to those of olanzapine. These heterocyclic compouds represent new leads for the development of novel antipsychotic drugs with atypical properties.

Synthesis and pharmacological evaluation of triflate-substituted analogues of clozapine: identification of a novel atypical neuroleptic.

The trifluoromethanesulfonyloxy (TfO) analogues 3 and 4 of 8-chloro-11-(4-methyl-1-piperazinyl)-5H-dibenzo[b,e][1,4]diazepine (clozapine, 1) and its 2-chloro isomer (iso-clozapine, 2), respectively, were synthesized via their OMe and OH analogues with the conventional synthetic method of the tricyclic dibenzodiazepines and evaluated pharmacologically along with their parent drugs. The binding profile of the 2-OTf analogue (4) is comparable to the binding profile of 1, although the affinity for the dopamine (DA) D2 receptors is higher (IC50 values are 31 nM and 330 nM for compounds 4 and 1, respectively). Interestingly, no notable affinity for muscarinic receptors could be detected in compound 4. On the contrary, the 8-OTf analogue 3 only displayed affinity for muscarinic M1 receptors (IC50 value 35 nM) and no affinity (IC50 value > 500 nM) for the other receptors tested. The 10 mumol/kg sc dose, but not the 10 mumol/kg po dose, of compound 4 stimulated the output of DA. Increases of 80% and 35% in DOPAC output from the dorsal striatum were seen after sc and po administrations of 10 mumol/kg of compound 4, respectively. Doses up to 100 mumol/kg of compound 3 had no effect on either parameter. Doses up to 100 mumol/kg of compound 4 were not cataleptogenic, but significantly decreased apomorphine-induced locomotor activity. In conclusion, compound 4 (GMC1-169) is a new clozapine-like neuroleptic candidate, which is lacking anticholinergic properties and displays a higher potency, as compared to clozapine (1) itself.

Phospholipase A 2 and the hypofrontality hypothesis of schizophrenia

Phospholipase A 2 (PLA 2) catalyzes the hydrolysis of membrane phospholipids to release cytotoxic products such as lysophosphatidylcholine. In schizophrenia increased PLA 2 activity and an accelerated breakdown of membrane phospholipids have been reported. In neuronal do membranes PLA 2 modulates dopamine (DA) release and DA receptor sensitivity. Using two different animal models we show inhibition of dopaminergic activity in vivo by intracerebral PLA 2 application. (1) Unilateral stereotaxic injection of PLA 2 into the substantia nigra pars compacta causes an apomorphine-induced ipsilateral rotational asymmetry. (2) Intracerebroventricular PLA 2 application reduces apomorphine-induced locomotion. NMR spectroscopy studies show a disordered phospholipid metabolism in the prefrontal cortex (PFC) of schizophrenics. The hypofrontality hypothesis of schizophrenia postulates hypodopaminergic activity in the prefrontal cortex. We speculate that increased PLA 2 activity in the PFC of schizophrenics might be related to both abnormalities and could thus contribute to hypofrontality in schizophrenia.

Intracerebroventricular injection of phospholipase A 2 inhibits apomorphine-induced locomotion in rats

Phospholipase A 2 (PLA 2) is a key enzyme in phospholipid metabolism. In neurons, membrane-bound PLA 2 plays an essential role in signal transduction by affecting neurotransmitter release and receptor sensitivity. There are some reports of increased PLA 2 activity in schizophrenia. We investigated the effects of intracerebroventricular (i.c.v.) injections of PLA 2 on dopamine-mediated behavior in rats. Ten days after i.c.v. injection, PLA 2 significantly inhibited apomorphine-induced locomotion as compared with i.c.v. saline injections. The inhibition of apomorphine-induced locomotion by PLA 2 was reversible within 4 weeks after stereotaxic surgery. These findings suggest a functional inhibition of dopaminergic postsynaptic receptors by PLA 2. Accelerated phospholipid metabolism and reduced dopaminergic activity in the prefrontal cortex have been postulated to play a role in schizophrenia. Increased PLA 2 activity may be related to both abnormalities and could thus play a role in the pathophysiology of schizophrenia.

Transmitter release from transplants of fetal ventral mesencephalon or locus coeruleus in the rat frontal cortex and nucleus accumbens: effects of pharmacological and behaviorally activating stimuli

The present study was performed in order to establish whether dopamine (DA) release from behaviorally functional intracerebral DA transplants is dependent on changes in neuronal impulse flow, and is under control of the host brain. Rats were subjected to combined intraventricular and ventral tegmental injections of 6-hydroxydopamine (6-OHDA) in order to obtain a severe bilateral lesion of the ascending mesocorticolimbic DA projections. Cell suspension grafts of fetal ventral mesencephalic neurons were thereafter implanted into the medial frontal cortex (MFC) and the nucleus accumbens (NAc). Since the neurotoxin injections removed also the ascending noradrenergic systems, fetal locus coeruleus neurons were added to the graft suspension in one group of animals. Age-matched lesion-only and normal animals served as controls. The lesion-induced alterations in spontaneous, amphetamine- and apomorphine-induced locomotor activity and in a skilled paw reaching task were evaluated before transplantation, and at 3 and 6 months post-grafting. Microdialysis probes were finally implanted in the MFC and NAc in order to monitor extracellular DA and noradrenaline (NA) levels ( i) during administration of pharmacological agents which augment or depress catecholamine release in the intact brain; ( ii) during exposure of the rats to stressful manipulations ( handling and immobilization) or appetitive stimuli ( eating) known to enhance cortical and limbic DA or NA release in intact animals. The lesion-induced reduction in amphetamine-induced locomotor activity was reversed in all grafted animals, which also showed a higher than normal spontaneous overnight activity. Daytime spontaneous locomotor activity (which was reduced in the lesion-only rats) as well as apomorphine-induced hyperactivity was reversed by the grafts of DA neurons only. By contrast, the lesion-induced impairment in skilled forelimb use was not alleviated by the grafts. The grafted DA neurons restored normal steady-state DA overflow in the NAc, whereas they enhanced cortical DA overflow to significantly higher than normal levels. Restoration of both cortical and striatal NA overflow was observed in the group that received mixed DA and NA grafts, whereas animals that received DA grafts only did not differ from the lesioned controls. The changes in extracellular DA and NA levels measured in the grafted MFC and NAc under potassium depolarization (100 mM KCl), inhibition of terminal catecholamine reuptake (10 μM nomifensine), and sodium channel blockade (1 μM TTX) indicated that graft-derived DA or NA release had normal neuronal properties, and was dependent on an intact axonal impulse flow. The grafted DA and NA neurons lacked the ability to respond to immobilization stress with enhanced transmitter release. However, most of the animals that received DA grafts exhibited an enhanced DA overflow in the NAc following handling stimulation. When the animals were engaged in eating, a significant 20% increase in DA overflow was detected in the grafted cortices, and a marginally significant 58% increase in DA overflow was measured in the DA-grafted NAc. By contrast, neither cortical nor accumbens DA overflow was significantly affected during eating behavior in the lesion-only controls. NA overflow from the grafted locus coeruleus neurons was also unaffected by the appetitive stimulus. The present results indicate that DA or NA release from intracortically and intrastriatally grafted DA and NA neurons is dependent on changes in axonal impulse flow. The findings of enhanced DA release from intracortically grafted DA neurons during eating and from intrastriatally grafted DA neurons during rewarded behavior and handling stimulation, suggest that the transplanted DA neurons are reached by physiologically relevant inputs from the host brain. On the other hand, the lack of response to forced immobilization (i.e. a stimulus which is normally a powerful activator of cortical DA release), reveals that the functional integration of the grafted DA neurons with the host brain is incomplete.