Non-habituated Rats Research Articles

Evaluation of Parameters Which Influence Voluntary Ingestion of Supplements in Rats.

Drug safety and efficacy studies frequently use oral gavage, but repetitive usage may cause problems. Administration through voluntary ingestion represents an opportunity for refinement. We aimed to develop a protocol for voluntary ingestion of gelatin-based supplements in rats, assessing the influence of age, sex, fasting (4 h), and additives (vanilla, VF; sucralose, S), and to test it in lactating dams. Three-week-old and 5-month-old Sprague-Dawley rats were placed individually in an empty cage containing a gelatin cube and trained daily (5 days/week), recording the day the whole cube was consumed (latency). Rats trained prior to gestation were offered a gelatin containing 250 mg/kg cocoa shell extract (CSE) during lactation. Rats that did not eat the cube after 8 training days were considered non-habituated, with a proportion similar in young males (7.1%), young females (11.1%), and adult females (10.3%), but significantly higher in adult males (39.3%). Excluding non-habituated rats, latency was 2-3 days, without differences between young and adult rats (p = 0.657) or between males and females (p = 0.189). VF or VF + S in the gelatin did not modify latency, while fasting significantly reduced it in females (p = 0.007) but not in males (p = 0.501). During lactation, trained females ate the CSE-gelatin within 1-5 min without litter problems. Conclusions: Acceptance of a gelatin-based supplement is negatively influenced by male sex, facilitated by fasting, and not modified by additives. Training is remembered after 2 months and does not interfere with lactation. Gelatin-based voluntary ingestion is suitable to administer drugs that need to pass through the digestive system, ensuring adequate dosage, and is important to detect non-habituated rats prior to the study. The current protocol may be implemented by training the rats in their own cage.

Exercise-Induced Hypoalgesia Profile in Rats Predicts Neuropathic Pain Intensity Induced by Sciatic Nerve Constriction Injury

The aim of this study was to investigate the predictive value of exercise-induced hypoalgesia (EIH) profile on pain intensity induced by nerve injury in a rat model. EIH was tested by evaluating the percentage of withdrawal responses to a train of 30 mechanical stimuli on the hind paw before and after 180 seconds of exercise on a rotating rod. The rats were grouped into low, medium, and high EIH based on their reduction in the percentage of withdrawal responses before and after exercise. Rats from each group then underwent left sciatic nerve constriction injury. Mechanical allodynia, mechanical hyperalgesia, and heat allodynia were assessed in the affected and contralateral hind paws prior to and 3 and 7 days following the procedure. The low EIH rats demonstrated increased hypersensitivity at baseline and developed significantly more severe heat allodynia, mechanical allodynia, and hyperalgesia 3 and 7 days following the injury compared to the medium and high EIH rats. Moreover, the low EIH rats developed contralateral heat allodynia following the injury. The EIH of habituated and nonhabituated rats was compared to study the role of stress on the hypoalgesic effect. No significant differences were found between the habituated and nonhabituated rats at baseline and 1 and 5 minutes after the exercise. PerspectiveEIH profile was found to be predictive of pain severity following nerve injury. It may suggest that selected patients with faulty pain modulation are at risk for developing chronic pain following injury or surgical procedures. EIH may represent a preoperative means to detect this predisposition and enable proactive management.

WIN55,212-2 impairs non-associative recognition and spatial memory in rats via CB1 receptor stimulation

Endogenous and exogenous cannabinoids modulate learning and memory primarily via the cannabinoid type 1 receptor (CB1R). A variety of experimental procedures has focused on the role of CB1R in various aspects of learning and memory processes. However, the picture still remains unclear as there is a lack of information on the effects of relatively low doses of CB1R agonists in relation to their effects on locomotion. The present study sought to investigate CB1R activation, using a range of relatively low doses of the CB1R agonist WIN55,212-2, on multiple aspects of learning and memory in rats. For this purpose, non-associative learning was examined using the habituation of locomotion paradigm, recognition memory was evaluated with the novel object recognition task, and the radial water maze test was selected to assess rats’ spatial memory. The ability of the CB1R antagonist, SR141716A, to counteract WIN55,212-2-induced behavioral effects was also tested. WIN55,212-2 (0.3, but not 0.03 or 0.1mg/kg) disrupted non-associative learning, different aspects of short- and long-term recognition memory (storage and retrieval) and retention of spatial memory. The 0.3mg/kg dose of WIN55,212-2 also decreased ambulatory, but not vertical (rearing), activity in non-habituated rats. These effects appeared to be CB1R dependent since pretreatment with SR141716A (0.03mg/kg) prevented the WIN55,212-2-induced behavioral effects. The present findings further support and extend the complex impact of exogenous cannabinoids on learning and memory in relation to their effects on locomotion.

Novelty-Induced Emotional Arousal Modulates Cannabinoid Effects on Recognition Memory and Adrenocortical Activity

Although it is well established that cannabinoid drugs can influence cognitive performance, the findings-describing both enhancing and impairing effects-have been ambiguous. Here, we investigated the effects of posttraining systemic administration of the synthetic cannabinoid agonist WIN55,212-2 (0.1, 0.3, or 1.0 mg/kg) on short- and long-term retention of object recognition memory under two conditions that differed in their training-associated arousal level. In male Sprague-Dawley rats that were not previously habituated to the experimental context, WIN55,212-2 administered immediately after a 3-min training trial, biphasically impaired retention performance at a 1-h interval. In contrast, WIN55,212-2 enhanced 1-h retention of rats that had received extensive prior habituation to the experimental context. Interestingly, immediate posttraining administration of WIN55,212-2 to non-habituated rats, in doses that impaired 1-h retention, enhanced object recognition performance at a 24-h interval. Posttraining WIN55,212-2 administration to habituated rats did not significantly affect 24-h retention. In light of intimate interactions between cannabinoids and the hypothalamic-pituitary-adrenal axis, we further investigated whether cannabinoid administration might differently influence training-induced glucocorticoid activity in rats in these two habituation conditions. WIN55,212-2 administered after object recognition training elevated plasma corticosterone levels in non-habituated rats whereas it decreased corticosterone levels in habituated rats. Most importantly, following pretreatment with the corticosterone-synthesis inhibitor metyrapone, WIN55,212-2 effects on 1- and 24-h retention of non-habituated rats became similar to those seen in the low-aroused habituated animals, indicating that cannabinoid-induced regulation of adrenocortical activity contributes to the environmentally sensitive effects of systemically administered cannabinoids on short- and long-term retention of object recognition memory.

Differences in the locomotor-activating effects of indirect serotonin agonists in habituated and non-habituated rats

The indirect serotonin (5-HT) agonist 3,4-methylenedioxymethamphetamine (MDMA) produces a distinct behavioral profile in rats consisting of locomotor hyperactivity, thigmotaxis, and decreased exploration. The indirect 5-HT agonist α-ethyltryptamine (AET) produces a similar behavioral profile. Using the Behavioral Pattern Monitor (BPM), the present investigation examined whether the effects of MDMA and AET are dependent on the novelty of the testing environment. These experiments were conducted in Sprague–Dawley rats housed on a reversed light cycle and tested during the dark phase of the light/dark cycle. We found that racemic MDMA (RS-MDMA; 3mg/kg, SC) increased locomotor activity in rats tested in novel BPM chambers, but had no effect on locomotor activity in rats habituated to the BPM chambers immediately prior to testing. Likewise, AET (5mg/kg, SC) increased locomotor activity in non-habituated animals but not in animals habituated to the test chambers. These results were unexpected because previous reports indicate that MDMA has robust locomotor-activating effects in habituated animals. To further examine the influence of habituation on MDMA-induced locomotor activity, we conducted parametric studies with S-(+)-MDMA (the more active enantiomer) in habituated and non-habituated rats housed on a standard or reversed light cycle. Light cycle was included as a variable due to reported differences in sensitivity to serotonergic ligands during the dark and light phases. In confirmation of our initial studies, rats tested during the dark phase and habituated to the BPM did not show an S-(+)-MDMA (3mg/kg, SC)-induced increase in locomotor activity, whereas non-habituated rats did. By contrast, in rats tested during the light phase, S-(+)-MDMA increased locomotor activity in both non-habituated and habituated rats, although the response in habituated animals was attenuated. The finding that habituation and light cycle interact to influence MDMA- and AET-induced hyperactivity demonstrates that there are previously unrecognized complexities associated with the behavioral effects of these drugs.

Establishment of a novel objective and quantitative method to assess pain-related behavior in monosodium iodoacetate-induced osteoarthritis in rat knee

IntroductionPain in osteoarthritis (OA) patients can be present at rest but typically worsens with movement of the affected joint. However, useful assessment methods of movement-induced pain in animal models are limited. Here, we describe the reduction of spontaneous activity in a rat model of OA as an objective and quantifiable behavioral pain that can predict the analgesic activity of a variety of agents following single-dose administration. MethodsOA was induced in male Sprague–Dawley (SD) rats by intra-articular injection of monoiodoacetate (MIA), and the joint degeneration was assessed with histologic and radiographic analyses. Spontaneous activities were measured in nonhabituated rats using standard, photocell-based monitor systems in the dark. To investigate the potential of the OA model to predict analgesic activity, a number of nonsteroidal anti-inflammatory drugs (NSAIDs) and atypical analgesic drugs were used. ResultsBiphasic reduction of total distance and number of rears was observed during the course of experiment after administering 1mg and 0.3mg of MIA, respectively. We found that number of rears was the most sensitive to MIA-induce OA and displayed the greatest percentage decrease in activity. Joint degeneration was observed with decreased bone mineral density and loss of articular cartilage 28days post-MIA injection. Appropriate dosage of opioids reversed MIA-induced decrease of number of rears indicating that reduction of this vertical spontaneous activity reflects pain-associated behavior. As high-doses of opioids reduced spontaneous activity, the sedative effect can be distinguished from the analgesic effect. Analgesic treatment indicates the coexistence of an inflammatory pain state (early phase) sensitive to NSAIDs and a non-inflammatory pain state (late phase) resistant to NSAID treatment. DiscussionThis study indicates that unlike standard measures of analgesia such as alteration in thermal or mechanical sensitivity, measurement of spontaneous activity is a validated method for measuring the effects of analgesics in rats with OA knee joints. Moreover, the animals require no habituation, and thus behavioral observation subjectivity is eliminated.

I. In vivo evidence for partial agonist effects of (−)-OSU6162 and (+)-OSU6162 on 5-HT2A serotonin receptors

The locomotor effects of (-)- and (+)-OSU6162 were evaluated in 'low activity' animals (reserpinized mice and habituated rats) and 'high activity' animals (drug-naive mice and non-habituated rats). Both enantiomers of OSU6162 had dual effects on behavior, stimulating locomotor activity in 'low activity' animals and inhibiting locomotor activity in 'high activity' animals. There were also certain differences between the two enantiomers in their behavioral profiles. The stimulatory effects of both enantiomers in reserpinized mice were blocked by the 5-HT2A selective antagonist M100907, but not by the D2-selective antagonists haloperidol or raclopride, or by the D1-selective antagonists SCH23390 or SCH39166. The stimulatory effect in mice was more pronounced for (+)- than for (-)-OSU6162. In drug-naive mice, both enantiomers of OSU6162 produced head twitches, albeit to a much lesser extent than DOI, and both enantiomers inhibited DOI-induced head twitches, the (-)-form more effectively so than the (+)-form. These results suggest that (-)- and (+)-OSU6162 are partial agonists on 5-HT2A receptors and that the (+)-form has a higher intrinsic activity than the (-)-form. At high doses, both enantiomers inhibited locomotor activity in drug-naive mice, with (-)-OSU6162 being more potent than (+)-OSU6162. Similarly, in high-active rats, both enantiomers inhibited locomotor activity, with the (-)-enantiomer being more potent than the (+)-enantiomer. Conversely, in habituated rats, both enantiomers stimulated locomotor activity, and here, as opposed to the case in low-active mice, (-)-OSU6162 was more effective than (+)-OSU6162. The stimulatory effects in habituated rats of both enantiomers could be antagonized with either haloperidol or M100907. Overall, these results indicate that the dual effects on behavior of (-)- and (+)-OSU6162 are mediated through D2 and 5-HT2A receptors, consistent with their in vitro functional selectivity profiles (see Burstein et al., accompanying paper). Thus, both enantiomers of OSU6162 seem to act as stabilizers not only on dopaminergic, but also on serotonergic brain signaling. These discoveries have important implications for the potential clinical utility of both compounds, as well as for several of their congeners.

II. In vitro evidence that (−)-OSU6162 and (+)-OSU6162 produce their behavioral effects through 5-HT2A serotonin and D2 dopamine receptors

(-)-OSU6162 has promise for treating Parkinson's disease, Huntington's disease and schizophrenia. Behavioral tests evaluating the locomotor effects of (-) and (+)-OSU6162 on 'low activity' animals (reserpinized mice and habituated rats) and 'high activity' animals (drug naive mice and non-habituated rats) revealed that both enantiomers of OSU6162 had dual effects on behavior, stimulating locomotor activity in 'low activity' animals and inhibiting locomotor activity in 'high activity' animals. To elucidate a plausible mechanism of action for their behavioral effects, we evaluated the intrinsic actions of (-)- and (+)-OSU6162, and a collection of other antipsychotic and antiparkinsonian agents at 5-HT2A and D2 receptors in functional assays with various degrees of receptor reserve, including cellular proliferation, phosphatidyl inositol hydrolysis, GTPγS and beta-arrestin recruitment assays. We also tested for possible allosteric actions of (-)-OSU6162 at D2 receptors. Both enantiomers of OSU6162 were medium intrinsic activity partial agonists at 5-HT2A receptors and low intrinsic activity partial agonists at D2 receptors. (+)-OSU6162 had higher efficacy at 5-HT2A receptors, which correlated with its greater stimulatory activity in vivo, but (-)-OSU6162 had higher potency at D2 receptors, which correlated with its greater inhibitory activity in vivo. (-)-OSU6162 did not display any convincing allosteric properties. Both (+)- and (-)-OSU6162 were significantly less active at 27 other monoaminergic receptors and reuptake transporters tested suggesting that D2 and 5-HT2A receptors play crucial roles in mediating their behavioral effects. Compounds with balanced effects on these two receptor systems may offer promise for treating neuropsychiatric diseases.

A Rodent Model of Schizophrenia Reveals Increase in Creatine Kinase Activity with Associated Behavior Changes

Schizophrenia is a debilitating mental disorder characterized by positive (delusions, hallucinations, disorganized speech) and negative (affective flattering, avolition and social withdrawal) symptoms as well as cognitive deficits. The frequency, severity and topography characterize the disorder as heterogeneous, the pathophysiology of schizophrenia is poorly understood. Sub-anesthetic doses of ketamine produce hyperactivity, stereotypy and abnormal social interaction and it is used as a model of schizophrenia. In this study, we induced an animal model by acute sub-anesthetic doses of ketamine and tested different behavioral parameters. We also evaluated the activity of creatine kinase (CK) in brain of rats treated with ketamine. Our results demonstrated that administration of 10, 25 and 50 mg/kg of ketamine induced an increase of covered distance in habituated and non-habituated rats to the behavioral apparatus. Ketamine administration induced significant social deficits and stereotypic behavioral in all doses tested. Finally we evaluated the effect of different doses of ketamine on creatinine kinase (CK) activity and we observed that CK activity is increased inspecific regions of the brain. Our study suggests that our animal model may be used as a model of schizophrenia and that cerebral energy metabolism might be altered in the brain of schizophrenic patients, probably leading to alterations that might be involved in the pathogenesis of schizophrenia.

W1725 Long Term Capsaicin Ingestion Decreases Intestinal Mechano- and Chemosensibility in Healthy Subjects

Endogenous and exogenous cannabinoids modulate learning and memory primarily via the cannabinoid type 1 receptor (CB1R). A variety of experimental procedures has focused on the role of CB1R in various aspects of learning and memory processes. However, the picture still remains unclear as there is a lack of information on the effects of relatively low doses of CB1R agonists in relation to their effects on locomotion. The present study sought to investigate CB1R activation, using a range of relatively low doses of the CB1R agonist WIN55,212-2, on multiple aspects of learning and memory in rats. For this purpose, non-associative learning was examined using the habituation of locomotion paradigm, recognition memory was evaluated with the novel object recognition task, and the radial water maze test was selected to assess rats’ spatial memory. The ability of the CB1R antagonist, SR141716A, to counteract WIN55,212-2-induced behavioral effects was also tested. WIN55,212-2 (0.3, but not 0.03 or 0.1 mg/kg) disrupted non-associative learning, different aspects of short- and long-term recognition memory (storage and retrieval) and retention of spatial memory. The 0.3 mg/kg dose of WIN55,212-2 also decreased ambulatory, but not vertical (rearing), activity in non-habituated rats. These effects appeared to be CB1R dependent since pretreatment with SR141716A (0.03 mg/kg) prevented the WIN55,212-2-induced behavioral effects. The present findings further support and extend the complex impact of exogenous cannabinoids on learning and memory in relation to their effects on locomotion.

Arousal and Stress Effects on Consolidation and Reconsolidation of Recognition Memory

This study examined the effects of the arousal level of the rat and exposure to a behavioral stressor on consolidation and reconsolidation of a nonaversive learning paradigm, the object recognition task. Learning was tested under two arousal conditions: no previous habituation to the experimental context (high novelty stress/arousal level) or extensive prior habituation (reduced novelty stress/arousal level). Results indicated that in the habituated rats, exposure to an out-of-context stressor (ie, elevated platform stress) impaired long-term consolidation and reconsolidation of object recognition. RU-486, a glucocorticoid receptor (GR) antagonist, infused into the basolateral amygdala (BLA), reversed the impairing effects of the stressor. In contrast, the nonhabituated aroused rats were impaired when consolidation was examined, but their memory was intact following reactivation of the memory trace. Exposure of nonhabituated rats to an out-of-context stressor enhanced the long-term consolidation of recognition memory, but impaired reconsolidation, and the effects were reversed by a GR antagonist infused into the BLA. Additionally, nonhabituated control rats showed intact retrieval following microinfusion of propranolol to the BLA immediately after the training, suggesting an involvement of beta-adrenoceptors in the BLA in the arousal-induced impairment of consolidation. These findings demonstrate opposite effects, detrimental and facilitative, of arousal and stress on memory consolidation and reconsolidation. In addition, the data suggests that although some general features underlie consolidation and reconsolidation, there is a possible dissimilarity between the two processes, which is dependent on the arousal level of the animal during training.

Inflammation-Induced Reduction of Spontaneous Activity by Adjuvant: A Novel Model to Study the Effect of Analgesics in Rats

The majority of rodent models used to evaluate analgesic drug effects rely on evoked measures of nociceptive thresholds as primary outcomes. These approaches are often time-consuming, requiring extensive habituation sessions and repeated presentations of eliciting stimuli, and are prone to false-positive outcomes due to sedation or tester subjectivity. Here, we describe the reduction of spontaneous activity by adjuvant (RSAA) model as an objective and quantifiable behavioral model of inflammatory pain that can predict the analgesic activity of a variety of agents following single-dose administration. In the RSAA model, activity was measured in nonhabituated rats using standard, photocell-based monitors. Bilateral inflammation of the knee joints by complete Freund's adjuvant (CFA) reduced the normal level of activity (horizontal locomotion and vertical rearing) by approximately 60% in a novel environment. This reduction in activity was dose-dependently reversed by ibuprofen, rofecoxib, celecoxib, piroxicam, and dexamethasone, whereas gabapentin and amitriptyline were inactive. Morphine significantly reversed the activity-suppressing effects of CFA, at 1 mg/kg s.c., but at higher doses locomotor activity progressively declined, coincident with the induction of sedation. In contrast to morphine and anti-inflammatory therapies, amphetamine did not affect vertical rearing, even though it increased horizontal locomotion. Thus, unlike standard measures of analgesia such as alteration in thermal or mechanical sensitivity, the RSAA model operationally defines analgesia as a drug-induced increase in spontaneous behavior (vertical rearing in a novel environment). We conclude that the RSAA model is valuable as an objective measure of analgesic efficacy that is not dependent on an evoked stimulus response.

Hypericum perforatum L (St John's wort) preferentially increases extracellular dopamine levels in the rat prefrontal cortex.

The effects of hydro-alcoholic extracts of Hypericum perforatum L on extracellular serotonin (5-HT), noradrenaline (NA) and dopamine (DA) levels and the acidic metabolites (3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxy-3-indoleacetic acid (5-HIAA)) were examined by in vivo microdialysis in the prefrontal cortex of awake rats. Thus, a single dose (60 mg kg(-1) i.p. or 300 mg kg(-1) p.o.) of H. perforatum increased DA concentrations to 165 and 140% of control values, respectively, and increased locomotor activity in nonhabituated rats. DOPAC and HVA levels were markedly reduced. 5-HT concentrations were elevated only moderately, while the NA levels were not affected by any treatment. The whole-tissue analysis revealed that hypericum increased, whereas the monoamine oxidase (MAO) A/B inhibitor phenelzine decreased DA and 5-HT turnover. The present data indicate that the mechanism of action of hypericum extract in vivo is more complex than the inhibition of monoamine reuptake or metabolism observed in vitro. The finding of preferential enhancement of DA transmission is in agreement with human studies measuring DA-mediated neuroendocrine responses.

Glucocorticoid effects on object recognition memory require training-associated emotional arousal.

Considerable evidence implicates glucocorticoid hormones in the regulation of memory consolidation and memory retrieval. The present experiments investigated whether the influence of these hormones on memory depends on the level of emotional arousal induced by the training experience. We investigated this issue in male Sprague-Dawley rats by examining the effects of immediate posttraining systemic injections of the glucocorticoid corticosterone on object recognition memory under two conditions that differed in their training-associated emotional arousal. In rats that were not previously habituated to the experimental context, corticosterone (0.3, 1.0, or 3.0 mg/kg, s.c.) administered immediately after a 3-min training trial enhanced 24-hr retention performance in an inverted-U shaped dose-response relationship. In contrast, corticosterone did not affect 24-hr retention of rats that received extensive prior habituation to the experimental context and, thus, had decreased novelty-induced emotional arousal during training. Additionally, immediate posttraining administration of corticosterone to nonhabituated rats, in doses that enhanced 24-hr retention, impaired object recognition performance at a 1-hr retention interval whereas corticosterone administered after training to well-habituated rats did not impair 1-hr retention. Thus, the present findings suggest that training-induced emotional arousal may be essential for glucocorticoid effects on object recognition memory.

(−)-1-(Benzofuran-2-yl)-2-propylaminopentane enhances locomotor activity in rats due to its ability to induce dopamine release

“Catecholaminergic and serotoninergic activity enhancer” effects are newly found mechanisms of action of a class of compound that enhance impulse propagation-mediated release of catecholamines and serotonin in the brain. In the present study, (−)-1-(benzofuran-2-yl)-2-propylaminopentane hydrochloride [(−)-BPAP HCl], a compound with selective and potent “catecholaminergic and serotoninergic activity enhancer” effects, was tested for its efficacy to potentiate locomotor activity in normal rats and to attenuate hypolocomotion in reserpine-treated rats. (−)-BPAP HCl potentiated locomotor activity in non-habituated rats during a 2-h observation period dose-dependently (0.3–10 mg/kg). (−)-BPAP HCl (1–3 mg/kg) was also effective to reverse reserpine-induced hypolocomotion. The effects of (−)-BPAP HCl in normal and reserpine-treated rats were attenuated by the dopamine D1 receptor antagonist, R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1 H-3-benzazepine (SCH 23390), suggesting that the effects of (−)-BPAP HCl were mediated by activation of the dopaminergic system. In addition, the administration of (−)-BPAP HCl increased ipsilateral turning in unilaterally 6-hydroxydopamine-lesioned rats, implying presynaptic activation of nigrostriatal dopaminergic terminals by (−)-BPAP HCl. Furthermore, although antiparkinsonian agents, such as apomorphine and amantadine, failed to improve reserpine-induced ptosis, (−)-BPAP HCl significantly improved ptosis. These findings suggested that a “catecholaminergic and serotoninergic activity enhancer” compound, (−)-BPAP, stimulates motor function in rats and improves motor deficits in animal models of Parkinson's disease due to its ability to induce dopamine release.

The effects of δ agonists on locomotor activity in habituated and non-habituated rats

The effects of the δ agonists SNC80 and deltorphin II on ambulation and rearing activity were measured in habituated and non-habituated rats. SNC80 (30, 100, 200, 400 nmol, i.c.v.) and deltorphin II (3, 15, 30, 60 nmol, i.c.v.) induced similar, dose-dependent biphasic locomotor effects in non-habituated subjects. An initial decrease in exploratory activity was associated with anxiogenic signs such as pilo-erection, freezing behaviour and pupil dilation for each drug. Pre-treatment with the δ antagonist naltrindole (10 nmol, i.c.v.) inhibited the depressant effect, but not the subsequent stimulant effect, on locomotor activity in response to 30 nmol deltorphin II in this assay ( P<0.05). In habituated rats, deltorphin II (0.03, 0.1, 0.3, 3 nmol, i.c.v.) caused significant, naltrindole-reversible increases in locomotor activity ( P<0.05 for all doses) at 1000-fold lower doses than those required for a similar response to SNC80 (10, 30, 100, 300 nmol, i.c.v.). Pharmacokinetic studies suggest that these compounds penetrate the brain to similar extents following i.c.v. injection. The substantial potency difference between deltorphin II and SNC80 in stimulating locomotor activity in habituated rats suggests pharmacological heterogeneity for these δ opioid receptor agonists.

Central administration of dopamine D3 receptor antisense to rat: effects on locomotion, dopamine release and [3H]spiperone binding.

A 15-mer, all-phosphorothioate-modified antisense oligodeoxynucleotide (ASO) targeted against rat dopamine D3 receptor mRNA (4 microM, 5 days) significantly reduced (28%) the amount of binding sites labelled with [3H]spiperone in monolayer cultured Chinese hamster ovary (CHO) cells transfected with the complementary desoxyribonucleic acid (cDNA) for the rat D3 receptor. In contrast, D3-ASO treatment did not reduce the amount of bound [3H]spiperone in CHO cells transfected with D2 receptor cDNA. Intracerebroventricular infusion of D3-ASO (osmotic minipump, 10 microg/microl/h, 7 days) influenced dopamine receptor density in the limbic forebrain such that the upper part of the dopamine/[3H]spiperone displacement curve--tentatively representing the D3 receptor--was altered significantly. Spontaneous locomotor activity of non-habituated rats was increased significantly in D3-ASO-treated animals; in addition, in vivo microdialysis revealed a moderate increase in dopamine release in the nucleus accumbens in these animals. In all experiments, an oligodeoxynucleotide comprising the same nucleotides as the antisense sequence, but in random order, was used as control. It is concluded that the antisense strategy is useful for investigating the functional role of dopamine D3 receptors and that the dopamine D3 receptor is involved in rat locomotor behaviour.

Pharmacological Characterization of PD 152255, a Novel Dimeric Benzimidazole Dopamine D 3 Antagonist

152255 ( E-1, 1′-(2-butene-1,4-diyl)bis [2-[4-[3-(1-piperidinyl)propoxy]-phenyl]-1H-benzimidazole]) exhibited high affinity ( K i = 12.7 nM) for human dopamine (DA) D 3 receptors expressed in CHO K1 cells but not for DA D 2L receptors ( K i = 565 nM), DA D 4.2 or DA D 1 receptors ( K i > 3 μM) and a number of other neurotransmitter receptors. Affinity for human muscarinic receptors was seen in vitro but no functional muscarinic agonist and/or antagonist action was observed in vivo. Antagonist activity at DA D 3 receptors was demonstrated by blockade of quinpirole-stimulated [ 3H]-thymidine uptake in D 3 transfected cells, an effect that was 28-fold more potent than in D 2-transfected cells. Unlike classical DA D 2 antagonists, PD 152255 did not increase rat brain DA synthesis and it increased locomotion in habituated rats. However, like antipsychotics, PD 152255 reduced locomotor activity in mice and reduced spontaneous and amphetamine-stimulated locomotion in nonhabituated rats. These results demonstrate that PD 152255 is a DA D 3 antagonist that may have antipsychotic activity.

Effects of intra‐accumbens blockade of NMDA receptors in two models of anxiety, in rats.

In this study the anxiolytic-like effects of intra-accumbens administration of (+)-MK-801, [(+)-5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine hydrogen maleate; noncompetitive N-methyl-D-aspartate receptor antagonist] and AP-7, [(+)-2-amino-7-phosphonoheptanoic acid; competitive NMDA receptor antagonist] in the test of neophobia and the Vogel conflict test, were assessed in rats. Both compounds, although in a limited dose range, disinhibited exploratory activity of non-habituated rats and increased responding suppressed by punishment, independently from changes in animals motility. The obtained results indicate an anxiolytic-like effect of local inhibition of NMDA receptor complex within this brain limbic structure, and support the proposal that accumbal glutamate innervation may be involved in processing of emotional input.

Effects of antagonists at the NMDA receptor complex in two models of anxiety

The effects of an antagonist at the strychnine insensitive glycine site (5,7-dichlorokynurenic acid, i.c.v.), and of noncompetitive (MK-801, i.p.) and competitive (CGP 37849, i.p.; CGP 39551, i.p.; AP-7, i.c.v.) NMDA antagonists were compared with diazepam (i.p.) in two animal models of anxiety (the open field exploratory behavior of non-habituated rats, and the Vogel conflict test). All drugs when applied in appropriate doses increased punished drinking in the Vogel test, without producing any significant changes in free drinking and the stimulus threshold at their lowest anticonflict doses. The effective doses were as follows: diazepam 1.5 and 2.5 mg/kg; MK-801 0.005 and 0.01 mg/kg; CGP 39551 5.0 and 20.0 mg/kg; CGP 37849 1.0 and 2.5 mg/kg; 5,7-dichlorokynurenic acid 5.0 μg (i.c.v); AP-7 0.5 μg (i.c.v.). In the open field diazepam (0.05 mg/kg), MK-801 (0.1 mg/kg), CGP 37849 (0.01, 0.1, 1.0 mg/kg), and AP-7 2.5 μg (i.c.v.) significantly increased exploratory activity in the central sectors of the open field (anti-neophobic reaction), without changing motor activity of the rat. MK-801 at the highest tested dose of 0.2 mg/kg significantly stimulated animal locomotor activity. CGP 37849 in the largest dose examined (10 mg/kg) significantly depressed the motor behavior of rats. Overall, it appeared that different NMDA antagonists showed an anxiolytic-like profile, similar to that of the benzodiazepine diazepam. Among different NMDA receptor complex antagonists studied, CGP 37849 was characterized by the largest distinction between the doses showing an anxiolytic-like action in the open field test, and changing rat motor behavior.