Adam J. Prus

Discriminative stimulus properties of 1.25 and 5.0 mg/kg doses of clozapine in rats: examination of the role of dopamine, serotonin, and muscarinic receptor mechanisms

Clozapine (CLZ), an atypical antipsychotic drug (APD), produces minimal extrapyramidal side effects (EPS) and has significant advantages for treating both positive and negative symptoms in schizophrenic patients. CLZ has been established as a discriminative cue in the drug discrimination paradigm and in generalization tests the CLZ cue is more selective for atypical, rather than typical, APDs. However, greater selectivity for atypical antipsychotics has been demonstrated with a lower (1.25 mg/kg) CLZ training dose in rats [Psychopharmacology, 149 (2000) 189], rather than the traditional, higher training dose (5.0 mg/kg). It is therefore of interest to evaluate the properties mediating the 1.25 mg/kg CLZ discriminative cue. In the present study, rats were trained to discriminate either 1.25 mg/kg (N=7) or 5.0 mg/kg (N=7) CLZ from vehicle in a two-lever drug discrimination task. The typical antipsychotic haloperidol (0.1-0.4 mg/kg) did not substitute for either CLZ cue, whereas the atypical antipsychotic melperone (0.37-3.0 mg/kg) provided full substitution in both groups (>80% CLZ-appropriate responding). The 5-HT(1A) receptor agonist (+)-8-OH-DPAT (0.04-0.16 mg/kg), and the selective 5-HT(2A) receptor antagonist M100907 (0.03-1.0 mg/kg) did not produce substitution in either group. (+)-8-OH-DPAT combined with haloperidol (0.05 mg/kg) engendered only partial substitution (>60% CLZ-appropriate responding) for both CLZ cues, and M100907 combined with haloperidol (0.05 and 0.1 mg/kg doses) failed to provide substitution in either group. Trihexyphenidyl (0.18-6.0 mg/kg), a muscarinic M(1)-preferring receptor antagonist, engendered full substitution for the 1.25 mg/kg CLZ cue, but only partial substitution for the 5.0 mg/kg CLZ cue. These results provide evidence that antagonism at the muscarinic M(1) receptor is sufficient to provide 1.25 mg/kg CLZ-like discriminative stimulus effects.

The neurotensin analog NT69L enhances medial prefrontal cortical dopamine and acetylcholine efflux: Potentiation of risperidone-, but not haloperidol-, induced dopamine efflux

NT69L is a novel neurotensin (8-13) analog that produces atypical antipsychotic-like effects in animal models. Because atypical antipsychotic drugs increase dopamine (DA) and acetylcholine (ACh) efflux in the medial prefrontal cortex and DA efflux in the nucleus accumbens, the present study sought to further evaluate the putative antipsychotic-like effects of NT69L by assessing DA and ACh efflux in these regions. Dual probe microdialysis was conducted in awake freely moving male rats, without using an acetylcholinesterase inhibitor in the perfusion medium. NT69L (1.0 and 3.0 mg/kg) produced significant increases in extracellular DA and ACh efflux in the medial prefrontal cortex. NT69L (1.0 mg/kg, but not 3.0 mg/kg) produced a significant increase of DA, but not ACh, efflux in the nucleus accumbens. Pretreatment with the serotonin (5-HT)(1A) receptor antagonist WAY100635 (0.2 mg/kg) significantly attenuated the 3.0 mg/kg NT69L-induced increase in medial prefrontal cortical DA efflux. Pretreatment with NT69L (1.0 mg/kg) significantly potentiated the effects of the atypical antipsychotic drug risperidone (0.1 mg/kg) on DA, but not ACh, efflux in the medial prefrontal cortex, while pretreatment with NT69L 1.0 mg/kg failed to alter the effects of haloperidol (0.1 mg/kg) on DA or ACh efflux in either region. These findings further suggest that NT analogs may be useful alone or adjunctively for the treatment of schizophrenia.

Acute, but not repeated, administration of the neurotensin NTS1 receptor agonist PD149163 decreases conditioned footshock-induced ultrasonic vocalizations in rats.

Neurotensin is an endogenous neuropeptide that has significant interactions with monoamine neurotransmitter systems. To date, neurotensin NTS1 receptor agonists, such as PD149163, have been primarily evaluated for the treatment for schizophrenia, drug addiction, and pain. Recently, PD149163 was found to attenuate fear-potentiated startle in rats, an experimental procedure used for screening anxiolytic drugs. The present study sought to assess these findings through testing PD149163 in a conditioned footshock-induced ultrasonic vocalization (USV) model. Conditioning was conducted in male Wistar rats using chambers equipped with shock grid floors and an ultrasonic vocalization detector. PD149163 and the 5-HT1A receptor partial agonist buspirone produced a statistically significant reduction of 22kHz USV counts. The typical antipsychotic haloperidol also reduced 22kHz USV counts, but did so at cataleptic doses. Ten days of repeated administration of PD149163 abolished the inhibitory effects of PD149163 on 22kHz USVs. These findings further support an anxiolytic profile for PD149163. However, tolerance to these effects may limit the utility of these drugs for the treatment of anxiety.

Discriminative stimulus properties of atypical and typical antipsychotic drugs: a review of preclinical studies

BackgroundDrug discrimination is an increasingly valuable behavioral assay for the preclinical development of antipsychotic drugs. The majority of studies have used the atypical antipsychotic clozapine because it displays robust discriminative stimulus properties and is the “prototypical” or “gold standard” atypical antipsychotic against which other antipsychotics will undoubtedly be compared for many years.ObjectivesPharmacological mechanisms mediating the discriminative stimulus properties of antipsychotics used as training drugs and the usefulness of drug discrimination for distinguishing typical and atypical antipsychotics were reviewed.ResultsClozapine appears to have a compound cue involving antagonism of two or more receptors. While muscarinic receptor antagonism is a prominent factor for mediation of clozapine’s cue in rats with a 5.0-mg/kg training dose, there are differences in clozapine’s cue with a low training dose and in pigeons and mice. With a low training dose, clozapine has consistently produced full or partial generalization to atypical but not to typical antipsychotics. Although not evaluated as extensively, the atypical antipsychotics quetiapine and ziprasidone also appear to generalize to atypical but not typical antipsychotics. This has not been the case for other antipsychotic drugs (olanzapine, chlorpromazine, haloperidol) used as training drugs.ConclusionsThere are important differences in discriminative stimulus properties both between and within atypical and typical antipsychotics and across species. While low-dose clozapine discrimination in rats appears to provide a more sensitive behavioral assay for distinguishing atypical from typical antipsychotics, the extent to which clozapine’s discriminative stimulus properties are predictive of its antipsychotic effects remains to be determined.

Acute nicotine reduces and repeated nicotine increases spontaneous activity in male and female Lewis rats

The Lewis (LEW) strain of rat appears more sensitive to nicotine than other strains in self-administration, conditioned place preference, and drug discrimination behavioral studies. The present study sought to further evaluate the behavioral effects of chronic nicotine treatment in the LEW strain by assessing spontaneous activity, which has consistently revealed sensitization to chronic nicotine administration in Sprague Dawley (SD) rats. High active and low active male and female LEW rats (N=8 per group) were treated twice daily with either nicotine (0.4 mg/kg, sc) or vehicle for 14 consecutive days. Regardless of baseline activity level or sex, spontaneous activity was significantly decreased, compared to saline-treated rats, after a single nicotine injection. However, spontaneous activity increased in both low- and high-activity rats (both sexes) over the two weeks of nicotine administration to levels that were significantly higher than saline-treated rats. Based on these findings, acute and chronic nicotine administration had greater suppressive and enhancing effects on spontaneous activity in LEW rats compared to other strains of rats previously studied. These results further clarify the behavioral sensitivity of the LEW strain of rat to nicotine exposure and lend credence to the role of genetics in the individual susceptibility to nicotine dependence.

Systemic administration of the neurotensin NTS₁-receptor agonist PD149163 improves performance on a memory task in naturally deficient male Brown Norway rats.

Agonists for the neurotensin NTS₁ receptor consistently exhibit antipsychotic effects in animal models without producing catalepsy, suggesting that NTS₁-receptor agonists may be a novel class of drugs to treat schizophrenia. Moreover, studies utilizing NTS₁ agonists have reported improvements in some aspects of cognitive functioning, including prepulse inhibition and learning procedures, which suggest an ability of NTS₁-receptor agonists to diminish neurocognitive deficits. The present study sought to assess both baseline delay-induced memory performance and the effects of NTS₁-receptor activation on learning and memory consolidation in male Long-Evans and Brown Norway rats using a delayed nonmatch-to-position task radial arm-maze task. In the absence of drugs, Brown Norway rats displayed a significant increase in spatial memory errors following 3-, 7-, and 24-hr delay, whereas Long-Evans rats exhibited an increase in spatial memory errors following only a 7-, and 24-hr delay. With Brown Norway rats, administration of PD149163 before or after an information trial significantly reduced errors during a retention trial after a 24 hr delay. Administration of the NTS(1/2)-receptor antagonist SR142948 prior to the information trial did not affect retention-trial errors. These data are consistent with previous findings that Brown Norway rats have natural cognitive deficits and that they may be useful for assessing putative antipsychotic drugs for cognitive efficacy. Moreover, the results of this study support previous findings suggesting that NTS₁-receptor agonists may improve some aspects of cognitive functioning.

Assessment of attention in male and female Brattleboro rats using a self-paced five-choice serial reaction time task

The Brattleboro rat is a mutant variation of the Long-Evans strain that exhibits negligible central nervous system levels of vasopressin, a neuropeptide that may influence behavioral and cognitive processes. Compared to Long-Evans rats, Brattleboro rats exhibit diminished fear conditioning and have impairments in spatial memory and sensory gating. The present study sought to further evaluate the cognitive profile of vasopressin-deficient rats by studying attention in male and female Brattleboro and heterozygous rats using a self-paced version of the five-choice serial reaction time task. Male Brattleboro rats required significantly more sessions to meet the training criteria than those by heterozygotic and Long-Evans (wild type) rats. Female Brattleboro rats displayed significantly poorer attention accuracy compared to heterozygotic and Long-Evans rats. Taken together, the present findings add further evidence that vasopressin deficiency diminishes cognitive functioning.

Drug discrimination: 30 years of progress

EditorialThe publication of this Special Issue of Psychopharmacol-ogy on the Stimulus Properties of Drugs marks the 30-yearanniversary in 2008 of the founding of the Society forStimulus Properties of Drugs (SSPD). As previouslydescribed by the society’s first three presidents (see Overtonet al. 1999), the decision to create an organization ofresearchers interested in the discriminative stimulus prop-erties of drugs occurred at a meeting in 1978 of theFederation of American Societies for Experimental Biology(FASEB). The first official meeting of SSPD was held a fewmonths later on June 3, 1978 in conjunction with themeeting of the Committee for Problems on Drug Depen-dence (CPDD) in Baltimore, MD, USA. SSPD has metannually since then and SSPD’s 30th anniversary meetingwas held on November 14, 2008 as a satellite meeting at themeeting of the Society for Neuroscience in Washington, D.C.In addition to the annual national meetings of thesociety, a number of international meetings have been heldsince 1978 with the most recent being in 2005 as a satellitemeeting of the European Behavioral Pharmacology Societyin Barcelona, Spain. There also have been a number ofbooks (often based on these symposia) on the stimulusproperties of drugs published over the years (Colpaert andBalster 1988, Colpaert and Slangen 1982, Glennon et al.1991, Lal 1977, Thompson and Pickens 1971) as well as acollection of articles in Volume 64 (No. 2) of Pharmacol-ogy, Biochemistry and Behavior in 1999.What are the stimulus properties of drugs?A very basic definition of a stimulus is something thataffects an organism by causing a response. In experimentalpsychology, stimuli have traditionally been such things astones, lights, etc., and typically, a laboratory situationwould be set up to assess changes in responding thatoccurred when changes in the stimulus were produced. Astimulus is generally defined by its characteristics, orproperties, and again, in a traditional sense, stimulusproperties would consist of such things as the frequencyof a tone or the intensity of a light. Clearly, drugs also canserve as stimuli and the properties of drug stimuli aremediated by the drug’s biological actions on the centralnervous system (e.g., functioning as an agonist or anantagonist at a specific neurotransmitter receptor). Much ofthe early research on the stimulus properties of drugs usedthe state-dependent paradigm (Overton 1974), but in the1970s the drug discrimination paradigm was developed (seereview by Colpaert 1999). As Colpaert (1999) describes(and the articles in this Special Issue mirror), the drugdiscrimination paradigm has largely replaced the state-dependent paradigm in the study of the stimulus propertiesof drugs. As Balster (1988)states,“One of the majorconceptual advances in behavioral pharmacology was theappreciation that drug effects could subserve stimulusproperties in stimulus–response learning theory.” Balsteralso points out that the first major recognition that drugscould function as stimuli and produce responses in organ-isms was in a collection of articles from the proceedings of a

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 1.25mg/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 1.25mg/kg clozapine in rats, and further confirm that clozapine produces a complex compound discriminative stimulus.

Evaluation of the effects of α2 adrenoceptor antagonism with the D2 receptor antagonist raclopride on conditioned avoidance responding in rats.

The α2 adrenoceptor antagonist idazoxan, when combined with a subeffective dose of the D2 receptor antagonist raclopride or other D2 receptor antagonists, produces inhibition of conditioned avoidance responding (CAR) in rats, an effect predictive of antipsychotic effects. In other models, this treatment combination indicates putative atypical antipsychotic effects as well, and has led to a α2/D2 receptor hypothesis for atypicality. However, this hypothesis would be better supported if other α2 adrenoceptor antagonists were investigated and the role of the alternative mechanisms, particularly 5-HT1A receptor agonism, for the behavioral effects of idazoxan were evaluated. This study sought to further test the α2/D2 receptor hypothesis by assessing the effects of α2, D2 and 5-HT1A receptor ligands on CAR in rats. Raclopride significantly reduced CAR. Administration of idazoxan or the α2 adrenoceptor antagonist yohimbine with a subeffective dose of raclopride also significantly reduced CAR. Pretreatment with the 5-HT1A receptor antagonist WAY100635 failed to significantly reverse the inhibition of CAR produced by the idazoxan and raclopride treatment combination. To the extent that 5-HT1A receptor antagonism failed to block the effects of idazoxan in combination with raclopride on CAR, α2 adrenoceptor antagonism alone appears to potentiate the putative antipsychotic effects produced through D2 receptor antagonism.