Nonhuman primates: translational models for predicting antipsychotic-induced movement disorders.

Repeated haloperidol treatment administered to nonhuman primates (NHPs) over several months or even years leads to the gradual appearance of drug-induced dystonic reactions in the orofacial region (mouth opening, tongue protrusion or retraction, bar biting) and in the whole body (writhing of the limbs and trunk, bar grasping). The propensity of antipsychotics to induce dystonia in NHPs is not correlated with their propensity to induce catalepsy in rodents, suggesting that the two types of effects are dissociated and may represent distinct aspects of the extrapyramidal symptoms induced by antipsychotics. In view of the clear homology to clinically observed phenomena, antipsychotic-induced dystonias in antipsychotic-primed NHPs would appear to possess a high degree of translational validity. These NHP phenomena could therefore serve as a useful model for predicting the occurrence of similar abnormal movements with novel substances developed for the treatment of schizophrenia or other psychotic disorders. Moreover, the NHP dystonia model could possibly serve as a biomarker for substances that will eventually cause tardive dyskinesia in patients.

How good are current approaches to nonclinical evaluation of abuse and dependence?

Nonclinical assessment of drug abuse and dependence is the subject of several recent regulatory guidelines, which are generally aligned on the methods to be employed. The most direct approach to assessing reinforcing properties of a drug is the self-administration procedure whereby animals can initiate intravenous injections of the test substance, something they readily do with prototypic drugs of abuse. Complications arise because there is no standardized procedure for evaluating substances with differing potencies, reinforcement properties, or pharmacokinetics. Moreover, the choice of training substance, species, and procedural parameters can radically affect the outcome. Apart from the lower cost of rats, primates present several advantages for self-administration studies with similarity to human pharmacokinetics in particular. The most powerful method for assessing similarities between a test substance and a prototypic drug of abuse is the drug discrimination procedure. In contrast to self-administration, drug discrimination is pharmacologically very specific, often reflecting functional activity at receptor level. Dependence is assessed by the occurrence of withdrawal effects on drug discontinuation. Although conceptually simple, interpretation can be complicated by factors such as duration and frequency of administration and observations as well as the choice of end points. Telemetry allows continuous observation of multiple parameters during withdrawal, thereby increasing sensitivity. Presently available tools identify all substances known to cause abuse or dependence, with little risk of false-positives. It remains unclear, however, how predictive these models are with entirely novel substances. Nonetheless, drug abuse/dependence is an area of safety pharmacology where the predictive value of animal models is remarkably high.

Behavioral indices in antipsychotic drug discovery.

Schizophrenia is characterized by three major symptom classes: positive symptoms, negative symptoms, and cognitive deficits. Classical antipsychotics (phenothiazines, thioxanthenes, and butyrophenones) are effective against positive symptoms but induce major side effects, in particular, extrapyramidal symptoms (EPS). The discovery of clozapine, which does not induce EPS and is thought effective against all three classes of symptom, has driven research for novel antipsychotics with a wider activity spectrum and lower EPS liability. To increase predictiveness, current efforts aim to develop translational models where direct parallels can be drawn between the processes studied in animals and in humans. The present article reviews existing procedures in animals for their ability to predict compound efficacy and EPS liability in relation to their translational validity. Rodent models of positive symptoms include procedures related to dysfunction in central dopamine and glutamatergic (N-methyl-D-aspartate) and serotonin (5-hydroxytryptamine) neurotransmission. Procedures for evaluating negative symptoms include rodent models of anhedonia, affective flattening, and diminished social interaction. Cognitive deficits can be assessed in rodent models of attention (prepulse inhibition) and of learning/memory (object and social recognition, Morris water maze and operant-delayed alternation). The relevance of the conditioned avoidance response is also discussed. A final section reviews procedures for assessing EPS liability, in particular, parkinsonism (catalepsy in rodents), acute dystonia (purposeless chewing in rodents, dystonia in monkeys), akathisia (defecation in rodents), and tardive dyskinesia (long-term antipsychotic treatment in rodents and monkeys). It is concluded that, with notable exceptions (attention, learning/memory, EPS liability), current predictive models for antipsychotics fall short of clear translational validity.

Continuous stimulation of dopaminergic receptors by rotigotine does not interfere with the sleep-wake cycle in the rat.

Rotigotine is a new, non-ergoline dopamine receptor agonist developed for the treatment of Parkinson's disease in a transdermal formulation (Neupro ) to provide sustained drug delivery for 24 h with a once daily dosing. The aim of the present study was to determine whether or not continuous dopaminergic stimulation can interfere with the sleep-wake cycle. To achieve this, rotigotine was administered as a slow release formulation to provide stable plasma and brain levels over a period of 6 days and the sleep-wake cycle was evaluated in the freely-moving male rat using electroencephalagraphic recording. For comparison, the mixed dopamine/noradrenaline reuptake inhibitor nomifensine (16 mg/kg p.o.) was administered once daily for 6 days. In contrast to nomifensine, rotigotine (0.5 and 5 mg/kg s.c.) had no clear effects on the sleep-wake cycle. Nomifensine delayed the onset of rapid eye movement (REM) sleep and, to a lesser extent, also that of slow wave sleep (SWS). In addition, it increased the duration of waking time and decreased the duration of SWS and REM sleep. These effects were observed on all days and repeated administration lead neither to potentiation nor attenuation of the effects. It is concluded that a continuous dopaminergic stimulation of dopamine receptors by rotigotine may not only be beneficial for the treatment of the motor symptoms of Parkinson's disease but also have additional benefits by not compromising either sleep architecture or circadian rhythm.

The use of the dog electroencephalogram (EEG) in safety pharmacology to evaluate proconvulsant risk.

INTRODUCTION: The dog is frequently used for cardiovascular safety pharmacology and for toxicology studies, but is not often used for central nervous system (CNS) safety pharmacology purposes. We have therefore examined the electroencephalogram (EEG) in conscious dogs by means of radio-telemetry methods using the proconvulsant agent pentylenetetrazole (PTZ) as reference substance. Assessment of proconvulsant risk is an important aspect of CNS safety evaluation and the EEG is a sensitive technique for identifying pathologic brain activity, most importantly paroxysmal activity. METHODS: Dogs were implanted with epidural electrodes wired to subcutaneously placed radiotransmitters. Following baseline recording, the test substance was administered and the EEG and electromyogram (EMG) activities were recorded from dogs placed in slings. The EEG was assessed visually for abnormal activity and dogs were also continuously observed for the appearance of overt convulsive activity. The PTZ infusion was stopped and diazepam was administered as soon as clear and sustained EEG effects and/or behavioural symptoms occurred. RESULTS: Slow i.v. infusion of PTZ (1.5 mg/kg/min) induced clear paroxysmal effects on the EEG trace in the form of spike and wave trains of 4-5 Hz. Paroxysmal activity associated with clonic convulsions occurred between 17 and 36 min after the start of infusion (a mean of 24 min) but in most cases paroxysmal activity was observed approximately 60 s prior to any overt convulsive activity. DISCUSSION: These data show the usefulness of the telemetered dog EEG in safety pharmacology. The dog EEG is appropriate in situations where results from cardiovascular and CNS safety tests in the same species are required, or where the use of other species is contraindicated because of metabolic or pharmacokinetic particularities.

New perspectives in CNS safety pharmacology.

International requirements for central nervous system (CNS) safety pharmacology are reviewed. Procedures for initial CNS safety screening (core battery studies) can be conducted from the beginning of the drug discovery process, but at latest before first studies in man. They should include assessment of general behaviour, locomotor activity and motor coordination, but can also include studies of pain sensitivity, convulsive threshold and interaction with hypnotics. Follow-up studies, to be conducted later in the drug development process but before product approval, cover assessment of higher cognitive function, electroencephalogram (EEG) and drug dependence/abuse liability. Procedures for assessing cognitive function can include, in order of complexity, passive avoidance, Morris and radial mazes and operant behaviour tasks (delayed alternation, repeated acquisition). EEG can include the quantified EEG (QEEG) and studies of the sleep/wakefulness cycle. Drug dependence/abuse procedures can include precipitated and nonprecipitated withdrawal (drug dependence), and place preference, drug discrimination and self-administration (drug abuse). In contrast to core battery CNS procedures, conducted exclusively in rodents, follow-up studies can include higher species, in particular primates.

The usefulness of non-human primates in central nervous system safety pharmacology.

INTRODUCTION: The present paper will suggest, on the basis of experimental evidence, that several non-human primate (NHP) procedures can be uniquely useful and relevant for central nervous system (CNS) safety pharmacology purposes. METHODS AND RESULTS: Classical antipsychotics (e.g. haloperidol) but not atypical antipsychotics (e.g. clozapine), in contrast to rodents, induce behavioral signs in NHP which are clearly homologous to those observed in humans and thus have high translational value. Operant techniques (delayed matching/non-matching-to-sample) and non-operant techniques (object retrieval) can be used in NHP to assess the facilitating and impairing effects of drugs on cognition. Brain structures sub-serving these functions are closer to humans in NHP than in rodents suggesting that drug data from NHP translate better to humans. Biting into a rubber tube can be induced in squirrel monkeys by exposure to non-reinforcement (frustration). This model is close to human notions of frustration/aggression, and is ethically more acceptable than methods using shock or animal fighting. It could therefore serve as a model of drug-induced irritability with potentially high translational value. CONCLUSION: There are cogent scientific reasons for selecting NHP in CNS and other safety pharmacology areas.

Rodent models of depression: forced swim and tail suspension behavioral despair tests in rats and mice.

The development of antidepressants requires simple rodent behavioral tests for initial screening before undertaking more complex preclinical tests and clinical evaluation. Presented in the unit are two widely used screening tests used for antidepressants, the forced swim (also termed behavioral despair) test in the rat and mouse, and the tail suspension test in the mouse. These tests have good predictive validity and allow rapid and economical detection of substances with potential antidepressant-like activity. The behavioral despair and the tail suspension tests are based on the same principle: measurement of the duration of immobility when rodents are exposed to an inescapable situation. The majority of clinically used antidepressants decrease the duration of immobility. Antidepressants also increase the latency to immobility, and this additional measure can increase the sensitivity of the behavioral despair test in the mouse for certain classes of antidepressant. Testing of new substances in the behavioral despair and tail suspension tests allows a simple assessment of their potential antidepressant activity by the measurement of their effect on immobility.

Continuous evaluation of drug withdrawal in the rat using telemetry: effects of morphine and chlordiazepoxide.

INTRODUCTION: The procedures used to assess withdrawal must be sensitive and widely applicable, i.e. not specific to any particular drug class. Furthermore, the measurements should not be affected by repeat testing. METHODS: We have used implanted telemetry devices to continuously follow body temperature, locomotor activity (LMA), heart rate (HR) and mean arterial blood pressure (mean ABP) in addition to food intake and body weight gain over 20days of treatment and 8days of withdrawal. The effects of morphine (32 and 64mg/kg p.o., b.i.d.) and chlordiazepoxide (16, 32 and 64mg/kg p.o., b.i.d.) were studied in rats. RESULTS: The results show that during the treatment phase chronic morphine reduced food intake and body weight gain, increased body temperature, HR, mean ABP and LMA. These effects continued over the 20days of treatment. In contrast, chlordiazepoxide slightly increased food intake and body weight gain throughout the treatment period. It also decreased body temperature and LMA but increased HR and mean ABP after the first few administrations but these effects disappeared over the 20days of treatment. Following discontinuation, both morphine- and chlordiazepoxide-treated rats showed a dose-related decrease in food intake and loss of weight on days 2 and 3 of discontinuation. Morphine discontinuation also induced a nocturnal hypothermia and a diurnal hypertension (i.e. during the light phase) which lasted for 4-5days and also moderate diurnal increases in locomotor activity and heart rate over the first 3days of discontinuation. Chlordiazepoxide discontinuation induced small increases in telemetry parameters some of which, such as the effect on locomotor activity, lasted for more than 5days. The intensity and duration of effects for both substances were broadly dose-related. DISCUSSION: These data show that telemetry can increase the sensitivity of withdrawal experiments to changes that might otherwise be missed and allows a better definition of the time-course of withdrawal effects. This technique is therefore useful as part of safety pharmacology abuse liability evaluation of novel test substances across a broad range of pharmacological and therapeutic classes.

Rodent models of depression: forced swim and tail suspension behavioral despair tests in rats and mice.

The development of antidepressants requires simple rodent behavioral tests for initial screening before undertaking more complex preclinical tests and clinical evaluation. Presented in the unit are two widely used screening tests used for antidepressants, the forced swim (also termed behavioral despair) test in the rat and mouse, and the tail suspension test in the mouse. These tests have good predictive validity and allow rapid and economical detection of substances with potential antidepressant-like activity. The behavioral despair and the tail suspension tests are based on the same principle: measurement of the duration of immobility when rodents are exposed to an inescapable situation. The majority of clinically used antidepressants decrease the duration of immobility. Antidepressants also increase the latency to immobility, and this additional measure can increase the sensitivity of the behavioral despair test in the mouse for certain classes of antidepressant. Testing of new substances in the behavioral despair and tail suspension tests allows a simple assessment of their potential antidepressant activity by the measurement of their effect on immobility.

Use of latency to immobility improves detection of antidepressant-like activity in the behavioral despair test in the mouse.

The behavioral despair test (BDT), also called the forced swim test, is an economic, reliable and sensitive test for the detection of potential antidepressant-like activity of new test substances. The vast majority of clinically active antidepressants are active in the BDT, although substances specifically acting on serotonin transmission are generally reported to be less easily detected. Substances active in the BDT decrease the duration of immobility at doses considered as relatively high. In contrast, some psychostimulants are considered as potential false positives since they are also active in the BDT although they are not recognized as clinically active antidepressants. In the present study we have evaluated the usefulness of latency to the first immobility period as an additional parameter in the BDT to further evaluate the effects of antidepressants and psychostimulants administered intraperitoneally in the mouse. The results show that this measure increases the sensitivity of the test for detecting the effects of tricyclic antidepressants (imipramine, desipramine) and selective serotonin/norepinephrine reuptake inhibitors (duloxetine and venlafaxine) but not of serotonin reuptake inhibitors (fluoxetine and escitalopram). In contrast with previous reports, psychostimulants (amphetamine and modafinil) did not affect the duration or the latency to immobility in the BDT. The mouse strain used in the BDT seems to be an important parameter to discriminate between antidepressants and psychostimulants. These results suggest that the measure of the latency to the first immobility improves the predictive validity of the BDT.

Preclinical behavioral models for predicting antipsychotic activity.

Schizophrenia is a major psychiatric disease that is characterized by three distinct symptom domains: positive symptoms, negative symptoms, and cognitive impairment. Additionally, treatment with classical antipsychotic medication can be accompanied by important side effects that involve extrapyramidal symptoms (EPS). The discovery of clozapine in the 1970s, which is efficacious in all three symptom domains and has a reduced propensity to induce EPS, has driven research for new antipsychotic agents with a wider spectrum of activity and a lower propensity to induce EPS. The following chapter reviews existing behavioral procedures in animals for their ability to predict compound efficacy against schizophrenia symptoms and liability to induce EPS. Rodent models of positive symptoms include procedures related to hyperfunction in central dopamine and serotonin (5-hydroxytryptamine) systems and hypofunction of central glutamatergic (N-methyl-d-aspartate) neurotransmission. Procedures for evaluating negative symptoms include rodent models of anhedonia, affective flattening, and diminished social interaction. Cognitive deficits can be assessed in rodent models of attention (prepulse inhibition (PPI), latent inhibition) and of learning and memory (passive avoidance, object and social recognition, Morris water maze, and operant-delayed alternation). The relevance of the conditioned avoidance response (CAR) is also discussed. A final section reviews animal procedures for assessing EPS liability, in particular parkinsonism (catalepsy), acute dystonia (purposeless chewing in rodents, dystonia in monkeys), akathisia (defecation in rodents), and tardive dyskinesia (long-term antipsychotic treatment in rodents and monkeys).

Overview of safety pharmacology.

Safety pharmacology assesses the potential risks of novel pharmaceuticals for human use. ICH S7A guidelines recommend a core battery of studies on three vital systems: central nervous system (CNS), cardiovascular (CV) and respiratory. Primary CNS studies look for effects on general behavior, locomotion, neuromuscular coordination, the convulsive threshold, and vigilance. The primary CV test battery evaluates proarrhythmic risk using in vitro approaches (hERG channel and Purkinje fiber assays) and in vivo measurements in conscious animals via telemetry. Comprehensive cardiac risk assessment also includes full hemodynamic evaluation, most suitably conducted in the anesthetized large animal. Basic respiratory function can be approached via whole-body plethysmography in conscious animals where sensitivity to respiratory-depressant effects can be enhanced by exposure to increased CO(2). Other aspects of safety pharmacology studies discussed are the timing of studies, ethical and animal welfare issues, and statistical evaluation.

Primary observation (Irwin) test in rodents for assessing acute toxicity of a test agent and its effects on behavior and physiological function.

The Irwin observation test is commonly used to evaluate the effects of a new substance on behavior and physiological function. The results of the Irwin test are used to determine potential toxicity and to select doses for specific therapeutic activity. The Irwin test can also be used in a safety approach for detecting untoward effects of a new compound on general behavior and for evaluating its acute neurotoxicity. In particular, data obtained in the Irwin Test can help to determine the dose range to be tested in other safety tests. Furthermore, the Irwin Test can furnish a first but pertinent orientation towards a specific therapeutic indication, a specific mechanism of action or a specific physiological function.

Pharmacological properties of ABT-239 [4-(2-{2-[(2R)-2-Methylpyrrolidinyl]ethyl}-benzofuran-5-yl)benzonitrile]: II. Neurophysiological characterization and broad preclinical efficacy in cognition and schizophrenia of a potent and selective histamine H3 receptor antagonist.

Acute pharmacological blockade of central histamine H3 receptors (H3Rs) enhances arousal/attention in rodents. However, there is little information available for other behavioral domains or for repeated administration using selective compounds. ABT-239 [4-(2-{2-[(2R)-2-methylpyrrolidinyl]ethyl}-benzofuran-5-yl)benzonitrile] exemplifies such a selective, nonimidazole H3R antagonist with high affinity for rat (pK(i) = 8.9) and human (pK(i) = 9.5) H3Rs. Acute functional blockade of central H3Rs was demonstrated by blocking the dipsogenia response to the selective H3R agonist (R)-alpha-methylhistamine in mice. In cognition studies, acquisition of a five-trial, inhibitory avoidance test in rat pups was improved with ABT-239 (0.1-1.0 mg/kg), a 10- to 150-fold gain in potency, with similar efficacy, over previous antagonists such as thioperamide, ciproxifan, A-304121 [(4-(3-(4-((2R)-2-aminopropanoyl)-1-piperazinyl)propoxy)phenyl)(cyclopropyl) methanone], A-317920 [N-((1R)-2-(4-(3-(4-(cyclopropylcarbonyl) phenoxy)propyl)-1-piperazinyl)-1-methyl-2-oxoethyl)-2-furamide], and A-349821 [(4'-(3-((R,R)2,5-dimethyl-pyrrolidin-1-yl)-propoxy)-biphenyl-4-yl)-morpholin-4-yl-methanone]. Efficacy in this model was maintained for 3 to 6 h and following repeated dosing with ABT-239. Social memory was also improved in adult (0.01-0.3 mg/kg) and aged (0.3-1.0 mg/kg) rats. In schizophrenia models, ABT-239 improved gating deficits in DBA/2 mice using prepulse inhibition of startle (1.0-3.0 mg/kg) and N40 (1.0-10.0 mg/kg). Furthermore, ABT-239 (1.0 mg/kg) attenuated methamphetamine-induced hyperactivity in mice. In freely moving rat microdialysis studies, ABT-239 enhanced acetylcholine release (0.1-3.0 mg/kg) in adult rat frontal cortex and hippocampus and enhanced dopamine release in frontal cortex (3.0 mg/kg), but not striatum. In summary, broad efficacy was observed with ABT-239 across animal models such that potential clinical efficacy may extend beyond disorders such as ADHD to include Alzheimer's disease and schizophrenia.

Place preference test in rodents.

Conditioned place preference is a behavioral model currently used to measure the rewarding properties induced by the administration of a drug. In this paradigm, the rewarding properties of a compound are associated with the particular characteristics of a given environment. Due to the recent availability of multiple lines of genetically modified mice, this unit includes procedures for testing place preference in both rats and mice.

Place preference test in rodents.

Conditioned place preference is a behavioral model currently used to measure the rewarding properties induced by the administration of a drug. In this paradigm, the rewarding properties of a compound are associated with the particular characteristics of a given environment. Advantages of this test are the absence of any instrumental learning and the possibility of evaluating reinforcing effects in the absence of the test substance. Due to the recent availability of multiple lines of genetically modified mice, this unit has been updated to include procedures for testing place preference in both rats and mice. Details are also provided for evaluating drug effects using this procedure.