Oral Health in the Population Assessment of Tobacco and Health Study.

Tobacco use is a well-established risk factor for multiple adverse oral conditions. Few nationally representative oral health data sets encompass the current diversity of tobacco and nicotine products. This investigation examines the validity of oral health measures in the Population Assessment of Tobacco and Health (PATH) Study to assess relationships between tobacco use and oral health. Cross-sectional data from PATH Study wave 4 (N = 33,643 US adults, collected 2016-2018) were used to obtain estimates for 6 self-reported oral conditions (e.g., bone loss around teeth, tooth extractions) and compared with analogous estimates from the National Health and Nutrition Examination Survey (NHANES) cycle 2017-2018 (N = 5,856). Within the PATH Study, associations were calculated between tobacco use status and lifetime and past 12-mo experience of adverse oral conditions using survey-weighted multivariable logistic regression. Nationally representative estimates of oral conditions between the PATH Study and NHANES were similar (e.g., ever-experience of bone loss around teeth: PATH Study 15.2%, 95% CI, 14.4%-15.9%; NHANES 16.6%, 95% CI, 14.9%-18.4%). In the PATH Study, combustible tobacco smoking was consistently associated with lifetime and past 12-mo experience of adverse oral health (e.g., exclusive cigarette smoking vs. never tobacco use, adjusted odds ratio [AOR] for loose teeth in past 12 mo: 2.02; 95% CI, 1.52-2.69). Exclusive smokeless tobacco use was associated with greater odds of loose teeth (AOR, 1.93; 95% CI, 1.15-3.26) and lifetime precancerous lesions (AOR, 3.85; 95% CI, 1.73-8.57). Use of other noncigarette products (e.g., pipes) was inconsistently associated with oral health outcomes. PATH Study oral health measures closely align with self-reported measures from NHANES and are internally concurrent. Observed associations with tobacco use and the ability to examine emerging tobacco products support application of PATH Study data in dental research, particularly to examine potential oral health effects of novel tobacco products and longitudinal changes in tobacco use behaviors.

The neuropharmacology of prolactin secretion elicited by 3,4-methylenedioxymethamphetamine ("ecstasy"): a concurrent microdialysis and plasma analysis study.

3,4-methylenedioxymethamphetamine (MDMA) is a substituted phenethylamine that is widely abused as the street drug "ecstasy". Racemic MDMA (S,R(+/-)-MDMA) and its stereoisomers elicit complex spectrums of psychobiological, neurochemical, and hormonal effects. In this regard, recent findings demonstrated that S,R(+/-)-MDMA and its stereoisomer R(-)-MDMA elicit increases in striatal extracellular serotonin levels and plasma levels of the hormone prolactin in rhesus monkeys. In the present mechanistic study, we evaluated the role of the serotonin transporter and the 5-HT(2A) receptor in S,R(+/-)-MDMA- and R(-)-MDMA-elicited prolactin secretion in rhesus monkeys through concurrent microdialysis and plasma analysis determinations and drug interaction experiments. Concurrent neurochemical and hormone determinations showed a strong positive temporal correlation between serotonin release and prolactin secretion. Consistent with their distinct mechanisms of action and previous studies showing that the serotonin transporter inhibitor fluoxetine attenuates the behavioral and neurochemical effects of S,R(+/-)-MDMA, pretreatment with fluoxetine attenuated serotonin release elicited by either S,R(+/-)-MDMA or R(-)-MDMA. As hypothesized, at a dose that had no significant effects on circulating prolactin levels when administered alone, fluoxetine also attenuated prolactin secretion elicited by S,R(+/-)-MDMA. In contrast, combined pretreatment with both fluoxetine and the selective 5-HT(2A) receptor antagonist M100907 was required to attenuate prolactin secretion elicited by R(-)-MDMA, suggesting that this stereoisomer of S,R(+/-)-MDMA elicits prolactin secretion through both serotonin release and direct agonism of 5-HT(2A) receptors. Accordingly, these findings inform our understanding of the neuropharmacology of both S,R(+/-)-MDMA and R(-)-MDMA and the regulation of prolactin secretion.

Effects of the monoamine uptake inhibitors RTI-112 and RTI-113 on cocaine- and food-maintained responding in rhesus monkeys.

Cocaine blocks uptake of the monoamines dopamine, serotonin and norepinephrine, and monoamine uptake inhibitors constitute one class of drugs under consideration as candidate "agonist" medications for the treatment of cocaine abuse and dependence. The pharmacological selectivity of monoamine uptake inhibitors to block uptake of dopamine, serotonin and norepinephrine is one factor that may influence the efficacy and/or safety of these compounds as drug abuse treatment medications. To address this issue, the present study compared the effects of 7-day treatment with a non-selective monoamine uptake inhibitor (RTI-112) and a dopamine-selective uptake inhibitor (RTI-113) on cocaine- and food-maintained responding in rhesus monkeys. Monkeys (N=3) were trained to respond for cocaine injections (0.01 mg/kg/inj) and food pellets under a second-order schedule [FR2(VR16:S)] during alternating daily components of cocaine and food availability. Both RTI-112 (0.0032-0.01 mg/kg/hr) and RTI-113 (0.01-0.056 mg/kg/h) produced dose-dependent, sustained and nearly complete elimination of cocaine self-administration. However, for both drugs, the potency to reduce cocaine self-administration was similar to the potency to reduce food-maintained responding. These findings do not support the hypothesis that pharmacological selectivity to block dopamine uptake is associated with behavioral selectivity to decrease cocaine- vs. food-maintained responding in rhesus monkeys.

CART peptides are modulators of mesolimbic dopamine and psychostimulants.

CART peptide produces behavioral effects when injected into the VTA or nucleus accumbens. In the VTA, the peptide behaves like an endogenous psychostimulant and produces increased locomotor activity and conditioned place preference. Since this is blocked by dopamine receptor blockers, it presumably involves release of dopamine. But in the nucleus accumbens, CART peptide reduces the locomotor-increasing effects of cocaine. This suggests that the peptide is an interesting target for medications development.

Activity of various CART peptides in changing locomotor activity in the rat.

Several CART (cocaine- and amphetamine-regulated transcript) peptides have been identified in the brain. One peptide, rlCART 55-102, has been previously characterized in locomotor and feeding assays in rodents. The present study characterized the locomotor-stimulating effect of several additional CART peptides after intra-VTA administration in rats. The results confirm earlier findings that intra-VTA administration of rlCART 55-102 dose-dependently increased locomotor activity. Intra-VTA administration of rlCART 62-102 increased activity comparable with that produced by rlCART 55-102. However, intra-VTA administration of rsCART 10-89 and rlCART 55-59 did not increase motor activity. These results suggest that the portion of rlCART 55-102 responsible for increasing motor activity must reside within the portion of the protein also encompassed by rlCART 61-102, as the initial 5-amino acid sequence of rlCART 55-102 did not increase motor activity. In summary, rlCART 55-102 and rlCART 62-102 are behaviorally active in this locomotor assay, while rsCART 10-89 and rlCART 55-59 are not.

Locomotor stimulant effects of novel phenyltropanes in the mouse.

With the hypothesis that 3-phenyltropane analogs of cocaine might be useful as cocaine medications, 17 analogs (RTI-51, RTI-55, RTI-108, RTI-112, RTI-113, RTI-116, RTI-120, RTI-121, RTI-126, RTI-139, RTI-141, RTI-150, RTI-171, RTI-177, RTI-199, RTI-204, and RTI-219) were characterized for their potency and selectivity at the monoamine transporters in a previous study. Based on their affinities to the transporters in this earlier study, the analogs were classified as nonselective (cocaine, RTI-51, RTI-55, RTI-108, RTI-112, RTI-116, RTI-126, and RTI-139) or dopamine transporter (DAT) selective (RTI-113, RTI-120, RTI-121, RTI-141, RTI-150, RTI-171, RTI-177, RTI-199, RTI-204, and RTI-219). In the present study, the locomotor stimulating effects of these analogs were compared to those of cocaine to obtain a measure of in vivo activity. Each analog was more potent than cocaine in the in vivo assay, as observed in the earlier in vitro studies. Most of these compounds were as efficacious as cocaine, but RTI-51, RTI-108, RTI-113, RTI-121, RTI-139, RTI-141, RTI-177, RTI-204, and RTI-219 were longer acting. Although no correlation between chemical structure and transporter selectivity was found, the short-acting DAT-selective analogs, RTI-120, RTI-150, RTI-171, and RTI-199, all contained a methyl group in the X position of the WIN 35,065-2 molecule. The positive correlation of the IC(50)s for the DAT to potencies for increasing locomotor activity suggested that binding to DAT was responsible for some, if not most, of the locomotor effects of these compounds. Several compounds, including RTI-113 and RTI-177, exhibited properties ideal for medications for cocaine abusers, such as an equivalent efficacy, a higher potency, and a longer duration of action as compared to cocaine.

Repeated cocaine administration does not alter morphine-induced rotational behavior in nigrally denervated rats.

Repeated administration of morphine to rats increases their sensitivity to behavioral effects of morphine as well as to those of psychomotor stimulants, such as cocaine and amphetamine. Conversely, stimulant-induced sensitization to behavioral effects of stimulants often results also in sensitization to behavioral effects of morphine. However, in nigrally lesioned rats, repeated injections of morphine produce sensitization to morphine-induced turning but not to turning induced by cocaine or amphetamine. The present study was performed to determine whether giving repeated cocaine injections to nigrally lesioned rats would produce cross-sensitization to morphine-induced turning. Daily injections of 10 mg/kg cocaine (i.p.) enhanced the turning response to cocaine by day 8, but not the turning response to 3.0 mg/kg morphine (s.c.). The response to morphine increased equally in both cocaine- and saline-treated animals after they had received morphine once. Dose-response curves for morphine (1.0-10 mg/kg) and for cocaine (3.0-30 mg/kg), determined during weeks 3 and 4, were the same in rats receiving daily injections of cocaine or daily injections of saline. Thus, although repeated exposure to cocaine or morphine resulted in sensitization to turning induced by each drug, respectively, there was no cross-sensitization between the two drugs. In contrast to other behaviors, rotational behavior does not seem to exhibit cross-sensitization between morphine and psychomotor stimulants.

Dopamine D1 and D2 receptors influence dopamine transporter synthesis and degradation in the rat.

Neurotransmitter transporters play an important role in maintaining synaptic homeostasis and in the actions of many drugs. Utilizing a technique for measuring the kinetics (synthesis, degradation, and half-life) of the dopamine transporter (DAT) protein in the rat striatum and nucleus accumbens, we have investigated the effects of systemic administration of dopamine receptor agonists and antagonists upon DAT kinetics in these brain regions. In the striatum, the dopamine D1 receptor agonist SKF38393 and the dopamine D1 receptor antagonist SCH23390 were without effect. However, the dopamine D2 receptor agonists R-(-)-propylnorapomorphine hydrochloride (NPA) and quinpirole decreased the half-life of DAT. This effect was blocked by the dopamine D2 antagonist eticlopride, which, by itself, increased the half-life of DAT. In the nucleus accumbens, the agonist SKF38393 increased the DAT half-life, whereas the antagonist SCH23390 decreased the half-life. In contrast to the striatum, NPA and quinpirole increased the DAT half-life, which was blocked by eticlopride and by itself had no effect on DAT kinetics. Cocaine increased the half-life of DAT in both the striatum and the nucleus accumbens. The results of the present study suggest that, through dopamine receptors, dopamine indirectly influences DAT protein turnover in the striatum and in the nucleus accumbens, but in different ways.

RTI-76, an irreversible inhibitor of dopamine transporter binding, increases locomotor activity in the rat at high doses.

An earlier study in our laboratory showed that 24 h after intracerebroventricular administration of the irreversible dopamine transporter inhibitor RTI-76, [3H]GBR12935 binding to the dopamine transporter (DAT) protein was inhibited in both the striatum and nucleus accumbens of the rat in a dose-dependent fashion (0.05-5.0 micromol). The rate of return of binding to control levels was used to calculate the half-life of DAT. Since changes in behavior could conceivably influence the half-life, the effects of various doses of RTI-76 on locomotor activity 1 and 3 days after RTI-76 administration were examined. During the first day after i.c.v. administration, 1.25 micromol RTI-76 had no effect on locomotor activity, but 2.5 micromol RTI-76 significantly increased locomotor activity in rats, a time at which this dose inhibited 41 and 42% of [3H]GBR12935 binding in the striatum and in the nucleus accumbens, respectively. These results agree with earlier reports showing that significant blockade of the dopamine transporter protein in the striatum is required for increases in motor activity in rodents. However, 5.0 micromol RTI-76 did not increase locomotor activity, even though binding was inhibited to 38 and 37% of control levels in the striatum and nucleus accumbens, respectively. Furthermore, our present results suggest that locomotor activity does not continue to increase as the blockade of DAT increases. Notably, there were no increases in locomotor activity at the dose of RTI-76 (100 nmol) used to measure DAT half-life.

Intra-ventral tegmental area injection of rat cocaine and amphetamine-regulated transcript peptide 55-102 induces locomotor activity and promotes conditioned place preference.

Cocaine- and amphetamine-regulated transcript (CART) is a novel mRNA that has been reported to be increased by acute psychostimulant administration, and that may be involved in the effects of psychostimulants. In this study, we examined the effect of centrally administered CART peptides on locomotor activity and conditioned place preference in the rat. CART peptide fragments were bilaterally injected into the ventral tegmental area. CART 55-102 (0.2-5.0 microg/side), an endogenously occurring peptide, dose dependently increased locomotor activity, whereas CART 1-26 (0.1-2.5 microg/side; not found endogenously) did not. The locomotor effects of CART 55-102 were dose dependently blocked by the dopamine D(2) receptor antagonist haloperidol (0.03-1.0 mg/kg i.p.). Four injections of 1.0 microg/side CART 55-102 induced a significant place preference, suggesting that CART 55-102 is reinforcing. Increases in locomotor activity after each of these CART 55-102 injections were similar and did not show tolerance or sensitization. This treatment regimen of CART 55-102 also did not produce sensitization to locomotor activity after a subsequent challenge with cocaine or amphetamine. When CART 55-102 (0.2-1.0 microg/side) was injected into the substantia nigra, no significant change in motor activity was observed. However, a higher dose of CART 55-102 (5.0 microg/side) induced a delayed increase in motor activity, suggesting a possible diffusion from the substantia nigra into the ventral tegmental area. Our findings suggest that CART 55-102 is behaviorally active and may be involved in the actions of psychostimulants. This is the first demonstration of the psychostimulant-like effects of CART peptides.

Dopamine transporter synthesis and degradation rate in rat striatum and nucleus accumbens using RTI-76.

Intracerebroventricular injections of the irreversible dopamine transporter (DAT) inhibitor, RTI-76 [3beta-(3-p-chlorophenyl) tropan-2beta-carboxylic acid p-isothiocyanatophenylethyl ester hydrochloride], decreased DAT binding in both the striatum and nucleus accumbens as measured by both [3H]GBR12935 and by [3H]WIN35,428. This decrease was dose-related, with 100 nmol RTI-76 producing approximately a 50% decrease in both regions. The maximal inhibition of DAT binding was observed 24 h after RTI-76 injection, and binding was fully restored 7 days after injection. The DAT protein half-life determined under these conditions was about 2 days. [3H]Nisoxetine binding at norepinephrine transporters in the cortex was not altered by RTI-76 administration at any time point or dose examined.

Sensitization to daily morphine injections in rats with unilateral lesions of the substantia nigra.

Morphine indirectly enhances dopaminergic activity in the nigrostriatal system, and repeated administration of morphine progressively increases the locomotor activity of rats. We used the rotational behavior model to determine if daily morphine produces an increase in turning and produces cross-sensitization to d-amphetamine and cocaine. Rats with unilateral nigrostriatal lesions received daily injections of saline or morphine (10 mg/kg). Repeated morphine administration produced a progressive increase in turning over 13 days. Next, a morphine dose-response curve (1.0-30 mg/kg) was determined. Both the saline and morphine-treated groups showed dose-dependent increases in turning, but, the peak effect in the morphine group was higher than that in the saline group, indicating sensitization to morphine. The morphine-treated group did not show cross-sensitization to either d-amphetamine (0.1-3 mg/kg) or cocaine (1.0-30 mg/kg); in fact, it showed less cocaine-induced turning than the saline group. Seventy-one days after saline or morphine injections began, the morphine group was still significantly more sensitive to turning induced by 10 mg/kg morphine than the saline group was (200 vs. 750). Therefore, repeated daily injections of morphine produce a progressive sensitization to turning induced by morphine in the absence of cross-sensitization to turning induced by psychomotor stimulants.

Dissociation of morphine-induced potentiation of turning and striatal dopamine release by amphetamine in the nigrally-lesioned rat.

Morphine has been reported to increase extracellular levels of dopamine in the brain of intact rats and to potentiate turning induced by amphetamine in nigrally-lesioned rats. The present study tested the hypothesis that there is a causal relationship between these two effects of morphine. We tested morphine alone, amphetamine alone, and the combination in separate groups of nigrally-lesioned rats for effects on turning and, by microdialysis, on extracellular dopamine levels. Morphine (3.0 or 10 mg/kg) did not produce significant turning but amphetamine (1.0 mg/kg) did. The lower dose, but not the higher dose, of morphine potentiated amphetamine-induced turning. Amphetamine, but not morphine, produced increases in extracellular dopamine levels. In contrast to what occurred with turning, 10 mg/kg but not 3.0 mg/kg morphine potentiated amphetamine-induced increases in extracellular dopamine levels. These results show that the potentiation of amphetamine-induced turning by morphine in nigrally-lesioned rats is not due to the potentiation of dopamine release in the intact striatum.

Opioid receptor agonists and antagonists alter GBR12909-induced turning in the rat.

We extended observations on cocaine-induced turning and its interactions with mu-opioid receptor agonists in nigrally-lesioned rats to GBR12909 (1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-[3-phenyl-propyl]-piperazine) , a selective dopamine reuptake inhibitor. GBR12909 produced turning that was potentiated by the mu-opioid receptor agonists morphine and methadone. The effects of these opioids were blocked by the general opioid receptor antagonist naloxone, which did not affect the action of GBR12909. The reuptake inhibitors nisoxetine (norepinephrine) and fluoxetine (serotonin) did not produce turning alone or in combination with morphine. Antagonists selective for each opioid receptor subtype did not alter GBR12909-induced turning. However, naltrexone, another general opioid receptor antagonist, potentiated turning induced by GBR12909. This was blocked by naloxone, suggesting that naltrexone has opioid receptor agonist actions, in contrast to naloxone. These results indicate that cocaine-induced turning and its potentiation by mu-opioid receptor agonists are dependent upon the inhibition of dopamine reuptake.

Naloxone does not alter amphetamine-induced rotational behavior or striatal dopamine levels of nigrally-lesioned rats.

Previous studies on rats have shown that the opioid antagonist naloxone attenuates amphetamine-induced stimulation of locomotor activity and increases in extracellular dopamine in the brain. However, in this study, naloxone did not attenuate amphetamine-induced rotational behavior or increases of extracellular dopamine in the intact striatum of nigrally-lesioned rats. These results suggests differences in the way in which endogenous opioids contribute to the behavioral and neurochemical effects of amphetamine in nigrally-lesioned compared to intact rats.

Mu opioid agonists potentiate amphetamine- and cocaine-induced rotational behavior in the rat.

Opioids modulate brain dopaminergic function in various experimental paradigms. This study used the rotational model of behavior in rats with unilateral 6-hydroxydopamine-induced lesions of the nigrostriatal pathway to investigate this interaction. Doses of two presynaptically acting dopaminergic drugs, amphetamine and cocaine, were coadministered with several doses of the mu opioid agonist, morphine. Morphine, at 3.0 mg/kg, potentiated rotational behavior induced by each dose of the stimulants. To determine the receptor specificity of the actions of morphine, the mu opioid agonists buprenorphine, fentanyl, levorphanol, meperidine, and methadone, and dextrorphan, the non-opioid isomer of levorphanol, were administered alone and with 1.0 mg/kg amphetamine. Each of these drugs, as well as morphine, produced circling behavior on its own. All of the mu opioid agonists and dextrorphan increased amphetamine-induced turning; the coadministration of dextrorphan, levorphanol, meperidine, methadone and morphine with amphetamine produced turning greater than predicted by simple additivity. To determine whether an opioid receptor was involved in these interactions, the opioid antagonist, naltrexone, was administered before the amphetamine/mu opioid receptor agonist combination. Naltrexone blocked the potentiating effects of morphine, but not those of the other drugs. Moreover, naltrexone alone dose-dependently increased amphetamine-induced rotational behavior. These studies show that some mu opioid receptor agonists can potentiate stimulant-induced rotational behavior and that blockade of opioid receptors can also produce a potentiation. The role of mu opioid receptors in these effects remains unclear.

Synergism between buprenorphine and cocaine on the rotational behavior of the nigrally-lesioned rat.

Buprenorphine, a partial mu-opioid receptor agonist, has been proposed as a treatment for cocaine abuse. However, studies in animals have produced conflicting results on the nature of the interaction between buprenorphine and cocaine. In some studies, buprenorphine attenuated the effects of cocaine and in others it enhanced them. The purpose of the present study was to evaluate the interaction of buprenorphine and cocaine on the rotational behavior of the nigrally-lesioned rat. Both buprenorphine (0.003-0.1 mg/kg) and cocaine (1.0-30 mg/kg) alone produced dose-dependent increases in rotational behavior. Buprenorphine produced long-lasting turning with a peak at 60 min after administration, while cocaine produced turning that peaked immediately after administration and lasted for about 2 h. To distinguish simple additivity from other possible outcomes, we determined the relative potency of each drug alone, using a defined level of effect: 150 turns above the saline control in 4 h. This effect was produced by 10.0 mg/kg cocaine alone and by 0.0175 mg/kg buprenorphine alone. Based on these results, fixed ratio combinations were tested and the experimentally derived effects were compared to the theoretically additive values, using an isobolographic analysis. The fixed ratio combinations of the two drugs tested produced turning greater than predicted from simple additivity. This finding provides statistically-supported evidence for synergism between the actions of buprenorphine and cocaine.

Theory and statistics of detecting synergism between two active drugs: cocaine and buprenorphine.

This article discusses the theory and statistical aspects in the design and analysis of experiments to detect synergism between two drugs that produce overtly similar effects. The current analysis extended and simplifies previously published work in this area. Application is made to a study by Kimmel et al. in this issue that examined the combined action of buprenorphine and cocaine in producing turning in rats having unilateral nigrostriatal lesions produced by 6-hydroxydopamine. The use of turning as an endpoint is unusual in quantitative studies of synergism in that no clear maximum effect (turning), could be elicited. Data from the turning study are analyzed statistically and reveal that the combination of buprenorphine and cocaine in each of two fixed ratio mixtures tested is synergistic for this effect.

Effects of acute and chronic morphine on rotational behavior in nigral-lesioned rats.

Stimulation of mu-opioid receptors located on dopaminergic neurons in the striatum and the nucleus accumbens increases dopamine release, which may account for some of the behavioral effects of morphine. In this study, we examined the effects of acute and chronic morphine treatment on rotational behavior in rats with unilateral 6-hydroxydopamine dopamine (6-OHDA)-induced lesions of the nigrostriatal pathway. Rats receiving morphine acutely (0.3-10 mg/kg) did not show a significant bias toward contralateral or ipsilateral turning. Mini osmotic pumps dispensing morphine continuously (20-24 mg/kg/day) were implanted s.c. in these animals. This treatment induced tolerance to the behavioral depression produced by the highest dose of morphine (10 mg/kg) when it was given acutely. A slight but significant increase in ipsilateral turning occurred over the range of morphine doses examined. The effects of morphine on rotational behavior are slight, and do not correlate well with the reported increase in locomotor activity or extraneural dopamine in the striatum that are produced by doses of morphine similar to the ones tested in this study.

Chimeric human and rat serotonin transporters reveal domains involved in recognition of transporter ligands.

The serotonin transporter (SERT) is a target for many clinically significant drugs, such as cocaine, amphetamine, and antidepressants. The relationship between the structure of SERT and the binding of substrates and antagonists is virtually unknown, despite a large body of data describing the structure-activity relationships of transporter ligands. The cloning of multiple species homologs of SERT affords a unique opportunity for molecular comparisons to identify potential domains and residues involved in ligand recognition. We have conducted pharmacological comparisons of the cloned rat and human SERTs in transiently transfected HeLa cells. Serotonin uptake and radioligand binding assays revealed that rat and human SERTs show different sensitivities to some but not all transporter ligands; most tricyclic antidepressants were significantly more potent at the human SERT, relative to rat SERT, whereas d-amphetamine was a more potent inhibitor of rat SERT. Several other ligand such as fluoxetine, paroxetine, (+)-methylenedioxymethamphetamine, cocaine, and the substrate 5-hydroxytryptamine, shows no significant species selectivity. Cross-species chimeras between rat and human SERTs were constructed to track the species-specific pharmacologies through the SERT molecule. These chimeric SERTs were expressed in HeLa cells and transported serotonin similarly to parental SERTs. Using these chimeras, we have isolated a region distal to amino acid 532 the imparts species preferences for both the tricyclic imipramine and d-amphetamine. Our results support the prediction of distinct binding sites for SERT ligands and implicate a restricted region in or near putative transmembrane domain 12 of the transport as being involved in both substrate and antagonist recognition.