Therapeutic doses of amphetamine or methylphenidate differentially increase synaptic and extracellular dopamine.

Methylphenidate (MP) and amphetamine (AMP) are first-line treatments for attention-deficit hyperactivity disorder. Although both drugs have similar therapeutic potencies, the stimulatory effect of AMP on extracellular dopamine (ECF DA) is greater than that of MP. We compared extracellular effects directly against synaptic changes. ECF DA was assessed by microdialysis in freely moving rodents and synaptic dopamine (DA) was measured using PET and [11C]-raclopride displacement in rodents and baboons. Microdialysis data demonstrated that MP (5.0 mg/kg, i.p.) increased ECF DA 360% +/- 31% in striatum, which was significantly less than that by AMP (2.5 mg/kg, i.p.; 1398% +/- 272%). This fourfold difference was not reflected by changes in synaptic DA. In fact, rodent PET studies showed no difference in striatal [11C]-raclopride binding induced by AMP (2.5 mg/kg, i.p.; 25% +/- 4% reduction) compared with that by MP (5.0 mg/kg, i.p.; 21% +/- 4% reduction). Primate PET experiments also showed no differences between AMP (0.5 mg/kg, i.v.; 24% +/- 4% reduction) and MP (1.0 mg/kg, i.v.; 25% +/- 7% reduction) induced changes in [11C]-raclopride binding potential. The similar potencies of MP and AMP to alter synaptic DA, despite their different potencies in raising ECF DA, could reflect their different molecular mechanisms.

Study of brain uptake and biodistribution of [11C]toluene in non-human primates and mice.

Inhalant abuse is a rapidly growing health problem particularly among adolescents. Yet we know little about the neural mechanisms underlying the abuse liability of inhalants, particularly when compared to other addictive drugs. Specifically, our understanding of the relationship between the regional brain phamacokinetics and features classically associated with drug reinforcement is lacking. Under the hypothesis that the abuse liability of toluene can be related to its pharmacokinetic properties and the pattern of regional brain uptake, we developed the methodology for radiolabeling and purifying [11C]toluene for use in PET studies. Here we report the regional brain distribution and kinetics of the widely abused solvent toluene in non-human primates and the whole body biodistribution in mice. To our knowledge, this is the first reported study of the in vivo brain pharmacokinetics of labeled toluene in non-human primates. Rapid uptake of radioactivity into striatal and frontal regions was followed by rapid clearance from the brain. Concurrent findings in rodents indicate similar radio-tracer kinetics, with excretion through kidneys and liver. Taken together, our data provides insight into pharmacokinetic features possibly associated with the abuse liability of toluene.

Gamma vinyl-GABA differentially modulates NMDA antagonist-induced increases in mesocortical versus mesolimbic DA transmission.

To explore the role of endogenous GABA in NMDA antagonist induced dopamine (DA) release, we used in vivo microdialysis to study the effects of pretreatment with gamma-vinyl GABA (GVG) on phencyclidine (PCP)-induced DA release in terminal regions of midbrain DA neurons. GVG, an irreversible inhibitor of the GABA catabolizing enzyme GABA-AT, significantly reduced the DA response to PCP (7.0 mg/kg) in freely moving animals. Preferential increases in PCP-induced DA release in the PFC (four-fold those of NAcc) were dose-dependently inhibited by acute pretreatment with GVG at doses of 150 (51% inhibition), 300 (68% inhibition), and 500 (82% inhibition) mg/kg, whereas NAcc PCP-induced DA activity was unresponsive to 150 mg/kg and only partially inhibited by 300 and 500 mg/kg. Subchronic treatment with GVG did not enhance the inhibitory capacity of the GABAergic system. While GVG evidently modulates PCP-induced increases in mesocorticolimbic DA transmission, the character of this modulation is regionally specific, with cortical NMDA-antagonist induced increases appearing more sensitive to inhibition by endogenous GABA than subcortical areas.

The selective sigma(1) receptor agonist, 1-(3,4-dimethoxyphenethyl)-4-(phenylpropyl)piperazine (SA4503), blocks the acquisition of the conditioned place preference response to (-)-nicotine in rats.

We examined the effect of the sigma(1) receptor agonist, 1-(3,4-dimethoxyphenethyl)-4-(phenylpropyl)piperazine (SA4503), on the acquisition of the conditioned place preference response to subcutaneously administered (-)-nicotine in rats. (-)-Nicotine, but not SA4503 or vehicle, produced a significant conditioned place preference response. Pretreatment of animals with either 1 or 3 mg/kg of SA4503 significantly attenuated the conditioned place preference response to (-)-nicotine.

Gamma-vinyl GABA (GVG) blocks expression of the conditioned place preference response to heroin in rats.

We examined the effect of gamma-vinyl GABA (GVG) on the expression of the conditioned place preference response to intraperitoneally (i.p.) administered heroin in rats. Heroin, but not vehicle, produced a significant conditioned place preference response. Pretreatment of animals with 300 mg/kg of GVG significantly attenuated the expression of the heroin-induced conditioned place preference response. These results are the first to suggest that systemic GVG may provide an effective alternative to methadone maintenance in the treatment of heroin addiction, since it is without abuse potential and can be used for treatment outside an institutional setting.

A consideration of the dopamine D2 receptor monomer-dimer equilibrium and the anomalous binding properties of the dopamine D2 receptor ligand, N-methyl spiperone.

Some discrepancies between experimental results with the two D2 antagonists N-methyl spiperone (NMSP) and raclopride (RAC) have been observed. Among these are the observation that MK-801 increases NMSP binding but not RAC binding: pretreatment with reserpine increases RAC binding but decreases NMSP binding; and that the two ligands yield different values for Bmax. It has been observed that the D2 receptor can exist in both a monomer and dimer form and that a NMSP photolabel ligand binds primarily to the monomer form while a RAC-like photolabel ligand binds both. Using a model of the dimerization in which the equilibrium dissociation constant increases with increasing dopamine (DA) concentration, the free monomer concentration can be shown to go through a maximum value with increasing DA. Using this model with data from a baboon PET study, it can be shown that under certain conditions an increase in binding could be observed. Further research may show that there are clusters of D2 receptors forming oligomers with more than two receptors in which NMSP binds to more sites on clusters with fewer receptors. If increasing DA favors cluster with fewer receptors, an increase in NMSP binding sites may also occur under some circumstances with an increase in DA.

The effect of gamma-vinyl-GABA on the consumption of concurrently available oral cocaine and ethanol in the rat.

It has frequently been reported that a high percentage of individuals, identified as either alcohol- or cocaine-dependent, concurrently abuse both drugs. The experiments reported here represent a continuing effort to develop an animal model to predict the effects of a potential pharmacotherapeutic agent on concurrently available oral ethanol and cocaine. These experiments utilized drinkometer circuitry to assess the effects of gamma-vinyl-GABA (GVG), a gamma-aminobutyric acid (GABA) transaminase inhibitor, on the consumption and temporal pattern of responses for orally self-administered ethanol and cocaine. The results of these experiments showed that GVG, at doses of 100, 200 and 300 mg/kg, reduced both ethanol and cocaine consumption in a dose-related manner. When compared to vehicle, GVG at all doses significantly reduced ethanol consumption while consumption of cocaine was significantly reduced only at 300 mg/kg. This is consistent with data showing that GVG reduces consumption of these drugs when administered alone and data showing that GVG is more potent in reducing ethanol-induced compared to cocaine-induced extracellular dopamine in the nucleus accumbens. Analysis of the temporal pattern of drinking across the session suggests that GVG's effects are due to a disruption of the reinforcing properties of ethanol and cocaine rather than a more general reduction in motor behavior. These data suggest that GVG has potential for clinical use in populations that abuse either alcohol or cocaine alone or in combination.

GABAergic blockade of cocaine-associated cue-induced increases in nucleus accumbens dopamine.

Environments previously associated with drug use can become one of the most common factors triggering relapse to drug-seeking behavior. To better understand the neurochemical mechanisms potentially mediating these cues, we measured nucleus accumbens dopamine levels in animals exposed to environmental cues previously paired with cocaine administration. In animals exposed to a cocaine-paired environment nucleus accumbens dopamine increased by 25%. When administered 2.5 h prior to presentation of the environmental trigger, racemic vigabatrin (an irreversible inhibitor of gamma-aminobutyric acid (GABA)-transaminase) abolished this cue-induced increase. Conversely, R-(-)-vigabatrin, the inactive enantiomer, had no effect. Combined with our earlier findings, these studies support the potential therapeutic benefit of this enzyme-based GABAergic strategy to modulate brain dopamine and the subsequent treatment of drug addiction.

Synergistic interactions between nicotine and cocaine or methylphenidate depend on the dose of dopamine transporter inhibitor.

There is a greater prevalence of cigarette smoking among cocaine-dependent individuals and hyperactive children treated with stimulants (e.g., methylphenidate, MP). However, little is known about the neurochemical basis of the interaction between nicotine and cocaine or MP. It is thought that the reinforcing effects of cocaine and MP are due partly to increases in synaptic DA in the nucleus accumbens (NAc). These measurable increases are secondary to the blockade of the DA transporter. In contrast, nicotine stimulates acetylcholine receptors located presynaptically on dopaminergic projections from the ventral tegmental area (VTA) to the NAc and increases DA transmission. Here we investigate the effects of nicotine on NAc DA in animals simultaneously injected with cocaine or MP. Coadministration of nicotine (0.4 mg/kg s.c.) and cocaine (10 mg/kg i.p.) or MP (5 mg/kg i.p.) increased the extracellular NAc DA levels in an additive manner, while coadministration of nicotine (0. 4 mg/kg s.c.) and a higher dose of cocaine (20 mg/kg) or MP (10 mg/kg) clearly produced a synergistic elevation in NAc DA. These findings suggest that the degree of DA transporter (DAT) occupancy contributes to the synergistic interaction between nicotine and cocaine or MP.

Comparison between intraperitoneal and oral methylphenidate administration: A microdialysis and locomotor activity study.

The therapeutic and stimulant properties of methylphenidate (MP), a drug commonly prescribed for the treatment of attention deficit hyperactivity disorder, have been attributed to increases in synaptic dopamine (DA) concentrations resulting from the blockade of DA transporters. In addition to obvious difficulties inherent in any interspecies comparison, interpretation of preclinical studies done with MP is further complicated by different routes of administration in animals (i.v. and i.p.) compared with humans (oral). In the present study we compared the effects of i.p. and intragastric (oral) MP both on rat nucleus accumbens DA assessed by in vivo microdialysis and on locomotor activity measured in a photocell apparatus. We also compared regional brain uptake and plasma levels of [(3)H]MP after administration of 5 mg/kg via both routes. Intraperitoneal MP (5 and 10 mg/kg) was approximately twice as potent as intragastric MP in terms of increasing extracellular DA levels and in stimulating locomotion. This was consistent with the higher brain uptake of [(3)H]MP when given i.p. rather than intragastrically. The dose of 2 mg/kg produced significant increases in both measurements when administered i.p., but not intragastrically. This study shows that relatively low doses of MP (2 mg i.p. and 5 mg intragastric) significantly increase extracellular DA and locomotor activity and indicates that the differences in the neurochemical and behavioral effects of MP between the intragastric and the i.p. routes are due to central drug bioavailability.

gamma-aminobutyric acid mimetic drugs differentially inhibit the dopaminergic response to cocaine.

Dopaminergic activity in the mesocorticolimbic system is associated with reinforcing properties of psychostimulant drugs. We previously demonstrated that increased gamma-aminobutyric acid (GABA)-ergic activity produced by gamma-vinyl GABA [D,L-4-amino-hex-5-enoic acid (Vigabatrin(R))], an irreversible inhibitor of GABA-transaminase, attenuated cocaine, nicotine, heroin, alcohol, and methamphetamine-induced increases in extracellular nucleus accumbens dopamine as well as behaviors associated with these biochemical changes. In the present study, using in vivo microdialysis techniques, we compared three different strategies to increase GABAergic activity in order to modulate cocaine-induced increase in extracellular dopamine. Our data demonstrate that the anticonvulsant 1-(2-(((diphenylmethylene)amino)oxy)ethyl)-1,2,5, 6-tetrahydro-3-pyridinecarboxylic acid hydrochloride (NNC-711), a GABA uptake inhibitor, dose and time dependently diminished increases in extracellular dopamine following acute cocaine challenge. Furthermore, we demonstrated that cyclized analogue of vigabatrin, a competitive reversible GABA-transaminase inhibitor, is a more potent inhibitor of cocaine-induced dopamine increase than vigabatrin. Our data suggest that in addition to irreversible inhibition of GABA transaminase, inhibition of GABA uptake represent another potentially effective, indirect strategy for the treatment of cocaine abuse.

Stereoselective inhibition of dopaminergic activity by gamma vinyl-GABA following a nicotine or cocaine challenge: a PET/microdialysis study.

Elevation of endogenous GABA by the racemic mixture of gamma vinyl-GABA (GVG, Vigabatrin) decreases extracellular nucleus accumbens (NAc) dopamine (DA) levels and diminishes the response to many drugs of abuse known to elevate DA in the mesocorticolimbic system. We investigated the effects of the individual enantiomers (S(+)-GVG, R(-)-GVG) on cocaine-induced NAc DA in rodents as well as the effects of nicotine-induced increases in primates. In a series of microdialysis experiments in freely moving animals, S(+)-GVG (150 mg/kg), R(-)-GVG (150 mg/kg) or racemic (R, S) GVG (300 mg/kg) was administered 2.5 hours prior to cocaine (20 mg/kg) administration. When compared with cocaine alone, the R(-) enantiomer did not significantly inhibit cocaine induced NAc DA release. S(+)-GVG, at half the dose of the racemic mixture (150 mg/kg), inhibited cocaine-induced DA elevation by 40%, while the racemic mixture (300 mg/kg) inhibited cocaine-induced DA release by 31%. In addition, our PET studies in primates demonstrated that S(+)-GVG completely inhibits nicotine-induced increases in the corpus striatum, again at half the dose of the racemic mixture. The R(-) enantiomer was ineffective. Although the S(+) enantiomer has been well established as the active compound in the treatment of epilepsy, the efficacy of this enantiomer with regard to mesolimbic DA inhibition generates a complex series of clinical and neurochemical issues. Further investigations will determine the locus of action and physiologic properties of each enantiomer.

Gamma-vinyl gamma-aminobutyric acid attenuates the synergistic elevations of nucleus accumbens dopamine produced by a cocaine/heroin (speedball) challenge.

In the present study, we examined the effect of an acute administration of the selective suicide inhibitor of gamma-aminobutyric acid (GABA)-transaminase, gamma-vinyl GABA on increases in nucleus accumbens dopamine produced by a cocaine/heroin challenge in freely moving animals. Cocaine (20 mg/kg, i.p.) produced an elevation in extracellular nucleus accumbens dopamine of approximately 380% above baseline, while heroin produced only a moderate increase of 70%. Coadministration of these two drugs, however, produced a synergistic elevation in nucleus accumbens dopamine of 1000%. This response was reduced by 50% in animals pretreated with gamma-vinyl GABA (300 mg/kg, i.p.) 2.5 h prior to challenge. This same dose of gamma-vinyl GABA inhibited cocaine-induced increases in nucleus accumbens dopamine by 25% and completely abolished heroin-induced increase. These findings indicate that gamma-vinyl GABA can interfere with the synergistic effects produced by the combination of an indirect dopamine releaser (heroin) and a dopamine reuptake blocker (cocaine).

Dopamine-transporter occupancy after intravenous doses of cocaine and methylphenidate in mice and humans.

OBJECTIVES: Recent studies using positron emission tomography (PET) have established the relationship between an intravenous dose of cocaine and the percentage occupancy of the dopamine transporter in humans, and have documented the requirement of more than 50% occupancy for perception of the "high". The present experiments were conducted to examine dose-occupancy and dose-effect relationships in mice for cocaine and also for methylphenidate, a dopamine uptake blocker used in pediatric psychiatry. METHODS: Percentage occupancies of the dopamine transporter by cocaine and methylphenidate were estimated after intravenous injection in mice from the displacement of in vivo binding of [(3)H]cocaine from the striatum. Locomotor activity was measured in a photocell apparatus. RESULTS: The relationship between drug doses (milligrams of hydrochloride salt per kilogram body weight) and percentage occupancy of the dopamine transporter was indistinguishable for cocaine and methylphenidate, and corresponded to about 50% occupancy at 0.25 mg/kg and about 80% at 1 mg/kg. This was similar to the relationship between drug dose and transporter occupancy, previously measured in human and baboons using [(11)C]cocaine or [(11)C]d-threo-methylphenidate and PET. Methylphenidate increased locomotor activity in the mice substantially more than cocaine at the same dose and the same degree of dopamine-transporter receptor occupancy. CONCLUSIONS: The range of dopamine-transporter occupancy required for behavioral activation in the mice was thus similar to that previously reported for experience of a cocaine- or methylphenidate-induced "high" in human subjects. Our results are consistent with other studies in which both cocaine and methylphenidate were evaluated in animal behavioral assays and were found to have very similar psychopharmacological properties.

Gamma-vinyl GABA inhibits methamphetamine, heroin, or ethanol-induced increases in nucleus accumbens dopamine.

We examined the acute effect of the irreversible GABA-transaminase inhibitor, gamma-vinyl GABA (GVG, Sabril((R)), Vigabatrin((R))) on increases in nucleus accumbens (NAc) dopamine (DA) following acute administration of methamphetamine, heroin, or ethanol. Methamphetamine (2.5 mg/kg) produced a dose-dependent increase (2, 700%) in NAc DA. GVG preadministration (300 or 600 mg/kg), however, inhibited this response by approximately 39 and 61%, respectively. The lower dose of methamphetamine (1.25 mg/kg), increased DA by 1, 700%. This response was inhibited to a similar extent (44%) regardless of the GVG dose preadministered (300 or 600 mg/kg). In addition, heroin-induced increases in NAc DA (0.5 mg/kg, 170%) were inhibited or completely abolished by GVG (150 or 300 mg/kg, 65 and 100%, respectively). Finally, at half the dose necessary for heroin, GVG (150 mg/kg) also completely abolished ethanol-induced increases in NAc DA following a 0.25 g/kg challenge dose (140%). Taken with our previous findings using nicotine or cocaine as the challenge drug, these results indicate that GVG attenuates increases in NAc DA by a mechanism common to many drugs of abuse. However, it appears unlikely that an acute dose of GVG can completely inhibit increases in NAc DA following challenges with a drug whose mechanism of action is mediated primarily through the DA reuptake site.

The irreversible gamma-aminobutyric acid (GABA) transaminase inhibitor gamma-vinyl-GABA blocks cocaine self-administration in rats.

gamma-Vinyl gamma-aminobutyric acid (GABA) (GVG) is an irreversible inhibitor of GABA transaminase, the primary enzyme involved in GABA metabolism. Acute administration of GVG increases brain GABA levels and blocks cocaine-induced locomotor activity, cocaine-induced lowering of brain stimulation reward thresholds, and cocaine-induced conditioned place preference. To further evaluate the effects of GVG on cocaine-induced reward, we examined its effects on cocaine self-administration in male Wistar rats on fixed ratio 5 and progressive ratio schedules of reinforcement. Additionally, the effects of GVG on operant responding for a food reward were examined on the same two schedules to determine whether the effects of GVG were specific to cocaine reward or generalized to other types of reward. GVG dose dependently decreased responding for cocaine on both schedules of reinforcement, suggesting that GVG attenuated the reward value of the cocaine. Responding for food was also decreased by GVG, suggesting that the effects of increased GABA levels induced by GVG may have a general effect on central reward systems. Data from this and other studies indicate that GVG does not induce motor impairment, decrease spontaneous locomotor activity, or induce catalepsy. Taken together, these data suggest that increases in GABAergic activity induced by GVG have an attenuating effect on centrally mediated reward systems and that the GABA system may be a useful target in the development of new therapeutic strategies for cocaine addiction.

PET and drug research and development.

The use of PET to examine the behavioral, therapeutic and toxic properties of drugs and substances of abuse is emerging as a powerful new scientific tool. PET provides a new perspective on drug research by virtue of its ability to directly assess both pharmacokinetic and pharmacodynamic events in humans and in animals. These parameters can be assessed directly in the human body both in healthy volunteers and in patients. Moreover, the new generation of high-resolution, small-animal cameras hold the promise of introducing imaging in the early stages of drug development and make it possible to carry out longitudinal studies in animals and to study genetically altered animals. This places PET in a unique position to contribute significantly to the process of drug development through understanding the molecular mechanisms underlying drug action while addressing some very practical questions such as determining effective drug doses for clinical trials for new drugs, determining the duration of drug action and examining potential drug interactions.

Comparable changes in synaptic dopamine induced by methylphenidate and by cocaine in the baboon brain.

Though the blockade of dopamine transporters (DAT) is associated with cocaine's and methylphenidate's reinforcing effects, it is the stimulation of dopamine (DA) receptors, achieved by increases in synaptic DA, that enables these effects to occur. Positron emission tomography (PET) and [11C]raclopride were used to assess the levels of occupancy of DA D2 receptors by dopamine achieved by doses of cocaine or methylphenidate previously documented to block over 70% of DAT. Studies were performed in five baboons using a paired scan protocol designed to measure DA D2 receptor availability (Bmax/Kd) at baseline conditions and after intravenous administration of either cocaine or methylphenidate. Cocaine (1-2 mg/kg) or methylphenidate (0.5 mg/kg) administered 5 min prior to [11C]raclopride decreased Bmax/Kd by 29+/-3% and 32 + 4%, respectively. Smaller reductions in Bmax/Kd (13% for cocaine given 30 min before [11C]raclopride and 25+/-10% for methylphenidate given 40 min before [11C]raclopride) were seen with longer periods between drug and radioligand. These observations are consistent with the slower striatal clearance kinetics of [11C]methylphenidate than [1C]cocaine observed in previous PET experiments and with the approximately twofold higher potency of methylphenidate than cocaine in in vitro experiments. Though the elevation of synaptic DA induced by >70% occupancy of DAT by these drugs lead to a modest increase in occupancy of D2 receptors (25-30%), further studies are required to assess if this is an underestimation because of differences in D2 receptor binding kinetics between raclopride and DA.

A pharmacologic strategy for the treatment of nicotine addiction.

Like many psychostimulant drugs, nicotine elevates extracellular and synaptic dopamine (DA) concentrations in the nucleus accumbens (NAc). This elevation has been linked to its reinforcing properties. Dopaminergic transmission within the NAc is modulated by gamma-aminobutyric acid (GABA). Therefore, we examined the utility of gamma vinyl-GABA (GVG, Vigabatrin) for inhibiting nicotine's biochemical effects on NAc DA as well as its effects on behaviors associated with these biochemical changes. Given 2.5 hours prior to nicotine, GVG (75 mg/kg) had no effect on nicotine-induced increases in extracellular NAc DA. However, at 90 mg/kg, GVG significantly inhibited nicotine-induced increases by approximately 50% while at 100 or 150 mg/kg, GVG completely abolished nicotine-induced increases in both naive and chronically nicotine-treated animals. When given 12 or 24 hours prior to nicotine administration at a dose of 100 mg/kg, GVG-induced inhibition was diminished or abolished, respectively. In addition, at a dose of 18.75 mg/kg GVG abolished the expression of nicotine-induced conditioned place preference (CPP) while a dose of 75 mg/kg abolished the acquisition phase of CPP. Finally, using positron emission tomography (PET) and 11C-raclopride in primates, GVG (100 mg/kg) abolished nicotine-induced increases in synaptic DA while having no effect on the rate of metabolism of the radiotracer or its regional distribution. Together, these data suggest that GVG may be useful for the treatment of nicotine addiction and further support the strategy of targeting the GABAergic system with a suicide inhibitor of GABA-transaminase for the treatment of drug addiction.