Distinct pattern of c-fos mRNA expression after systemic and intra-accumbens amphetamine and MK-801.

Pharmacological manipulation of both dopamine and glutamate systems affects motor responses in laboratory animals. The two systems, however, seem to act in opposite ways, since direct or indirect activation of dopamine receptors induces similar stimulatory effects to those seen following blockade of N-methyl-D-aspartate receptors. In the present study we compared the pattern of c-fos activation induced by systemic and intra-accumbens administration of the non-competitive N-methyl-D-aspartate antagonist MK-801 and the indirect dopamine agonist amphetamine. Systemic MK-801 induced c-fos mRNA expression in the motor cortex and preferentially in the motor thalamus, i.e. ventrolateral nucleus. Systemic amphetamine, on the other hand, enhanced c-fos mRNA expression in the shell of the accumbens and in limbic thalamic nuclei such as the anteroventral and anterodorsal nuclei. The main effect observed after intra-accumbens administrations of either drug was enhanced c-fos expression in the thalamus, somewhat similar to what seen following systemic administration. In fact also in this case there was a preferential activation of the limbic thalamus by amphetamine and the motor thalamus by MK-801. The present results confirm that different neural substrates underlie behavioral effects induced by systemic administrations of N-methyl-D-aspartate receptor antagonists and dopamine agonists. Further they suggest that intra-accumbens manipulation of the two neural systems could affect different efferent pathways from this structure activating different thalamic targets.

The effects of isolation rearing on glutamate receptor NMDAR1A mRNA expression determined by in situ hybridization in Fawn hooded and Wistar rats.

Rats reared in social isolation exhibit a syndrome of behavioral and biochemical effects indicative of enhanced mesolimbic dopamine (DA) function. The precise nature of the neurodevelopmental changes that produce this state are unknown but result in enhanced DA neurotransmission in the nucleus accumbens (NAC). It was hypothesized that this may be the indirect result of chronic changes in glutamate NMDA receptor function. The same prediction has been made for Fawn hooded (FH) rats that exhibit some of the characteristic effects of isolation-reared rats when compared to Wistar rats. Therefore, mRNA levels of the NMDAR1A receptor subunit were determined by in situ hybridization and were quantified in the striatum, hippocampus and prefrontal cortex of FH and Wistar rats. Isolation rearing alone was not found to have an effect on the expression of NMDAR1A, while FH rats had reduced levels across most brain regions examined. In some areas of the striatum and prefrontal cortex, this effect was greater in FH isolates than in FH socials, while in the hippocampus, the opposite was observed.

Locomotor-stimulating effects of indirect dopamine agonists are attenuated in Fawn hooded rats independent of postweaning social experience.

The effects of the indirect dopamine (DA) agonists cocaine and D-amphetamine on locomotor activity were examined in Fawn hooded (FH) rats and Wistar rats. The effect of isolation rearing was also examined to determine if it might have different effects in these two strains. Contrary to previous findings in other rat strains, only small increases in locomotor-stimulating responses to low doses of cocaine were observed in the present study as a result of isolation rearing. However, at higher cocaine doses, locomotor activity was substantially attenuated in FH rats relative to Wistar rats. A similar pattern of effects was observed for amphetamine in FH rats but only at the intermediate dose. The effects of strain and rearing were independent. There was no evidence for interactions between these factors.

Enhanced corticosterone release after a modified forced swim test in Fawn hooded rats is independent of rearing experience.

Recent findings have demonstrated that Fawn hooded (FH/Har) rats exhibit enhanced plasma corticosterone (CORT) responses compared to Wistar rats after exposure to an open field, whereas this effect was not influenced by early social experience. In contrast, it was found that behavior in a modified version of the Porsolt Forced Swim Test (Porsolt FST) was affected by both strain and social experience. An important part of this study included modifications of the Porsolt FST that allowed separation of multiple behavioral endpoints. The present experiment was conducted to determine if FH/Har rats also exhibit enhanced CORT responses after exposure to the modified forced swim test, and whether CORT levels might predict the behavioral response in this context. After the initial exposure in the modified forced swim test FH/Har rats had higher CORT levels than Wistar rats, but this difference was not affected by isolation rearing. However, CORT levels were not correlated with the main behavioral measures assayed in this test. Nonetheless, the data confirm that FH/Har rats have altered HPA axis responses to stressors.

Locomotor activity induced by the non-competitive N-methyl-D-aspartate antagonist, MK-801: role of nucleus accumbens efferent pathways.

We have recently shown that focal administration of dizocilpine hydrogen maleate (MK-801, a non-competitive N-methyl-D-aspartate antagonist) within the nucleus accumbens increases locomotor activity in a dopamine-independent manner. The purpose of this study was to investigate the neural network underlying locomotor stimulation induced by N-methyl-D-aspartate receptor blockade in the accumbens. In the first experiment, we examined the effect of different doses (1, 5 and 25 nmol) of the active and inactive enantiomers of the N-methyl-D-aspartate antagonist, (+)- and (-)-MK-801, respectively, focally administered in the nucleus accumbens. Only the active enantiomer induced a significant increase in locomotor activity; furthermore, the effect induced by the two highest doses of (+)-MK-801 was significantly different from that induced by (-)-MK-801. In the second part of the study, we performed ibotenic acid lesions to the major output nuclei of the accumbens, the ventral pallidum, mediodorsal thalamus, ventrolateral/ventromedial thalamus and pedunculopontine tegmental nucleus, to observe their effect on locomotor activity induced by focal (+)-MK-801 (25 nmol) administration into the accumbens. None of the lesions had any effect on spontaneous locomotor activity. Hyperactivity induced by accumbens MK-801 administrations was unaffected by ibotenic acid lesions of the pedunculopontine tegmental nucleus, while lesions of the mediodorsal thalamus induced only a partial inhibition. In contrast, ibotenic acid lesions of the ventral pallidum and ventrolateral/ventromedial thalamus completely blocked the motor response induced by accumbens MK-801. These data indicate that the intact mediodorsal thalamus, which has been proposed as a part of the loop that relays accumbens information to the prefrontal cortex, does not seem to be a structure of primary importance in MK-801 locomotor activity. On the contrary, the motor nuclei of the thalamus appear to play a more relevant role, suggesting that different neural substrates may mediate dopamine and glutamate functional output from the nucleus accumbens.

Differential basis of strain and rearing effects on open-field behavior in Fawn Hooded and Wistar rats.

Open-field behavior was examined under several conditions in isolation-reared, and socially reared, Fawn Hooded (FH) and Wistar rats. Lighting conditions (red or white light) and complexity (object or no object) were varied: Experiment 1, white light, no object; Experiment 2, red light, no object; Experiment 3, white light, object; Experiment 4, red light, object. The plasma corticosterone (CORT) response to open-field exposure was examined two further experiments. Observation of differences in open-field behavior, resulting from strain or rearing condition, was dependent on both lighting condition and complexity. Differences in exploratory behavior exhibited by isolation-reared rats were best explained by changes in response to novelty, while those in FH, relative to Wistar, rats were primarily due to increased anxiety. Supporting these conclusions, FH rats had enhanced stimulated CORT levels, while isolation rearing was without effect.

Sensitization components of post-traumatic stress disorder: implications for therapeutics.

Behavioral sensitization to psychomotor stimulants and stressors involves increasing reactivity to repetition of the same dose (or intensity) of the stimulus over time. In cocaine-induced behavioral sensitization, conditioned components are prominent and the underlying pharmacological and neuroanatomical substrates differ as a function of stage of its evolution. Since there is cross-sensitization between cocaine-induced behavioral sensitization and stress-induced behavioral sensitization and, as well, a high comorbidity of stimulant abuse with post-traumatic stress disorder, many of the mechanisms and principles of cocaine-induced behavioral sensitization may be pertinent to post-traumatic stress disorder. The extent and duration of cocaine-induced behavioral sensitization/stress-induced behavioral sensitization are a function of age, genetic strain, and gender of the organism, as well as magnitude, frequency, duration, quality, and environmental contingencies of the inducing stimuli. These effect a spatiotemporal evolution of cascades of alterations in gene expression involving a host of immediate early and late effector genes, including neurotrophic and apoptotic factors. They may change the physiology, biochemistry, and neuroanatomy of the developing brain and, in adults, substrates critical for learning and memory. Suggestions are made for differential strategies of therapeutic intervention as primary prevention, in the immediate post-traumatic period, and in the late phases of post-traumatic stress disorder evolution.

Effects of 5,7-dihydroxytryptamine depletion of tissue serotonin levels on extracellular serotonin in the striatum assessed with in vivo microdialysis: relationship to behavior.

Effects of i.c.v. administration of 5,7-dihydroxytryptamine (5,7-DHT) on biochemistry and behavior were studied in awake Sprague-Dawley rats. It was found that 5,7-DHT depletion of striatal tissue levels of serotonin (5-HT) does not diminish extracellular levels until substantial depletions occur. This finding is similar to those observed after 6-hydroxydopamine lesions of the brain dopamine systems. Although varying amounts of 5,7-DHT produced serotonin depletions in striatal tissue, decreases in extracellular levels were only observed at tissue depletions greater than 60% compared to saline-injected control subjects. Thus, the effects of serotonin lesions which produce only moderate depletions may not be the result of decreased extracellular serotonin, but instead may be the result of compensatory changes in remaining neurons which maintain normal extracellular serotonin concentrations. Different degrees of striatal serotonin depletion were associated with opposite behavioral effects. Moderate levels of serotonin depletion (50-75%) produced evidence of increased anxiety, while these effects were no longer seen in rats with more severe 5-HT depletions (>75%).

Effects of isolation-rearing on locomotion, anxiety and responses to ethanol in Fawn Hooded and Wistar rats.

Voluntary ethanol (EtOH) consumption is increased by isolation-rearing in several rat strains. The following experiments examined the effects of isolation-rearing on basal and ethanol-stimulated behavior in Fawn Hooded rats, an alcohol-preferring rat strain, compared to Wistar rats. Locomotor activity and anxiety were examined under both conditions. Basal locomotor activity was higher in isolated subjects of both strains in low light conditions, but under bright light conditions, this difference was only observed in Wistar rats. Locomotor stimulant effects of EtOH were only observed in isolation-reared rats. In the elevated plus maze, Fawn Hooded rats were more anxious than Wistar rats under low light conditions, but under bright light conditions, Wistar socials were less anxious than all of the other groups. Administration of 1.5 mg/kg EtOH produced an anxiolytic response in the elevated plus maze under bright light conditions in Fawn Hooded rats, but to a lesser degree Wistar rats, particularly Wistar isolates. In conclusion, although both strain and isolation-rearing had effects on locomotion and anxiety as well as the stimulatory and anxiolytic effects of EtOH, these effects appeared to be independent.

Effects of isolation-rearing on voluntary consumption of ethanol, sucrose and saccharin solutions in Fawn Hooded and Wistar rats.

These experiments examined the hypothesis that isolation-rearing and strain influence hedonic mechanisms. In experiment 1, voluntary consumption of ethanol and water was monitored in the home cage of Fawn Hooded (FH) and Wistar rats. FH rats were found to consume more ethanol at low concentrations than Wistar rats, independent of rearing condition, and isolation-reared rats were found to consume more of high ethanol concentrations, independent of strain. In experiment 2, isolation-reared rats were found to consume more sucrose, independent of concentration, than socially reared rats. In experiment 3, Fawn Hooded rats were found to be more sensitive to low concentration solutions of saccharin, and to consume less of the high concentration solutions, while isolation-rearing was found to enhance consumption of high concentrations. Thus, hedonic processes are independently modulated by strain and rearing conditions, although the effects of isolation-rearing appear to be exacerbated in Fawn Hooded rats.

Conditioned increases in anxiogenic-like behavior following exposure to contextual stimuli associated with cocaine are mediated by corticotropin-releasing factor.

Although cocaine is a powerful reinforcer, it has been reported to produce anxiety in humans and anxiogenic-like behavior in animals. The goal of this study was three-fold: (1) to determine the doses of cocaine that induce anxiogenic-like behavior in the elevated plus-maze in rats, (2) to determine if cocaine-associated contextual cues are capable of eliciting anxiogenic-like behavior in the absence of the drug, and (3) to identify possible mechanisms through which cocaine-associated cues affect behavior in the elevated plus-maze. Measurement of the amount of time that the animals spend exploring the open arms of the maze provides a sensitive index of anxiogenic-like behavior in rats. In experiment 1, rats were injected with 10 mg/kg, 20 mg/kg, or 30 mg/kg cocaine HCl or saline for 6 days. On day 6, the rats were tested in the elevated plus-maze 25 min after injection with cocaine or saline. The animals chronically treated with the three doses of cocaine exhibited a dose-dependent increase in anxiogenic-like behavior in the elevated plus-maze, compared to the saline-treated group. In experiment 2, cocaine-induced (30 mg/kg) conditioning was achieved using a simple contextual design. On the final day of the experiment (day 6), after 5 days of conditioning, the rats were exposed for 25 min to the cocaine-associated contextual cues, then placed in the elevated plus-maze. Animals that had been exposed to cocaine-associated contextual cues prior to being placed in the elevated plus-maze exhibited a significant increase in anxiogenic-like behavior compared to the control groups. However, pretreatment of the rats with the CRF antagonist, alpha-helical CRF9-41 (1 microg, i.c.v.), on the test day, prior to exposure to cocaine-associated contextual cues, attenuated the subsequent anxiogenic-like behavioral response in the elevated plus-maze (experiment 3). The results suggest that contextual cues associated with repeated treatment with 30 mg/kg cocaine are capable of eliciting anxiogenic-like behavior in the absence of the drug and that CRF mediates the expression of anxiogenic-like behaviors in the elevated plus-maze following exposure to cocaine-associated cues. The conditioned anxiogenic action elicited by cocaine-associated cues may have relevance for understanding the complex addictive nature of this drug and some of the clinical phenomena related to its use.

Conditioned release of corticosterone by contextual stimuli associated with cocaine is mediated by corticotropin-releasing factor.

Elevated blood concentrations of corticosterone (CORT), an adrenal steroid associated with stress responses, is one of the endocrine correlates of cocaine treatment. Experiment 1 confirmed and extended previous findings that chronic cocaine treatment does not alter corticosteroid responses to cocaine. In Experiment 2, conditioned endocrine effects of cocaine were examined in three groups of rats after 7 consecutive days of treatment. Cocaine-induced conditioning was achieved using a simple contextual design. In group 1 (paired), rats were injected with cocaine (30 mg/kg), then immediately placed into a locomotor activity chamber for 30 min. One hour after the rats were returned to their home cages, they received an injection of saline. In group 2 (unpaired), rats were injected with saline, then immediately placed into a locomotor activity chamber for 30 min. One hour after the rats were returned to their home cages, they received an injection of cocaine (30 mg/kg). Rats in group 3 (control) received only saline injections, but otherwise were treated as animals in the other treatment groups. On the test day (Day 8), all rats were placed immediately into the locomotor apparatus for 30 min prior to collection of a blood sample. Blood CORT concentrations and locomotor activity in the paired group were significantly higher than in the unpaired and control groups. However, pretreatment of the rats in Experiment 3 with the corticotropin-releasing factor (CRF) antagonist, alpha-helical CRF9-41 (1 microg, i.c.v.), on the test day, prior to exposure to cocaine-associated contextual cues, attenuated the subsequent conditioned increase in blood CORT concentrations. These data represent the first demonstration of classical conditioning of a steroid hormone response to stimuli associated with a psychoactive drug in rats and suggest that the effect is mediated by endogenous CRF. Because the hypothalamic-pituitary-adrenal (HPA) axis has been implicated in modulating the actions of cocaine, it is plausible that such conditioned increases in CORT release by cocaine-associated cues may further predispose an organism to the reinforcing effects of the drug or enhance the susceptibility to drug-taking behavior. Alternatively, such conditioned effects may be related to the anxiogenic properties of cocaine. Further understanding of the conditioned effects of hormones in the development and expression of addictive behaviors may provide new insights into treatment of drug addiction.

The effects of social isolation on the forced swim test in Fawn hooded and Wistar rats.

Although the forced swim test (FST) has long been used as a preclinical screen of antidepressant efficacy, locomotor stimulants are known to produce confounding effects using the traditional dependent measure in this test: immobility. It has recently been suggested that measurement of struggling behavior may be a better index of antidepressant activity. The present experiments examined behavior in the forced swim test in two potential animal models of depression: the Fawn hooded rat, and the isolation-reared rat. No evidence was found to support these assertions, indeed immobility was decreased in Fawn hooded compared to Wistar rats, however this appeared to be caused by increased struggling behavior in Fawn hooded socials and increased swimming in Fawn hooded isolates. Although these differential results are highly suggestive of different underlying causes of decreased immobility in Fawn hooded rats depending on rearing conditions, the data suggests that the underlying psychological functions assumed to be represented by behavior assessed in this paradigm may not be adequately discriminated.

Different neural mechanisms underlie dizocilpine maleate- and dopamine agonist-induced locomotor activity.

This study evaluated and compared the role of mesoaccumbens dopamine and the ventral pallidal region in the locomotor stimulatory action of the non-competitive N-methyl-D-aspartate antagonist dizocilpine maleate and dopamine agonists. Intra-accumbens injections of both amphetamine (1, 5 and 25 nmol) and dizocilpine maleate (1, 5, 25 and 50 nmol) induced a dose-dependent increase in locomotor activity. The N-methyl-D-aspartate antagonist was somewhat less effective than amphetamine. 6-Hydroxydopamine dopamine-depleting lesions of the nucleus accumbens completely blocked locomotor stimulation induced by focal administrations of amphetamine (5 nmol), but were ineffective in altering the actions of dizocilpine maleate (50 nmol). Ibotenic acid lesions of the ventral pallidal region and muscimol injections into this area also prevented the stimulatory effects of systemic amphetamine (1 mg/kg), while having no effect on the locomotor-activating actions of systemic dizocilpine maleate (0.3 mg/kg). Microdialysis studies revealed that systemically administered apomorphine (2 mg/kg) significantly decreased extracellular GABA in the pallidum, which was accompanied by substantial increases in locomotor output. Systemically administered dizocilpine maleate (0.3 mg/kg), on the other hand, also increased locomotor activity without having any effect on pallidal GABA. These data, taken together, indicate that while the locomotor effects of dopamine agonists are dependent upon intact mesoaccumbens dopamine and involve GABAergic efferents from the nucleus accumbens to the ventral pallidum, dizocilpine maleate's stimulatory actions are independent of such mechanisms.

The role of striatal dopaminergic mechanisms in rotational behavior induced by phencyclidine and phencyclidine-like drugs.

Phencyclidine (PCP) and phencyclidine-like drugs (TCP, dexoxadrol, MK-801, and SKF 10,047) were evaluated for their ability to induce rotational behavior in rats with unilateral 6-OHDA lesions of the medial forebrain bundle and for their ability to alter striatal dopamine (DA) overflow with microdialysis procedures. All of the compounds tested produced rotational behavior ipsilateral to the lesion, suggesting that they were enhancing extracellular dopamine in the intact striatum. The microdialysis studies, however, did not support this contention. There appeared to be a complete dissociation between the ability of the five compounds to produce ipsilateral rotations and their ability to enhance extracellular dopamine levels in the striatum. PCP was the only compound able to elicit significant increases in striatal dopamine overflow following i.p. injections and also produce dramatic rotational behavior. MK-801 was the most potent compound in enhancing rotational output while it had no effect at all on striatal dopamine overflow. Dexoxadrol also produced significant rotational output without having any effect on extracellular levels of dopamine following i.p. injections. TCP and SKF 10,047, at doses which produced significant rotational behavior, only elevated dopamine 16% and 12%, respectively, at peak effect. It is most parsimonious to conclude that the effects of PCP-like drugs on nigro-striatal function are mediated through their ability to act as indirect NMDA receptor antagonists and not through their ability to alter striatal dopamine activity.

Alterations in striatal dopamine overflow during rotational behavior induced by amphetamine, phencyclidine, and MK-801.

Rats lesioned unilaterally in the medial forebrain bundle with 6-OHDA rotated ipsilateral to the lesion following injections of amphetamine, phencyclidine (PCP), and MK-801. Concurrent measurement of striatal dopamine (DA) in the intact striatum with in vivo microdialysis revealed a dissociation between rotational behavior and alterations in DA overflow induced by the three drugs. Amphetamine produced robust ipsilateral rotational behavior and a substantial elevation in striatal DA (approximately 130% increase at asymptote). PCP produced comparable increases in rotational behavior, but only approximately 30% increase in striatal DA. MK-801 also had a comparable behavioral effect but failed to alter DA overflow in the intact striatum. Since MK-801, a noncompetitive NMDA antagonist which does not enhance extracellular dopamine in the striatum, is able to produce ipsilateral rotational behavior in rats with unilateral nigrostriatal lesions, it is likely that the effects of PCP may also be determined predominantly through NMDA blockade in this model.

Heteroaromatic analogs of 1-[2-(diphenylmethoxy)ethyl]- and 1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazines (GBR 12935 and GBR 12909) as high-affinity dopamine reuptake inhibitors.

A new series of heteroaromatic GBR 12935 [1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)-piperazine] (I) and GBR 12909 [1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazine] (2) analogs was synthesized and evaluated as dopamine transporter (DAT) ligands. Analogs 5-16, in which the benzene ring in the phenylpropyl side chain of the GBR molecule had been replaced with a thiophene, furan, or pyridine ring, exhibited high affinity and selectivity for the DAT vs serotonin transporter (SERT) and stimulated locomotor activity in rats in a manner similar to the parent compound 2. In cocaine and food self-administration studies in rhesus monkeys, both thiophene-containing (6 and 8) and pyridine-containing (14 and 16) derivatives displayed potency comparable to 2 in decreasing the cocaine-maintained responding at the doses tested (0.8, 1.7, and 3 mg/kg). However, these compounds did not produce the degree of separation between food- and cocaine-maintained responding that was seen with 2. Among the bicyclic fused-ring congeners 17-38, the indole-containing analog of 2, 22, showed the greatest affinity for binding to the DAT, with IC50 = 0.7 nM, whereas the corresponding indole-containing derivative of 1, 21, displayed the highest selectivity (over 600-fold) at this site vs the SERT site.

Role of central dopaminergic and 5-hydroxytryptaminergic projections in the behavioral responses elicited by thyrotropin-releasing hormone in rats.

The systemic administration of thyrotropin-releasing hormone (TRH) to rats elicits locomotor activation, wet dog shakes, jaw movements, paw licking and tail rattle. Central dopamine (DA) and 5-hydroxytryptamine (5-HT) systems and peripheral vagal afferents have been implicated in these responses. To define this circuitry further, the effects of lesions of these pathways on the behavioral responses elicited by intraperitoneal (IP) injections of TRH were assessed in rats. Lesions of the DAergic innervation of the nucleus accumbens did not affect the locomotor activation, wet dog shakes, paw licking, jaw movements or tail rattle elicited by TRH. This is consistent with our in vivo microdialysis finding that TRH did not affect the release of DA in the nucleus accumbens at a dose that strongly increased locomotor activity. Depletion of spinal 5-HT significantly decreased the wet dog shakes induced by TRH, while depletion of forebrain 5-HT had no effect on any behavior. Bilateral vagotomy did not affect the locomotor response to TRH or any of the other behaviors measured. Taken together these results suggest that the DAergic mesolimbic, the 5-HTergic projections to the forebrain and vagal afferent systems are not mediators of the behavioral responses to systemic TRH. In contrast, the raphe-spinal 5-HTergic projection system may serve to modulate the wet dog shakes elicited by this peptide.

Development of novel, potent, and selective dopamine reuptake inhibitors through alteration of the piperazine ring of 1-[2-(diphenylmethoxy)ethyl]-and 1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazines (GBR 12935 and GBR 12909).

The design, synthesis, and biological evaluation of compounds related to the dopamine (DA) uptake inhibitors: 1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)piperazine (1) and 1-[2-[bis-(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazine (2) (GBR 12395 and GBR 12909, respectively), directed toward the development and identification of new ligands interacting with high potency and selectivity at the dopamine transporter (DAT) is reported. The substitution of the piperazine ring in the GBR structure with other diamine moieties resulted in the retention of the high affinity of new ligands for the DAT. Some of the modified GBR analogs (e.g. 8, 10, (-)-49, or (-)-50) displayed substantially higher selectivity (4736- to 693-fold) for the dopamine (DA) versus the serotonin (5HT) reuptake site than the parent compounds. The bis(p-fluoro) substitution in the (diphenylmethoxy)ethyl fragment slightly increased the affinity of the ligands at the DA reuptake site but reduced their selectivity at this site (e.g. 9 and 8, 11 and 10, or 17 and 16, respectively). Congeners, such as the series of monosubstituted and symmetrically disubstituted piperazines and trans-2,5-dimethylpiperazines, which lack the (diphenylmethoxy)ethyl substituent lost the affinity for the DAT yet exhibited very high potency for binding to the sigma receptors (e.g.28). The chiral pyrrolidine derivatives of 1, (-)-49, and (+)-49, exhibited an enantioselectivity ratio of 181 and 146 for the inhibition of DA reuptake and binding to the DAT, respectively.