The Discriminative Stimulus Properties of the Atypical Antipsychotic Clozapine in C57BL/6 Mice

Serotonin and α1 adrenergic receptor antagonism may contribute to atypical antipsychotic drug effects. Clozapine (2.5 mg/kg) drug discrimination in C57BL/6 mice may selectively screen atypical antipsychotic drugs. Previous data show that the atypical antipsychotics olanzapine, risperidone, ziprasidone but not the typical antipsychotic haloperidol fully substitutes for clozapine. The present study demonstrated that the atypical antipsychotics quetiapine, sertindole, zotepine, iloperidone, melperone fully substituted for clozapine but aripiprazole did not. The typical antipsychotics fluphenazine and perphenazine failed to fully substitute for clozapine but chlorpromazine and thioridazine fully substituted for clozapine. This model does not differentiate between atypical and typical antipsychotic drugs but it may be useful in the detection of antipsychotics with potent serotonin and α1 adrenergic receptor antagonist actions

Discriminative stimulus properties of 1.25 mg/kg clozapine in rats: Mediation by serotonin 5-HT2 and dopamine D4 receptors

The atypical antipsychotic drug clozapine remains one of most effective treatments for schizophrenia, given a lack of extrapyramidal side effects, improvements in negative symptoms, cognitive impairment, and in symptoms in treatment-resistant schizophrenia. The adverse effects of clozapine, including agranulocytosis, make finding a safe clozapine-like a drug a goal for drug developers. The drug dis- crimination paradigm is a model of interoceptive stimulus that has been used in an effort to screen experimental drugs for clozapine-like atypical antipsychotic effects. The present study was conducted to elucidate the receptor-mediated stimulus properties that form this clozapine discriminative cue by testing selective receptor ligands in rats trained to discriminate a 1.25 mg/kg dose of clozapine from vehicle in a two choice drug discrimination task. Full substitution occurred with the 5-HT2A inverse agonist M100907 and the two preferential D4/5-HT2/α1 receptor antagonists Lu 37-114 ((S)-1-(3-(2-(4- (1H-indol-5-yl)piperazin-1-yl)ethyl)indolin-1-yl)ethan-1-one) and Lu 37-254 (1-(3-(4-(1H-indol-5-yl) piperazin-1-yl)propyl)-3,4-dihydroquinolin-2(1H)-one). Partial substitution occurred with the D4 re- ceptor antagonist Lu 38-012 and the α1 adrenoceptor antagonist prazosin. Drugs selective for 5-HT2C, 5-HT6 muscarinic, histamine H1, and benzodiazepine receptors did not substitute for clozapine. The present findings suggest that 5-HT2A inverse agonism and D4 receptor antagonism mediate the dis- criminative stimulus properties of 1.25 mg/kg clozapine in rats, and further confirm that clozapine produces a complex compound discriminative stimulus

Generalization to Atypical Antipsychotic Drugs Depends On Training Dose in Rats Trained to Discriminate 1.25 Mg/Kg Clozapine Versus 5.0 Mg/Kg Clozapine Versus Vehicle in a Three-Choice Drug Discrimination Task

The prototypical atypical antipsychotic drug (APD) clozapine (CLZ) elicits a discriminative cue that appears to be similar to the stimulus properties elicited by atypical, but not typical, antipsychotic drugs in two-choice drug discrimination procedures. However, the ability of CLZ to generalize to atypical APDs depends on the training dose, since several atypical APDs (e.g. sertindole, risperidone) do not substitute for a 5.0 mg/kg CLZ training dose in rats, but do so for a 1.25 mg/kg CLZ training dose. Yet, a 1.25 mg/kg CLZ discriminative stimulus has not generalized to all atypical APDs either (e.g. quetiapine); thus, both 1.25 mg/kg and 5.0 mg/kg CLZ discriminative stimuli may be necessary to provide a better screen for atypical APDs. The present study sought to determine whether a three-choice 1.25 mg/kg CLZ versus 5.0 mg/kg CLZ versus vehicle drug discrimination task in rats might better distinguish atypical from typical APDs. Adult male Sprague-Dawley rats were trained in this three-choice drug discrimination task with a fixed ratio 30 reinforcement schedule for food. Clozapine produced full substitution (≥ 80% condition-appropriate responding) for both the 1.25 mg/kg CLZ dose (ED 50 = 0.09 mg/kg) and the 5.0 mg/kg CLZ dose (ED50 = 2.71 mg/kg). The atypical APD olanzapine produced full substitution for the 5.0 mg/kg CLZ dose, but not for the 1.25 mg/kg CLZ dose (ED50 = 1.55 mg/kg). In contrast, the atypical APD quetiapine produced full substitution for the 1.25 mg/kg CLZ dose (ED50 = 0.13 mg/kg), but not for the 5.0 mg/kg CLZ dose. Similarly, the atypical APD sertindole produced full substitution for only the 1.25 mg/kg CLZ dose (ED50 = 0.94 mg/kg). Risperidone, another atypical APD, produced partial substitution (≥ 60% and ≤, 80% condition-appropriate responding) for the 1.25 mg/kg CLZ dose, and failed to substitute for the 5.0 mg/kg CLZ dose. The atypical APD ziprasidone and the typical APDs haloperidol and chlorpromazine failed to substitute for either CLZ training dose. These results demonstrated that the 1.25 mg/kg CLZ training dose provides partial or full stimulus generalization to more atypical APDs than does the 5.0 mg/kg CLZ training dose. Full substitution by olanzapine for only the 5.0 mg/kg CLZ dose suggests that this higher training dose is also important for screening atypical APDs

Discriminative Stimulus Properties of the Atypical Antipsychotic Drug Clozapine in Rats Trained to Discriminate 1.25 Mg/Kg Clozapine Vs. 5.0 Mg/Kg Clozapine Vs. Vehicle

Clozapine, the prototype for atypical antipsychotic drugs, is used in the drug discrimination paradigm as a model for screening atypical from typical antipsychotic drugs. Previous drug discrimination studies in rats have shown that a 1.25 mg/kg clozapine training dose provides full stimulus generalization (i.e.) ≥ 80% condition-appropriate responding) to most atypical antipsychotic drugs, although a 5.0 mg/kg clozapine training dose appears necessary to provide stimulus generalization to other atypical antipsychotic drugs. The present study sought to characterize the pharmacological mechanisms that mediate these clozapine training doses. In rats trained to discriminate 1.25 vs. 5.0 mg/kg clozapine vs. vehicle in a three-choice drug discrimination task, various receptor-selective compounds were tested for stimulus generalization. The antidepressant mianserin was also tested. Full stimulus generalization from the 1.25 mg/kg clozapine training dose occurred only to mianserin (98.8%). Partial substitution (i.e. ≥ 60% and \u3c80% condition-appropriate responding) to the 5.0 mg/kg clozapine training dose occurred for the muscarinic receptor antagonist scopolamine. The combined total percentage of responding on the 1.25 and 5.0 mg/kg clozapine levers, however, was well above the full substitution criteria at the 0.25, 0.5, and 1.0 mg/kg scopolamine doses. The M1 agonist N-desmethylclozapine, the nicotinic antagonist mecamylamine, the D 1 antagonist SCH 23390, the D4 antagonist LU 38-012, the 5-HT1A agonist (+)-8-OH-DPAT, the 5-HT1A antagonist WAY 1 00 635, the 5-HT2A/2B/2C antagonist ritanserin, the 5-HT6 antagonist RO4368554, the α1 antagonist prazosin, the α2 antagonist yohimbine, and the histamine H1 antagonist pyrilamine all failed to substitute for either the 1.25 or the 5.0 mg/kg clozapine training doses. These results are consistent with previous evidence that antidepressant drugs have a tendency to substitute for clozapine and that muscarinic receptor antagonism may mediate the discriminative stimulus properties of 5.0 mg/kg clozapine. The lack of stimulus generalization from either clozapine training dose to other receptor-selective compounds, however, fails to explain how this model screens atypical from typical antipsychotic drugs and suggests that the discriminative stimulus properties of clozapine consist of a compound cue

Serotonin Receptor Mechanisms Mediate the Discriminative Stimulus Properties of the Atypical Antipsychotic Clozapine in C57BL/6 Mice

Rationale: The atypical antipsychotic drug (APD) clozapine (CLZ) has been shown to have a robust discriminative cue in rats, pigeons, and monkeys in two-choice drug discrimination procedures. Objectives: The present study determined whether a two-choice drug discrimination procedure with CLZ could be established in C57BL/6 mice and whether this procedure could distinguish between atypical and typical APDs. Methods: C57BL/6 male mice were trained to discriminate 2.5 mg/kg CLZ from vehicle in a two-lever drug discrimination procedure. Results: Generalization testing with CLZ produced full substitution at the 2.5- and 5.0-mg/kg doses with an ED50 of 1.14 mg/kg. The atypical APDs olanzapine (ED50=0.24 mg/kg), risperidone (ED 50=0.072 mg/kg), and ziprasidone (ED50=0.33 mg/kg) fully substituted for CLZ\u27s discriminative cue, while the typical APD haloperidol failed to substitute for CLZ. Generalization testing with selective ligands showed that the serotonin (5-HT)2A/2B/2C antagonist ritanserin fully substituted for CLZ (ED50=2.08 mg/kg) and that the 5-HT receptor agonist quipazine significantly attenuated CLZ\u27s discriminative cue without disrupting response rates. The muscarinic receptor antagonist scopolamine, the dopamine agonist amphetamine, and the 5-HT agonist quipazine failed to substitute for CLZ. Conclusions: These results demonstrated that antagonism of 5-HT receptors plays an important role in mediating the discriminative stimulus properties of the atypical APD CLZ in C57BL/6 mice. The atypical APDs olanzapine, risperidone, and ziprasidone fully substituted for CLZ, while the typical APD haloperidol did not. These results suggest that CLZ drug discrimination in C57BL/6 mice may be an effective preclinical behavioral assay for screening atypical from typical antipsychotic drugs

Striatal signal transduction and drug addiction

Drug addiction is a severe neuropsychiatric disorder characterized by loss of control over motivated behavior. The need for effective treatments mandates a greater understanding of the causes and identification of new therapeutic targets for drug development. Drugs of abuse subjugate normal reward-related behavior to uncontrollable drug-seeking and -taking. Contributions of brain reward circuitry are being mapped with increasing precision. The role of synaptic plasticity in addiction and underlying molecular mechanisms contributing to the formation of the addicted state are being delineated. Thus we may now consider the role of striatal signal transduction in addiction from a more integrative neurobiological perspective. Drugs of abuse alter dopaminergic and glutamatergic neurotransmission in medium spiny neurons of the striatum. Dopamine receptors important for reward serve as principle targets of drugs abuse, which interact with glutamate receptor signaling critical for reward learning. Complex networks of intracellular signal transduction mechanisms underlying these receptors are strongly stimulated by addictive drugs. Through these mechanisms, repeated drug exposure alters functional and structural neuroplasticity, resulting in transition to the addicted biological state and behavioral outcomes that typify addiction. Ca2+ and cAMP represent key second messengers that initiate signaling cascades, which regulate synaptic strength and neuronal excitability. Protein phosphorylation and dephosphorylation are fundamental mechanisms underlying synaptic plasticity that are dysregulated by drugs of abuse. Increased understanding of the regulatory mechanisms by which protein kinases and phosphatases exert their effects during normal reward learning and the addiction process may lead to novel targets and pharmacotherapeutics with increased efficacy in promoting abstinence and decreased side effects, such as interference with natural reward, for drug addiction

The Role of M1 Muscarinic Cholinergic Receptors in the Discriminative Stimulus Properties of N-Desmethylclozapine and the Atypical Antipsychotic Drug Clozapine in Rats

Rationale: The discriminative stimulus properties of clozapine (CLZ) have been studied for decades because it remains the prototype for atypical antipsychotic drug effects and yet is unique in many ways, including increased efficacy in treatment-resistant schizophrenia and in reducing suicidality. Recent studies have indicated that the active CLZ metabolite N-desmethylclozapine (NDMC) may play a role in mediating the cognitive efficacy of CLZ and may also have atypical antipsychotic properties. Objectives: The present study sought to determine if NDMC has discriminative stimulus properties similar to that of its parent drug CLZ. Materials and methods: Rats were trained to discriminate 1.25 mg/kg CLZ from vehicle in a two-choice drug discrimination task. Results: Although NDMC (2.5-20.0 mg/kg) failed to substitute for CLZ, the combination of NDMC (5.0 and 10.0 mg/kg) with a low dose (0.3125 mg/kg) of CLZ produced full substitution (\u3e80% CLZ-appropriate responding) for the 1.25 mg/kg CLZ training dose. Co-administration of the M1-preferring receptor antagonist trihexyphenidyl (6.0 mg/kg) with a 5.0 mg/kg dose of NDMC produced partial substitution (\u3e60% to \u3c80% CLZ-appropriate responding) for CLZ, while administration of trihexyphenidyl alone (0.3-12.0 mg/kg) failed to substitute for CLZ. Conclusions: These findings suggest that NDMC produces discriminative stimulus effects that are different from those elicited by its parent drug CLZ. This difference may be due to the agonist properties of NDMC at M1 muscarinic cholinergic receptors

The Effects of Acute and Repeated Nicotine Doses On Spontaneous Activity in Male and Female Sprague Dawley Ratsanalysis of Brain Area Epibatidine Binding and Cotinine Levels

Previous research in this laboratory has shown that nicotine\u27s effects on spontaneous activity are contingent on individual differences, attenuating activity in high active rats and increasing it in low active rats. This study was designed to further evaluate this phenomenon, and to compare it with nicotine\u27s effects on nicotinic acetylcholine receptor (nAChR) expression in several brain regions. Male and female Sprague-Dawley rats selected for differences in baseline activity were administered nicotine twice daily for 14 days, and its effects on spontaneous activity were evaluated following 1, 13 and 27 doses. Furthermore, [3H] epibatidine binding and plasma cotinine levels were evaluated 24 h after the 28th dose. Contrary to previous findings, the effects of repeated nicotine on spontaneous activity were minimally contingent on baseline activity levels. Following an initial attenuation, males, but not females, exhibited sensitization to nicotine\u27s effects on spontaneous activity. [3H] epibatidine was significantly increased in several brain regions in both male and female nicotine-treated animals, and in females selected for high activity at baseline. However, a clear relationship between these effects and spontaneous activity was not found, due to the lack of consistent effects of nicotine administration and baseline activity on spontaneous activity. Interestingly, significant correlations suggest that rats exhibiting higher spontaneous activity on the final test day were differentially marked by higher [3H] epibatidine. Cotinine levels were higher in low activity males than in high activity males, but no differences were observed between high and low activity females. Thus, no clear relationship between this variable and spontaneous activity could be discerned. Based on these data, no simple relationships between the effects of nicotine administration or baseline activity on [3H] epibatidine binding, nicotine metabolism, or spontaneous activity were observed. However, a relationship between [3H] epibatidine and spontaneous activity on the final test day is suggested

Discriminative Stimulus Properties of 1.25 Mg/Kg Clozapine in Rats: Mediation by Serotonin 5-HT2 and Dopamine D4 Receptors

The atypical antipsychotic drug clozapine remains one of most effective treatments for schizophrenia, given a lack of extrapyramidal side effects, improvements in negative symptoms, cognitive impairment, and in symptoms in treatment-resistant schizophrenia. The adverse effects of clozapine, including agranulocytosis, make finding a safe clozapine-like a drug a goal for drug developers. The drug discrimination paradigm is a model of interoceptive stimulus that has been used in an effort to screen experimental drugs for clozapine-like atypical antipsychotic effects. The present study was conducted to elucidate the receptor-mediated stimulus properties that form this clozapine discriminative cue by testing selective receptor ligands in rats trained to discriminate a 1.25 mg/kg dose of clozapine from vehicle in a two choice drug discrimination task. Full substitution occurred with the 5-HT2A inverse agonist M100907 and the two preferential D4/5-HT2/α1 receptor antagonists Lu 37-114 ((S)-1-(3-(2-(4-(1H-indol-5-yl)piperazin-1-yl)ethyl)indolin-1-yl)ethan-1-one) and Lu 37-254 (1-(3-(4-(1H-indol-5-yl)piperazin-1-yl)propyl)-3,4-dihydroquinolin-2(1H)-one). Partial substitution occurred with the D4 receptor antagonist Lu 38-012 and the α1 adrenoceptor antagonist prazosin. Drugs selective for 5-HT2C, 5-HT6 muscarinic, histamine H1, and benzodiazepine receptors did not substitute for clozapine. The present findings suggest that 5-HT2A inverse agonism and D4 receptor antagonism mediate the discriminative stimulus properties of 1.25 mg/kg clozapine in rats, and further confirm that clozapine produces a complex compound discriminative stimulus

Generalization Testing with Atypical and Typical Antipsychotic Drugs in Rats Trained to Discriminate 5.0 Mg/Kg Clozapine from Vehicle in a Two-Choice Drug Discrimination Task

Clozapine (CLZ) drug discrimination is used as a preclinical model to evaluate compounds for putative atypical antipsychotic properties. In rats, a 1.25 mg/kg CLZ training dose appears to have greater pharmacological specificity for atypical antipsychotic drugs than the traditional 5.0 mg/kg CLZ training dose; however, methodological differences among studies have precluded a direct comparison between these training doses. In the present study, rats were trained to discriminate a 5.0 mg/kg CLZ dose from vehicle in a two-choice drug discrimination task using methods similar to those in a previous study from our laboratory that used a 1.25 mg/kg CLZ training dose. Clozapine produced full substitution (≥80% CLZ-lever responding) for itself at the training dose (5.0 mg/kg). The atypical antipsychotics olanzapine, quetiapine, and ziprasidone also produced full substitution for 5.0 mg/kg CLZ, whereas the atypical antipsychotics risperidone and sertindole produced partial substitution (≥60% CLZ-lever responding). The typical antipsychotic, thioridazine, produced full substitution for the 5.0 mg/kg CLZ training dose, but the typical antipsychotics chlorpromazine, fluphenazine, and haloperidol failed to substitute for clozapine. In a subgroup of 1.25 mg/kg CLZ-trained rats, ziprasidone produced strong partial substitution (73.0% CLZ-lever responding) for the 1.25 mg/kg CLZ training dose. Based on these findings, some atypical antipsychotic drugs (i.e., quetiapine and ziprasidone) produce full substitution only for the 5.0 mg/kg CLZ training dose, whereas other atypical antipsychotic drugs (i.e., sertindole and risperidone) produce full substitution only for the 1.25 mg/kg CLZ training dose. Thus, both of these training doses are important for the screening of putative atypical antipsychotic drugs with the clozapine drug discrimination assay