Improvement of learning and increase in dopamine level in the frontal cortex by methylphenidate in mice lacking dopamine transporter.

The symptoms of attention-deficit/hyperactivity disorder (ADHD) are characterized by inattention and hyperactivity-impulsivity. It is a common childhood neurodevelopmental disorder that often persists into adulthood. Improvements in ADHD symptoms using psychostimulants have been recognized as a paradoxical calming effect. The psychostimulant methylphenidate (MPH) is currently used as the first-line medication for the management of ADHD. Recent studies have drawn attention to altered dopamine-mediated neurotransmission in ADHD, particularly reuptake by the dopamine transporter (DAT). This hypothesis is supported by the observation that DAT knockout mice exhibit marked hyperactivity that is responsive to acute MPH treatment. However, other behaviors relevant to ADHD have not been fully clarified. In the present study, we observed learning impairment in shuttle-box avoidance behavior together with hyperactivity in a novel environment in DAT knockout mice. Methylphenidate normalized these behaviors and enhanced escape activity in the tail suspension test. Interestingly, the effective dose of MPH increased extracellular dopamine in the prefrontal cortex but not striatum, suggesting an important role for changes in prefrontal dopamine in ADHD. Research that uses rodent models such as DAT knockout mice may be useful for elucidating the pathophysiology of ADHD.

Region-specific dendritic spine loss of pyramidal neurons in dopamine transporter knockout mice.

Dopamine transporter (DAT) knockout (KO) mice show numerous behavioral alterations, including hyperlocomotion, cognitive deficits, impulsivity and impairment of prepulse inhibition of the startle reflex (PPI), phenotypes that may be relevant to frontostriatal disorders such as schizophrenia. Dendritic spine changes of pyramidal neurons in the dorsolateral prefrontal cortex (DLPFC) are among the most replicated of findings in postmortem studies of schizophrenia. The mechanisms that account for dendritic changes in the DLPFC in schizophrenia are unclear. Here, we report basal spine density of pyramidal neurons in the medial prefrontal cortex (mPFC), the motor cortex, the CA1 region of the hippocampus, and the basolateral amygdala in DAT KO mice. Pyramidal neurons were visualized using DAT KO mice crossbred with a Thy1-GFP transgenic mouse line. We observed a significant decrease in spine density of pyramidal neurons in the mPFC and the CA1 region of the hippocampus in DAT KO mice compared to that in WT mice. On the other hand, no difference was observed in spine density of pyramidal neurons in the motor cortex or the basolateral amygdala between DAT genotypes. These results suggest that decreased spine density could cause hypofunction of the mPFC and the hippocampus, and contribute to the behavioral abnormalities observed in DAT KO mice, including cognitive deficits. This might suggest that aberrant dopaminergic signaling may trigger dystrophic changes in dendrites of hippocampal and prefrontocortical pyramidal neurons in schizophrenia.

Decreased vesicular monoamine transporter 2 (VMAT2) and dopamine transporter (DAT) function in knockout mice affects aging of dopaminergic systems.

Dopamine (DA) is accumulated and compartmentalized by the dopamine transporter (DAT; SLC3A6) and the vesicular monoamine transporter 2 (VMAT2; SLC18A2). These transporters work at the plasma and vesicular membranes of dopaminergic neurons, respectively, and thus regulate levels of DA in neuronal compartments that include the extravesicular cytoplasmic compartment. DA in this compartment has been hypothesized to contribute to oxidative damage that can reduce the function of dopaminergic neurons in aging brains and may contribute to reductions in dopaminergic neurochemical markers, locomotor behavior and responses to dopaminergic drugs that are found in aged animals. The studies reported here examined aged mice with heterozygous deletions of VMAT2 or of DAT, which each reduce transporter expression to about 50% of levels found in wild-type (WT) mice. Aged mice displayed reduced locomotor responses under a variety of circumstances, including in response to locomotor stimulants, as well as changes in monoamine levels and metabolites in a regionally dependent manner. Several effects of aging were more pronounced in heterozygous VMAT2 knockout (KO) mice, including aging induced reductions in locomotion and reduced locomotor responses to cocaine. By contrast, some effects of aging were reduced or not observed in heterozygous DAT KO mice. These findings support the idea that altered DAT and VMAT2 expression affect age-related changes in dopaminergic function. These effects are most likely mediated by alterations in DA compartmentalization, and might be hypothesized to be exacerbated by other factors that affect the metabolism of cytosolic DA. This article is part of the Special Issue entitled 'The Synaptic Basis of Neurodegenerative Disorders'.

Smoking quit success genotype score predicts quit success and distinct patterns of developmental involvement with common addictive substances.

Genotype scores that predict relevant clinical outcomes may detect other disease features and help direct prevention efforts. We report data that validate a previously established v1.0 smoking cessation quit success genotype score and describe striking differences in the score in individuals who display differing developmental trajectories of use of common addictive substances. In a cessation study, v1.0 genotype scores predicted ability to quit with P=0.00056 and area under receiver-operating characteristic curve 0.66. About 43% vs 13% quit in the upper vs lower genotype score terciles. Latent class growth analyses of a developmentally assessed sample identified three latent classes based on substance use. Higher v1.0 scores were associated with (a) higher probabilities of participant membership in a latent class that displayed low use of common addictive substances during adolescence (P=0.0004) and (b) lower probabilities of membership in a class that reported escalating use (P=0.001). These results indicate that: (a) we have identified genetic predictors of smoking cessation success, (b) genetic influences on quit success overlap with those that influence the rate at which addictive substance use is taken up during adolescence and (c) individuals at genetic risk for both escalating use of addictive substances and poor abilities to quit may provide especially urgent focus for prevention efforts.

Biomarkers for smoking cessation.

One way to enhance therapeutic development is through the identification and development of evaluative tools such as biomarkers. This review focuses on putative diagnostic, pharmacodynamic, and predictive biomarkers for smoking cessation. These types of biomarkers may be used to more accurately diagnose a disease, personalize treatment, identify novel targets for drug discovery, and enhance the efficiency of drug development. Promising biomarkers are presented across a range of approaches including metabolism, genetics, and neuroimaging. A preclinical viewpoint is also offered, as are analytical considerations and a regulatory perspective summarizing a pathway toward biomarker qualification.

Evaluation of an HIV prevention intervention designed for African American Women: results from the SISTA Community-Based Organization Behavioral Outcomes Project.

One of the Centers for Disease Control and Prevention's strategies for addressing racial disparities within the HIV epidemic is to support the implementation of HIV prevention behavioral interventions designed for African Americans. One such intervention is Sisters Informing Sisters about Topics on AIDS (SISTA), a culturally relevant and gender-specific, five-session, group-level, HIV prevention intervention designed for African American women. In 2008, the Centers for Disease Control and Prevention funded five community-based organizations to conduct outcome monitoring of SISTA to assess the outcomes associated with implementation in the field. Using a 90-day recall, demographic and sexual risk data were collected from participants at baseline and at 90 and 180 days post-intervention. Findings reveal that women participating in SISTA (n = 432) demonstrated a significant reduction in sexual risk between baseline and both follow-up time points for each of the six outcomes being measured (e.g., any unprotected sex, all protected sex).

Dopamine D4 receptor gene variation moderates the efficacy of bupropion for smoking cessation.

Smokers (≥10 cigarettes per day, N=331) of European ancestry taking part in a double-blind placebo-controlled randomized trial of 12 weeks of treatment with bupropion along with counseling for smoking cessation were genotyped for a variable number of tandem repeats polymorphism in exon III of the dopamine D4 receptor gene. Generalized estimating equations predicting point-prevalence abstinence at end of treatment and 2, 6 and 12 months after the end of treatment indicated that bupropion (vs placebo) predicted increased odds of abstinence. The main effect of Genotype was not significant. A Genotype × Treatment interaction (P=0.005) showed that bupropion predicted increased odds of abstinence in long-allele carriers (odds ratios (OR)=1.31, P<0.0001), whereas bupropion was not associated with abstinence among short-allele homozygotes (OR=1.06, P=0.23). The Genotype × Treatment interaction remained when controlling for demographic and clinical covariates (P=0.01) and in analyses predicting continuous abstinence (P's≤0.054). Bupropion may be more efficacious for smokers who carry the long allele, which is relevant to personalized pharmacogenetic treatment approaches.

Effects of MDMA on Extracellular Dopamine and Serotonin Levels in Mice Lacking Dopamine and/or Serotonin Transporters.

3,4-Methylendioxymethamphetamine (MDMA) has both stimulatory and hallucinogenic properties which make its psychoactive effects unique and different from those of typical psychostimulant and hallucinogenic agents. The present study investigated the effects of MDMA on extracellular dopamine (DA(ex)) and serotonin (5-HT(ex)) levels in the striatum and prefrontal cortex (PFC) using in vivo microdialysis techniques in mice lacking DA transporters (DAT) and/or 5-HT transporters (SERT). subcutaneous injection of MDMA (3, 10 mg/kg) significantly increased striatal DA(ex) in wild-type mice, SERT knockout mice, and DAT knockout mice, but not in DAT/SERT double-knockout mice. The MDMA-induced increase in striatal DA(ex) in SERT knockout mice was significantly less than in wildtype mice. In the PFC, MDMA dose-dependently increased DA(ex) levels in wildtype, DAT knockout, SERT knockout and DAT/SERT double-knockout mice to a similar extent. In contrast, MDMA markedly increased 5-HT(ex) in wildtype and DAT knockout mice and slightly increased 5-HT(ex) in SERT-KO and DAT/SERT double-knockout mice. The results confirm that MDMA acts at both DAT and SERT and increases DA(ex) and 5-HT(ex).

Evaluation of an HIV prevention intervention for African Americans and Hispanics: findings from the VOICES/VOCES Community-based Organization Behavioral Outcomes Project.

There is limited knowledge about whether the delivery of evidence-based, HIV prevention interventions in 'real world' settings will produce outcomes similar to efficacy trial outcomes. In this study, we describe longitudinal changes in sexual risk outcomes among African American and Hispanic participants in the Video Opportunities for Innovative Condom Education and Safer Sex (VOICES/VOCES) program at four CDC-funded agencies. VOICES/VOCES was delivered to 922 high-risk individuals in a variety of community settings such as substance abuse treatment centers, housing complex centers, private residences, shelters, clinics, and colleges. Significant risk reductions were consistently observed at 30- and 120-days post-intervention for all outcome measures (e.g., unprotected sex, self-reported STD infection). Risk reductions were strongest for African American participants, although Hispanic participants also reported reducing their risky behaviors. These results suggest that, over a decade after the first diffusion of VOICES/VOCES across the U.S. by CDC, this intervention remains an effective tool for reducing HIV risk behaviors among high-risk African American and Hispanic individuals.

A greater role for the norepinephrine transporter than the serotonin transporter in murine nociception.

Norepinephrine and serotonin involvement in nociceptive functions is supported by observations of analgesic effects of norepinephrine transporter (NET) and serotonin transporter (SERT) inhibitors such as amitriptyline. However, the relative contribution of NET and SERT to baseline nociception, as well as amitriptyline analgesia, is unclear. Amitriptyline and morphine analgesia in wild-type (WT) mice and littermates with gene knockout (KO) of SERT, NET or both transporters was conducted using the hotplate and tail-flick tests. Hypoalgesia was observed in NET KO mice, and to a lesser extent in SERT KO mice. The magnitude of this hypoalgesia in NET KO mice was so profound that it limited the assessment of drug-induced analgesia. Nonetheless, the necessary exclusion of these subjects because of profound baseline hypoalgesia strongly supports the role of norepinephrine and NET in basal nociceptive behavior while indicating a much smaller role for serotonin and SERT. To further clarify the role of NET and SERT in basal nociceptive sensitivity further experiments were conducted in SERT KO and NET KO mice across a range of temperatures. NET KO mice were again found to have pronounced thermal hypoalgesia compared to WT mice in both the hotplate and tail-flick tests, while only limited effects were observed in SERT KO mice. Furthermore, in the acetic acid writhing test of visceral nociception pronounced hypoalgesia was again found in NET KO mice, but no change in SERT KO mice. As some of these effects may have resulted from developmental consequences of NET KO, the effects of the selective NET blocker nisoxetine and the selective SERT blocker fluoxetine were also examined in WT mice: only nisoxetine produced analgesia in these mice. Collectively these data suggest that NET has a far greater role in determining baseline analgesia, and perhaps other analgesic effects, than SERT in mice.

Organic labeling influences food valuation and choice.

Everyday we choose between a variety of different food items trying to reach a decision that fits best our needs. These decisions are highly dependent on the context in which the alternatives are presented (e.g. labeling). We investigate the influence of cognition on food evaluation, using an fMRI experiment in which subjects saw and bid on different foods labeled with (or without) a widely known German emblem for organically produced food. Increased activity in the ventral striatum was found for foods labeled "organic" in comparison to conventionally labeled food. Between-subject differences in activity were related to actual everyday consumption behavior of organic food.

Effect of KEPI (Ppp1r14c) deletion on morphine analgesia and tolerance in mice of different genetic backgrounds: when a knockout is near a relevant quantitative trait locus.

We previously identified KEPI as a morphine-regulated gene using subtractive hybridization and differential display PCR. Upon phosphorylation by protein kinase C, KEPI becomes a powerful inhibitor of protein phosphatase 1. To gain insights into KEPI functions, we created KEPI knockout (KO) mice on mixed 129S6xC57BL/6 genetic backgrounds. KEPI maps onto mouse chromosome 10 close to the locus that contains the mu-opioid receptor (Oprm1) and provides a major quantitative trait locus for morphine effects. Analysis of single nucleotide polymorphisms in and near the Oprm1 locus identified a doubly-recombinant mouse with C57BL/6 markers within 1 Mb on either side of the KEPI deletion. This strategy minimized the amount of 129S6 DNA surrounding the transgene and documented the C57BL/6 origin of the Oprm1 gene in this founder and its offspring. Recombinant KEPIKO mice displayed (a) normal analgesic responses and normal locomotion after initial morphine treatments, (b) accelerated development of tolerance to analgesic effects of morphine, (c) elevated activity of protein phosphatase 1 in thalamus, (d) attenuated morphine reward as assessed by conditioned place preference. These data support roles for KEPI action in adaptive responses to repeated administration of morphine that include analgesic tolerance and drug reward.

Species differences in cannabinoid receptor 2 (CNR2 gene): identification of novel human and rodent CB2 isoforms, differential tissue expression and regulation by cannabinoid receptor ligands.

Cannabinoids, endocannabinoids and marijuana activate two well-characterized cannabinoid receptors (CB-Rs), CB1-Rs and CB2-Rs. The expression of CB1-Rs in the brain and periphery has been well studied, but neuronal CB2-Rs have received much less attention than CB1-Rs. Many studies have now identified and characterized functional glial and neuronal CB2-Rs in the central nervous system. However, many features of CB2-R gene structure, regulation and variation remain poorly characterized in comparison with the CB1-R. In this study, we report on the discovery of a novel human CB2 gene promoter transcribing testis (CB2A) isoform with starting exon located ca 45 kb upstream from the previously identified promoter transcribing the spleen isoform (CB2B). The 5' exons of both CB2 isoforms are untranslated 5'UTRs and alternatively spliced to the major protein coding exon of the CB2 gene. CB2A is expressed higher in testis and brain than CB2B that is expressed higher in other peripheral tissues than CB2A. Species comparison found that the CB2 gene of human, rat and mouse genomes deviated in their gene structures and isoform expression patterns. mCB2A expression was increased significantly in the cerebellum of mice treated with the CB-R mixed agonist, WIN55212-2. These results provide much improved information about CB2 gene structure and its human and rodent variants that should be considered in developing CB2-R-based therapeutic agents.

Cocaine-conditioned locomotion in dopamine transporter, norepinephrine transporter and 5-HT transporter knockout mice.

The behavioral effects of cocaine are affected by gene knockout (KO) of the dopamine transporter (DAT), the serotonin transporter (SERT) and the norepinephrine transporter (NET). The relative involvement of each of these transporters varies depending on the particular behavioral response to cocaine considered, as well as on other factors such as genetic background of the subjects. Interestingly, the effects of these gene knockouts on cocaine-induced locomotion are quite different from those on reward assessed in the conditioned place preference paradigm. To further explore the role of these genes in the rewarding effects of cocaine, the ability of five daily injections of cocaine to induce conditioned locomotion was assessed in DAT, SERT and NET KO mice. Cocaine increased locomotor activity acutely during the initial conditioning session in SERT KO and NET KO, but not DAT KO, mice. Surprisingly, locomotor responses in the cocaine-paired subjects diminished over the five conditioning sessions in SERT KO mice, while locomotor responses increased in DAT KO mice, despite the fact that they did not demonstrate any initial locomotor responses to cocaine. Cocaine-induced locomotion was unchanged over the course of conditioning in NET KO mice. In the post-conditioning assessment, conditioned locomotion was not observed in DAT KO mice, and was reduced in SERT KO and NET KO mice. These data reaffirm the central role of dopamine and DAT in the behavioral effects of cocaine. Furthermore, they emphasize the polygenic basis of cocaine-mediated behavior and the non-unitary nature of drug reward mechanisms, particularly in the context of previous studies that have shown normal cocaine-conditioned place preference in DAT KO mice.

Nicotine abstinence genotyping: assessing the impact on smoking cessation clinical trials.

Twin studies document substantial heritability for successful abstinence from smoking. A genome-wide association study has identified markers whose allele frequencies differ with nominal P<0.005 in nicotine-dependent clinical trial participants who were successful vs unsuccessful in abstaining from smoking; many of these results are also supported by data from two additional samples. More study is required to precisely determine the variance in quitting success that can be accounted for by the single-nucleotide polymorphisms that are currently identified and to precisely classify individuals who may display varying degrees of genetic vs environmental effects into quitters or nonquitters. However, the data at hand do allow us to model the effects of genotypic stratification in smoking cessation trials. We identify relationships between the costs of identifying and genotyping prospective trial participants vs the costs of performing the clinical trials. We quantitate the increasing savings that result from genetically stratified designs as recruiting/genotyping costs go down and trial costs increase. This model helps to define the circumstances in which genetically stratified designs may enhance power and reduce costs for smoking cessation clinical trials.

Sigma1 receptor antagonists determine the behavioral pattern of the methamphetamine-induced stereotypy in mice.

OBJECTIVE: The effects of sigma receptor antagonists on methamphetamine (METH)-induced stereotypy have not been examined. We examined the effects of sigma antagonists on METH-induced stereotypy in mice. RESULTS: The administration of METH (10 mg/kg) to male ddY mice induced stereotyped behavior consisting of biting (90.1%), sniffing (4.2%), head bobbing (4.1%), and circling (1.7%) during an observation period of 1 h. Pretreatment of the mice with BMY 14802 (alpha-(4-fluorophenyl)-4-(5-fluoro-2-pyrimidinyl)-1-piperazinebutanol; 1, 5, and 10 mg/kg), a non-specific sigma receptor antagonist, significantly increased METH-induced sniffing (19.2%, 30.5%, and 43.8% of total stereotypical behavior) but decreased biting (76.6%, 66.9%, and 49.3% of total stereotypical behavior) in a dose-dependent manner. This response was completely abolished by (+)-SKF 10,047 ([2S-(2alpha,6alpha,11R)]-1,2,3,4,5,6-hexahydro-6,11-dimethyl-3-(2-propenyl)-2,6-methano-3-benzazocin-8-ol; 4 and 10 mg/kg), a putative sigma(1) receptor agonist, and partially by PB 28 (1-cyclohexyl-4-[3-(1,2,3,4-tetrahydro-5-methoxy-1-naphthalen-1-yl)-n-propyl]piperazine; 1 and 10 mg/kg), a putative sigma(2) receptor agonist. The BMY 14802 action on METH-induced stereotypy was mimicked by BD 1047 (N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(dimethylamino)ethylamine; 10 mg/kg), a putative sigma(1) receptor antagonist, but not by SM-21 ((+/-)-tropanyl 2-(4-chlorophenoxy)butanoate; 1 mg/kg), a putative sigma(2) receptor antagonist. The BD 1047 effect on METH-induced stereotypy was also abolished completely by (+)-SKF 10,047 and partially by PB 28. The overall frequency of METH-induced stereotypical behavior was unchanged with these sigma receptor ligands, despite the alteration in particular behavioral patterns. The BMY 14802 action on METH-induced stereotypy was unaffected by pretreatment with centrally acting histamine H(1) receptor antagonists (pyrilamine or ketotifen, 10 mg/kg), suggesting that these effects are independent of histamine H(1) receptor signaling systems. CONCLUSION: In summary, modulation of central sigma(1) receptors alters the pattern of METH-induced stereotypy, producing a shift from stereotypical biting to stereotypical sniffing, without affecting the overall frequency of stereotypical behavior.

Behavioral effects of CB2 cannabinoid receptor activation and its influence on food and alcohol consumption.

Consumers of marijuana typically feel a strong, compulsive desire to consume food. Although past research revealed that the CB1 cannabinoid receptor is a potent regulator of food intake, the functional presence of neuronal CB2 cannabinoid receptors in the brain has been controversial. The role of CB2 receptors in food and alcohol consumption and the behavioral effects of CB2 receptor ligands are not well characterized. This is because CB2 cannabinoid receptors were thought to be absent from the brain and expressed primarily in immune cells and in the periphery. We tested the effects of peripheral injections of CB2 antagonist AM 630, CB2 agonist PEA, and CB1 antagonist AM 251 on male C57BL/6, Balb/c, and DBA/2 mice at the beginning of the night cycle and after overnight 12-hour fasts. We also investigated the effects of the putative CB2 agonist, JWH015, and CB2 antagonist, SR144528, in mouse motor function tests and in the two-compartment black and white box. Under standard conditions, the CB2 antagonist AM 630 inhibited food consumption in C57BL/6 mice and DBA/2 mice, but failed to block food intake of Balb/c mice. The CB2 agonist PEA had no significant effect on food consumption in Balb/c mice, and reduced food intake in C57BL/6 and DBA mice. The CB1 antagonist AM 251 inhibited food ingestion in the three mouse strains at variable times. After 12-hour food deprivation, the CB2 antagonist AM 630 increased food consumption in C57Bl/6 mice, but failed to produce significant changes in food intake for Balb/c and DBA/2 mice. The CB2 agonist PEA also reduced food consumption in all three mice strains at variable times. In comparison to the CB2 ligands, CB1 antagonist AM 251 inhibited food ingestion in the mouse strains. A general pattern of depression in locomotor activity was induced by JWH 015 in both males and females in the three mouse strains tested as the dose was increased. The development and enhancement of alcohol preference was observed after chronic treatment with CB2 agonist JWH 015 in stressed mice, but not in controls. In the DBA/2 strain, the spontaneous locomotor activity and stereotype behavior was enhanced by acute administration of low doses of SR144528. There was a reduction in CNR2 gene expression in the ventral mid-brain region of mice that developed alcohol preference, but not in those that did not develop alcohol preference. These effects of CB2 cannabinoid receptor ligands in in vivo behavioral tests are provided as functional evidence that CB2-Rs in the brain play a role in food and alcohol consumption and in the modification of mouse behavior.

Knockout of the mu opioid receptor enhances the survival of adult-generated hippocampal granule cell neurons.

Recent evidence suggests that mu opioid receptors (MOR) are key regulators of hippocampal structure and function. For example, exogenous MOR agonists morphine and heroin negatively impact hippocampal function and decrease adult hippocampal neurogenesis. Here we explored the role of MOR in the birth and survival of hippocampal progenitor cells by examining adult neurogenesis in mice that lack MOR. Adult male mice lacking exon 1 of MOR were injected with the S phase marker bromodeoxyuridine (BrdU) and killed either 2 hours or 4 weeks later to evaluate proliferating and surviving BrdU-immunoreactive (IR) cells, respectively, in the adult hippocampal granule cell layer. Wild-type (WT), heterozygote, and homozygote mice did not differ in the number of BrdU-IR cells at a proliferation time point. However, 4 weeks after BrdU injection, heterozygote and homozygote mice had 57% and 54% more surviving BrdU-IR cells in the hippocampal granule cell layer as compared with WT mice. A decrease in apoptosis in the heterozygote and homozygote mice did not account for the difference in number of surviving BrdU-IR cells since there were no alterations in number of pyknotic, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive, or activated caspase 3-IR cells compared with WT. In concordance with the increased numbers of granule cells maturing into neurons, heterozygote and homozygote mice had larger hippocampal granule cell layers and increased numbers of granule cells. These findings indicate that MOR may play a role in regulating progenitor cell survival and more generally encourage further exploration of how MOR activation can influence hippocampal structure and function.

Structure-activity relationship of trihexyphenidyl analogs with respect to the dopamine transporter in the on going search for a cocaine inhibitor.

A series of trihexyphenidyl (THP) analogs were used to search for a derivative that could serve as a cocaine inhibitor. A compound that blocks binding of the cocaine analog carboxyfluorotropane (CFT), allows dopamine uptake and exhibits low side effects could serve as a good candidate for that purpose. All analogs were tested for the extent to which they inhibit CFT binding, dopamine uptake and n-methyl scopolamine (NMS) binding. Several structure-function relationships emerged. Methylation/halogenation of THP's benzene ring enhanced the compound's ability to block CFT binding in comparison to its ability to block dopamine uptake (5a-e). Replacement of the cyclohexyl ring with a benzene ring tended to create compounds that had lower affinities to the dopamine transporter (7b compared to THP, 7d compared to 5h, 7c compared to 8c) and modification of THP's piperidine ring tended to enhance affinity to the dopamine transporter (5f-h, 8a, 8c). One analog (5f) that showed little muscarinic activity indicating that it would probably have few side effects was investigated for its effects as an in vivo cocaine inhibitor. However, it showed few antagonistic effects in vivo. Nevertheless, this work greatly elucidates the structure-function relationships required for potential cocaine inhibitors and so lays out promising directions for future research.