Dopamine neuronal transport kinetics and effects of amphetamine

The dopamine (DA) transporter (DAT) regulates DA neurotransmission by recycling DA back into neurons. Drugs that interfere with DAT function, e.g., cocaine and amphetamine, can have profound behavioral effects. The kinetics of DA transport by DAT in isolated synaptosomal or single cell preparations have been previously studied. To investigate how DA transport is regulated in intact tissue and to examine how amphetamine affects the DAT, the kinetics of DA uptake by the DAT were examined in tissue slices of the mouse caudate-putamen with fast-scan cyclic voltammetry. The data demonstrate that inward DA transport is saturable and sodium-dependent. Elevated levels of cytoplasmic DA resulting from disruption of vesicular storage by incubation with 10 microM Ro 4-1284 did not generate DA efflux or decrease its uptake rate. However, incubation with 10 microM amphetamine reduced the net DA uptake rate and increased extracellular DA levels due to DA efflux through the DAT. In addition, a new, elevated steady-state level of extracellular DA was established after electrically stimulated DA release in the presence of amphetamine, norepinephrine, and exogenous DA. These results from intact tissue are consistent with a kinetic model of the DAT established in more purified preparations in which amphetamine and other transported substances make the inwardly facing DAT available for outward transport of intracellular DA.     

High doses of amphetamine augment, rather than disrupt, exocytotic dopamine release in the dorsal and ventral striatum of the anesthetized rat

High doses of amphetamine (AMPH) are thought to disrupt normal patterns of action potential-dependent dopaminergic neurotransmission by depleting vesicular stores of dopamine (DA) and inducing robust non-exocytotic DA release or efflux via dopamine transporter (DAT) reversal. However, these cardinal AMPH actions have been difficult to establish definitively in vivo. Here, we use fast-scan cyclic voltammetry (FSCV) in the urethane-anesthetized rat to evaluate the effects of 10 and 20 mg/kg AMPH on vesicular DA release and DAT function in dorsal and ventral striata. An equivalent high dose of cocaine (40 mg/kg) was also examined for comparison to psychostimulants acting preferentially by DAT inhibition. Parameters describing exocytotic DA release and neuronal DA uptake were determined from dynamic DA signals evoked by mild electrical stimulation previously established to be reinforcing. High-sensitivity FSCV with nanomolar detection was used to monitor changes in the background voltammetric signal as an index of DA efflux. Both doses of AMPH and cocaine markedly elevated evoked DA levels over the entire 2-h time course in the dorsal and ventral striatum. These increases were mediated by augmented vesicular DA release and diminished DA uptake typically acting concurrently. AMPH, but not cocaine, induced a slow, DA-like rise in some baseline recordings. However, this effect was highly variable in amplitude and duration, modest, and generally not present at all. These data thus describe a mechanistically similar activation of action potential-dependent dopaminergic neurotransmission by AMPH and cocaine in vivo. Moreover, DA efflux appears to be a unique, but secondary, AMPH action.     

Effects of Glutamate Antagonists on Methamphetamine and 3,4-Methylenedioxymethamphetamine-Induced Striatal Dopamine Release In Vivo

Abstract: Several amphetamine analogues are reported to increase striatal glutamate efflux in vivo, whereas other data indicate that glutamate is capable of stimulating the efflux of dopamine (DA) in the striatum via a glutamate receptor-dependent mechanism. Based on these findings, it has been proposed that the ability of glutamate receptor-blocking drugs to antagonize the effects of amphetamine may be explained by their capacity to inhibit DA release induced by glutamate. To examine this possibility further, we investigated in vivo the ability of glutamate antagonists to inhibit DA release induced by either methamphetamine (METH) or 3,4-methylenedioxymethamphetamine (MDMA). Both METH and MDMA increased DA efflux in the rat striatum and, in animals killed 1 week later, induced persistent depletions of DA and serotonin in tissue. Pretreatment with MK-801 or CGS 19755 blocked the neurotoxic effects of METH and MDMA but, did not significantly alter striatal DA efflux induced by either stimulant. Infusion of 6-cyano-7-nitroquinoxaline-2,3-dione into the striatum likewise did not alter METH-induced DA overflow, and none of the glutamatergic antagonists affected the basal release of DA when given alone. The findings suggest that the neuroprotective effects of NMDA antagonists do not involve an inhibition of DA release, nor do the data support the proposal that glutamate tonically stimulates striatal DA efflux in vivo. Whether phasic increases in glutamate content might stimulate DA release, however, remains to be determined.     

Presynaptic control of striatal dopamine neurotransmission in adult vesicular monoamine transporter 2 (VMAT2) mutant mice

The vesicular monoamine transporter 2 (VMAT2) plays a pivotal role in regulating the size of vesicular and cytosolic dopamine (DA) storage pools within the CNS, and can thus influence extracellular DA neurotransmission. Transgenic mice have been generated with a dramatically reduced (by approximately 95%) expression of the VMAT2 gene which, unlike complete knockout lines, survive into adulthood. We compared the pre-synaptic regulation of both impulse-dependent (exocytotic) and carrier-mediated (via reversal of the DA transporter, DAT) DA release in the dorsolateral caudate putamen (CPu) of striatal slices derived from adult homozygous VMAT2 mutant and wild-type mice using fast cyclic voltammetry. Impulse-dependent DA release, evoked by a single electrical pulse, was lower in homozygous (116 nm) than wild-type mice (351 nm) indicating smaller vesicular DA stores, an observation supported by the evanescent effect of amfonelic acid (300 nm) in homozygous mice. Amphetamine (2 microm) increased extracellular DA via DAT reversal in both wild-type (by 459 nm) and VMAT2 mutant (by 168 nm, p < 0.01 vs. wild-type) mice. In both cases, the effect was blocked by the DAT inhibitor GBR12935 (1 microm). Simultaneously, amphetamine decreased impulse-dependent DA release, albeit less in homozygous (by 55%) than in wild-type (by 78%) mice. In wild-types, this decrement was largely reversed by GBR12935 but not by the D2/D3 autoreceptor antagonist (-)sulpiride (1 microm). Conversely, in homozygous VMAT2 mutant mice, it was attenuated by (-)sulpiride but not GBR12935. The D2/D3 receptor agonist quinpirole inhibited impulse-dependent DA release with a lower EC50 value in homozygous mice (12 nm) compared with wild-types (34 nm), indicating the compensatory presence of functionally supersensitive release-regulating autoreceptors. However, analysis of DA reuptake kinetics obtained in the absence and presence of DAT blockade (by cocaine and amfonelic acid) revealed only minor differences in DAT functionality. These results demonstrate that impaired vesicular DA storage constrains extracellular DA levels in the dorsolateral CPu whether induced by either impulse-dependent or carrier-mediated mechanisms and that the relative importance of the DAT and terminal autoreceptors as control mechanisms in the actions of amphetamine are reversed in VMAT2 mutant mice.     

A review of the effects of dopaminergic agents on humans,animals, and drug-seeking behavior,and its implications for medication development

Medication development for cocaine abuse has focused on potential mechanisms of action related to the abuse of cocaine. The hypothesis that mesolimbic dopamine (DA) is the key neurochemical mediator of cocaine’s addictive and reinforcing effects is well supported by a wide variety of data from animal studies. On the other hand, medications that increase DA or block its actions in humans can produce effects that appear incompatible with this hypothesis. This article reviews these incompatibilities between animal and human data with a focus on the DAergic actions of drugs, including DA reuptake inhibitors, direct DA agonists, DA increasers, and DA antagonists. Possible reasons for these discrepancies are discussed, and the potential role of high-affinity DA uptake inhibitors, such as GBR12909, for pharmacotherapeutic application to treat cocaine abuse is discussed. Since progress in developing pharmacotherapies for treating cocaine addiction in humans is likely to come from understanding its mechanisms of action, it is clear that further research on the effects of cocaine in humans and animals will be critical to the medication development effort. A shortened version of this paper was presented at the Satellite Meeting of the International Society for Neurochemistry on “Cellular and Molecular Mechanisms of Drugs of Abuse: Cocaine and Methamphetamine” held on August 19–20, 1993 in Nice, France.     

Differential effects of d-amphetamine, β-phenylethylamine,cocaine and methylphenidate on the rate of dopamine synthesis in terminals of nigrostriatal and mesolimbic neurons and on the efflux of dopamine metabolites into cerebroventricular perfusates of rats

The $\text{in}$$\text{vivo}$ of four psychomotor stimulants (d-amphetamine, β-phenylethylamine, cocaine and methylphenidate) were determined on: 1) the rate of dopamine (DA) synthesis, as measured by the accumulation of dihydroxyphenylalanine (DOPA) after aromatic L-amino acid decarboxylase inhibition, in the striatum (terminals of nigrostriatal neurons) and in the nucleus accumbens and olfactory tubercle (terminals of mesolimbic neurons) and 2) the efflux of the DA metabolite 3,4-dihydroxyphenylacetic acid (DOPAC) into cerebroventricular perfusates of conscious, freely-moving rats. d-Amphetamine and β-phenylethylamine produced biphasic responses with lower doses of each drug increasing both the efflux of DOPAC and the rate of DA synthesis in the striatum. Higher doses of each drug either had no effect or actually decreased the efflux of DOPAC and also decreased the rate of DA synthesis in the striatum. Higher doses of each drug either had no effect only decreased the efflux of DOPAC and the rate of DA synthesis in the striatum. The effects of the drugs on the rate of DA synthesis in the nucleus accumbens and olfactory tubercle were similar to, but less pronounced than those seen in the striatum. These results are consistent with the following suggestions: 1) low doses of d-amphetamine and β-phenylethylamine facilitate the neuronal release of DA while higher doses of both drugs facilitate release and inhibit neuronal reuptake of the amine, and 2) cocaine and methylphenidate preferentially block the neuronal reuptake of DA.     

Differential regulation of accumbal dopamine transmission in rats following cocaine, heroin and speedball self-administration

Cocaine/heroin combinations (speedball) exert synergistic neurochemical and behavioral effects that are thought to contribute to the increased abuse potential and subjective effects reported by polydrug users. In vivo fast-scan cyclic voltammetry was used to examine the effects of chronic intravenous self-administration (25 consecutive sessions) of cocaine (250 μg/inf), heroin (4.95 μg/inf) and speedball (250/4.95 μg/inf cocaine/heroin) on changes in electrically evoked dopamine (DA) efflux, maximal rate of DA uptake (V(max)) and the apparent affinity (K(m)) of the DA transporter in the nucleus accumbens. The increase in electrically evoked DA was comparable following cocaine and speedball injection; however, heroin did not increase evoked DA. DA transporter K(m) values were similarly elevated following cocaine and speedball, but unaffected by heroin. However, speedball self-administration significantly increased baseline V(max), while heroin and cocaine did not change baseline V(max), compared with the baseline V(max) values of drug-na?ve animals. Overall, elevated DA clearance is a likely consequence of synergistic elevations of nucleus accumbens extracellular DA concentrations by chronic speedball self-administration, as reported previously in microdialysis studies. The present results indicate neuroadaptive processes that are unique to cocaine/heroin combinations and cannot be readily explained by simple additivity of changes observed with cocaine and heroin alone.     

Ketanserin pretreatment attenuates MDMA-induced dopamine release in the striatum as measured by in vivo microdialysis

Systemic administration of the amphetamine analogue, 3,4-methylenedioxymethamphetamine (MDMA) produced a dose-dependent increase in the extracellular concentration of dopamine (DA) in the striatum as measured by in vivo microdialysis in awake, freely-moving rats. The extracellular concentration of the DA metabolite, 3,4-dihydroxyphenylacetic acid (DOPAC), was significantly decreased in dialysate samples following the administration of MDMA (10 and 20 mg/kg, i.p.). The serotonin-2 (5-HT2) antagonist ketanserin (3 mg/kg, i.p.) had no effect on the extracellular concentration of DA or DOPAC in the striatum of vehicle- treated rats. The administration of ketanserin (3 mg/kg) 1 hr prior to MDMA (20 mg/kg) significantly attenuated the MDMA- induced increase in the extracellular concentration of DA without affecting the decrease in DOPAC concentrations. These data are suggestive that MDMA administration increases DA release in the striatum of awake, freely-moving rats. In addition, MDMA-induced increase in the extracellular concentration of DA in the striatum is mediated, in part, via 5-HT2 receptor mechanisms.     

Differences in dopamine responsiveness to drugs of abuse in the nucleus accumbens shell and core of Lewis and Fischer 344 rats

The use of inbred rat strains provides a tool to investigate the role of genetic factors in drug abuse. Two such strains are Lewis and Fischer 344 rats. Although several biochemical and hormonal differences have been observed between Lewis and Fischer 344 strains, a systematic comparison of the effect of different drugs of abuse on dopamine (DA) transmission in the shell and core of the nucleus accumbens of these strains is lacking. We therefore investigated, by means of dual probe microdialysis, the effect of different doses of morphine (1.0, 2.5, and 5.0 mg/kg), amphetamine (0.25, 0.5, and 1.0 mg/kg) and cocaine (5, 10, and 20 mg/kg) on DA transmission in the shell and in the core of nucleus accumbens. Behavior was monitored during microdialysis. In general, Lewis rats showed greater DA responsiveness in the NAc core compared to F344 rats except after 2.5 mg/kg of morphine and 20 mg/kg of cocaine. In the NAc shell, different effects were obtained depending on drug and dose: after 1.0 mg/kg of morphine no strain differences were observed, at 2.5 and 5.0 mg/kg Lewis rats showed greater increase in DA in the NAc shell. Following amphetamine and cocaine challenge, Lewis rats showed greater DA increase in the shell after 0.25 mg/kg of amphetamine and 20 mg/kg of cocaine. Behavioral activation was greater in Lewis rats in response to the lowest dose of morphine (1.0 mg/kg), to the highest dose of amphetamine (1.0 mg/kg) and to all doses of cocaine. These differences might be the basis for the different behavioral responses of these strains to drugs of abuse.     

Slow potentials of the olfactory bulb in the carp

The slow negative potentials evoked in carp olfactory bulb (OB) by some odorants and slow positive potentials evoked by nonspecific irritation (water stream, NaCl solutions) of olfactory epithelium have been studied. The slow potentials of both types were not inverted in deep layers of OB and were resistant to blockade of synaptic transmission by manganese ions. The negative slow potentials were not also affected by hypoxia and associated with local increase of OB tissue resistance. Positive slow potentials were affected by hypoxia and associated with local decrease of OB tissue resistance. The electrical tetanization of local zones of olfactory epithelium evoked in OB steady potential shifts of negative polarity, but diffuse tetanization of olfactory nerve evoked shifts of positive polarity. The results support the hypothesis of glial origin of slow potentials. Possible mechanisms of slow negative and positive potential generation are discussed.     

New findings on nuclear gangliosides: overview on metabolism and function

Dopamine (DA) is an important transmitter in both motor and limbic pathways. We sought to investigate the role of D(1)-receptor activation in axonal DA release regulation in dorsal striatum using a D(1)-receptor antagonist, SKF-83566. Evoked DA release was monitored in rat striatal slices using fast-scan cyclic voltammetry. SKF-83566 caused a concentration-dependent increase in peak single-pulse evoked extracellular DA concentration, with a maximum increase of ～ 65% in 5 μM SKF-83566. This was accompanied by a concentration-dependent increase in extracellular DA concentration clearance time. Both effects were occluded by nomifensine (1 μM), a dopamine transporter (DAT) inhibitor, suggesting that SKF-83566 acted via the DAT. We tested this by examining [(3)H]DA uptake into LLc-PK cells expressing rat DAT, and confirmed that SKF-83566 is a competitive DAT inhibitor with an IC(50) of 5.7 μM. Binding studies with [(3)H]CFT, a cocaine analog, showed even more potent action of SKF-83566 at the DAT cocaine binding site (IC(50) = 0.51 μM). Thus, data obtained using SKF-83566 as a D(1) DA-receptor antagonist may be confounded by concurrent DAT inhibition. More positively, however, SKF-83566 might be a candidate to attenuate cocaine effects in vivo because of the greater potency of this drug at the cocaine versus DA binding site of the DAT.     

Differences in the in vivo dynamics of neurotransmitter release and serotonin uptake after acute para-methoxyamphetamine and 3,4-methylenedioxymethamphetamine revealed by chronoamperometry

Illicit use of p-methoxyamphetamine (PMA) is rapidly increasing. However, little is known about the acute effects of PMA on neurotransmission in vivo. High-speed chronoamperometry was used to monitor neurotransmitter release and clearance in anesthetized rats after local application of PMA or 3,4-methylenedioxymethamphetamine (MDMA). In striatum, PMA caused less neurotransmitter release than MDMA. PMA-evoked release could be partially blocked by pre-treatment with a serotonin (5-HT) reuptake inhibitor, suggesting that evoked 5-HT release contributed to the electrochemical signal and was mediated by the 5-HT transporter (SERT). MDMA-evoked release was not blocked by a SERT inhibitor, suggesting that primarily DA was released. To study the effect of these amphetamines on clearance of 5-HT mediated specifically by the SERT, clearance of exogenously applied 5-HT was measured in the CA3 region of the hippocampus. In contrast to the striatum where 5-HT is cleared by both the SERT and the dopamine transporter (DAT), 5-HT is cleared primarily by the SERT in the CA3 region. This is also a region where neither PMA nor MDMA evoked release of neurotransmitter. The maximal inhibition of 5-HT clearance was greater after PMA than MDMA. These data demonstrate in vivo (1) brain region variability in the ability of PMA and MDMA to evoke release of neurotransmitter; (2) that clearance of 5-HT in the striatum is mediated by both the SERT and the DAT; (3) distinct differences in the amount and nature of neurotransmitter released in the striatum after local application of PMA and MDMA and (4) that PMA is a more efficacious inhibitor of 5-HT clearance in the hippocampus than MDMA. These fundamental differences may account for the more severe adverse reactions seen clinically after PMA, compared to MDMA.     

Rhythmic activity of the olfactory bulb of the carp under nembutal anesthesia

The effect of nembutal intraperitoneal injections upon the induced waves and orthodromic potentials of the olfactory bulb (OB) induced by odor stimulation and paired electric stimuli of the olfactory nerve was examined in immobilized by diplacine carp (cyprinus carpio L.). The nembutal anesthesia resulted in a gradual decrease in frequency of induced waves tested by orthodromic response and amplitude of spontaneous electric activity ob OB. The changes found under nembutal anesthesia in evoked electric activity of OB were also observed in chronic and acute intersection of the olfactory tracts.     

Role of Synaptophysin in Exocytotic Release of Dopamine from Xenopus Oocytes Injected with Rat Brain mRNA

1. The role of synaptophysin in the exocytotic release of dopamine (DA) was examined in Xenopus laevis oocytes injected with rat brain mRNA.2. The mRNA-injected oocytes showed DA uptake which depended on the incubation time and external DA concentrations.3. Stimulation with KCl (10–50 mM) of mRNA-injected oocytes preloaded with DA evoked external Ca²⁺-dependent release of DA. The noninjected and water-injected oocytes did not produce uptake of DA and stimulation-evoked release of DA.4. The high-KCl (50 mM)-stimulated release of DA decreased in the oocytes injected with rat brain mRNA together with antibody to synaptophysin.5. Immunoblot analysis demonstrated that synaptophysin was expressed in the brain mRNA-injected oocytes but not in the noninjected and water-injected oocytes.6. Thus, uptake and release machinery similar to native dopaminergic nerve terminals was expressed in Xenopus oocytes by injecting mRNA-extracted from the rat brain, and synaptophysin may play a role in the exocytotic release of DA.     

In vivo electrochemical studies of the effects of cocaine on dopamine nerve terminals in the rat neostriatum

In vivo electrochemical measurements, involving chronoamperometric recordings using monoamine-selective Nafion-coated electrodes, were used to study the effects of locally applied cocaine (50-500 micromolar barrel concentrations) on dopamine (DA) nerve terminals in the neostriatum of the anaesthetized rat. Local application of cocaine did not elicit detectable increases in basal levels of extracellular DA. However, locally applied cocaine significantly augmented the concentration of DA detected following a potassium (K+)-evoked depolarization. Data obtained with a new high-speed chronoamperometric recording technique further support that DA is the predominant species detected electrochemically following potassium-evoked depolarizations both before and after local application of cocaine. Unlike other locally applied uptake inhibitors that we have studied, cocaine failed to augment the time dynamics of released DA. In addition, large doses of the highest concentration of cocaine caused an attenuation of K+-evoked DA release, presumably due to cocaine's local anaesthetic properties. These data suggest that cocaine elevates synaptic levels of DA, but in a manner that is not identical to other potent monoamine uptake inhibitors.     

Differential effects of the pharmacological manipulation of serotonin systems on cocaine and amphetamine self-administration in rats

The effects of the administration of serotonergic drugs on infusion rates of rats self-administering cocaine and amphetamine on an FR-10 schedule of reinforcement in daily 4 hour sessions were compared. Pretreatment with fluoxetine (2.5, 5, and 10 mg/kg), an inhibitor of serotonin reuptake, significantly decreased rates of responding maintained by amphetamine, but had no effect on responding maintained by cocaine at any of the doses tested. Pretreatment with cinanserin (3, 10, and 17.5 mg/kg), a serotonergic receptor antagonist, decreased rates of amphetamine self-administration at the highest dose tested, and also had no effect on cocaine self-administration. These data suggest a differential sensitivity of cocaine and amphetamine self-administration to pharmacological manipulation of central serotonin systems. They are consistent with biochemical data which demonstrates a negative correlation between the reinforcing potency of amphetamine-like drugs, but not cocaine-like drugs and their potency at serotonin binding sites.     

Effect of Decreased Oxygen on In Vitro Release of Endogenous 3,4-Dihydroxyphenylethylamine from Mouse Striatum

The effects of hypoxia on release of endogenous 3,4-dihydroxyphenylethylamine (DA, dopamine) were investigated in mouse striatal slices. Following a 60-min preincubation, potassium increased DA release 12 times between zero and 1 min. By 10 min, uptake processes exceeded release and DA levels in the media decreased. Hypoxia (low oxygen) and anoxia (no oxygen) increased DA in the media by 120 and 205%, respectively, but did not alter dihydroxyphenylacetic acid concentrations. Under similar conditions, anoxia increased [3H]DA uptake eight-fold. For the uptake studies, the amount of DA added to the media was critical; in the presence of high concentrations of DA, anoxia reduced reuptake. Regardless of exogenous DA, hypoxia and anoxia increased extracellular DA, which may play a role in ischemic cell damage.     

Mechanism of action of methylmercury on in vivo striatal dopamine release. Possible involvement of dopamine transporter

Methylmercury (MeHg) produces significant increases in the spontaneous output of dopamine (DA) from rat striatal tissue. The mechanism through MeHg produces such increase in the extracellular DA levels could be due to increased DA release or decreased DA uptake into DA terminals. One of the aims of this study was to investigate the role of DA transporter (DAT) in the MeHg-induced DA release. Coinfusion of 400 microM MeHg and nomifensine (50 microM) or amphetamine (50 microM) produced increases in the release of DA similar to those produced by nomifensine and amphetamine alone. In the same way, MeHg-induced DA release was not attenuated under Ca(2+)-free conditions or after pretreatment with reserpine (10 mg/kg i.p.) or tetrodotoxin (TTX), suggesting that the DA release was independent of calcium and vesicular stores, as well as it was not affected by the blockade of voltage sensitive sodium channels. Thus, to investigate whether depolarization of dopaminergic terminal was able to affect MeHg-induced DA release, we infused 75 mM KCl through the dialysis membrane. Our results clearly showed a decrease induced by MeHg in the KCl-evoked DA release. Taken together, these results suggest that MeHg induces release of DA via transporter-dependent, calcium- and vesicular-independent mechanism and it decreases the KCl-evoked DA release.     

Subchronic steroid administration induces long lasting changes in neurochemical and behavioral response to cocaine in rats

The abuse of anabolic androgenic steroids (AASs), such as nandrolone, is not only a problem in the world of sports but is associated with the polydrug use of non-athletes. Among other adverse effects, AAS abuse has been associated with long term or even persistent psychiatric problems. We have previously found that nandrolone decanoate treatment could produce prolonged changes in rats’ brain reward circuits associated to drug dependence. The aim in this study was to evaluate whether AAS-induced neurochemical and behavioral changes are reversible.The increases in extracellular dopamine (DA) and serotonin (5-HT) concentration, as well as stereotyped behavior and locomotor activity (LMA) evoked by cocaine were attenuated by pretreatment with nandrolone. The recovery period, which was needed for the DA system to return back to the basic level, was fairly long compared to the dosing period of the steroid. In the 5-HT system, the time that system needed to return back to the basal level, was even longer than in the DA system. The attenuation was still seen though there were no detectable traces of nandrolone in the blood samples.Given that accumbal outflow of DA and 5-HT, as well as LMA and stereotyped behavior are all related to reward of stimulant drugs, this study suggests that nandrolone decanoate has significant, long-lasting but reversible effects on the rewarding properties of cocaine.