Inhibition of Brain Type A Monoamine Oxidase and 5-Hydroxytryptamine Uptake by Two Amphetamine Metabolites, p-Hydroxyamphetamine and p-Hydroxynorephedrine

Two amphetamine metabolites, p-hydroxyamphetamine (p-OHA) and p-hydroxynorephedrine (p-OHN), selectively inhibited the A form of monoamine oxidase (MAO) in rat and mouse forebrain homogenates. Of these two metabolites, p-OHA inhibited MAO-A more strongly than p-OHN. This MAO-A-selective inhibition by p-OHA or p-OHN was found to be competitive with respect to deamination of its substrate, 5-hydroxytryptamine (5-HT). The degree of MAO-A inhibition was not changed by 90 min of preincubation of the enzyme preparations with either metabolite, and the activity inhibited by p-OHA after the preincubation recovered completely to the control level after repeated washing. Uptake of 5-HT or dopamine into mouse forebrain synaptosomes was highly reduced by both p-OHA and p-OHN. Both metabolites were more potent in reducing dopamine uptake than in reducing 5-HT uptake. In reduction of 5-HT and of dopamine uptake, p-OHA was more potent than p-OHN. These results indicate that p-OHA is a more selective inhibitor of brain MAO-A activity and 5-HT uptake than its subsequent metabolite, p-OHN. These two actions of p-OHA might, together with possible 5-HT efflux into the synaptic cleft, greatly contribute to head twitch, a brain 5-HT-mediated animal behavior induced by p-OHA.     

High-performance liquid chromatography of methamphetamine and its related compounds in human urine following derivatization with fluorescein isothiocyanate

A high-performance liquid chromatographic method with fluorescence detection for the determination of methamphetamine and its related compounds is reported. Methamphetamine, amphetamine, norephedrine, p-hydroxymethamphetamine and 1-phenylethylamine as an internal standard were extracted from human urine, derivatized with fluorescein-4-isothiocyanate, and then separated on a reversed-phase column within 36 min. The fluorescence intensity of the effluent was monitored at excitation and emission wavelengths of 496 and 518 nm, respectively. Calibration curves were confirmed to be linear up to at least 100 pmol on the column with a correlation coefficient (r) of 0.994–0.999 for the target compounds. The detection limits (S/N=3) were 55–105 fmol per 20-μl injection. The method was successfully applied to urine samples taken from methamphetamine addicts.     

Amphetamine-induced locomotion and gene expression are altered in BDNF heterozygous mice

Administration of amphetamine overstimulates medium spiny neurons (MSNs) by releasing dopamine and glutamate from afferents in the striatum. However, these afferents also release brain-derived neurotrophic factor (BDNF) that protects striatal MSNs from overstimulation. Intriguingly, all three neurochemicals increase opioid gene expression in MSNs. In contrast, striatal opioid expression is less in naive BDNF heterozygous (BDNF(+/-)) vs. wild-type (WT) mice. This study was designed to determine whether partial genetic depletion of BDNF influences the behavioral and molecular response to an acute amphetamine injection. An acute injection of amphetamine [5 mg/kg, intraperitoneal (i.p.)] or saline was administered to WT and BDNF(+/-) mice. WT and BDNF(+/-) mice exhibited similar locomotor activity during habituation, whereas BDNF(+/-) mice exhibited more prolonged locomotor activation during the third hour after injection of amphetamine. Three hours after amphetamine injection, there was an increase of preprodynorphin mRNA in the caudate putamen and nucleus accumbens (Acb) and dopamine D(3) receptor mRNA levels were increased in the Acb of BDNF(+/-) and WT mice. Striatal/cortical trkB and BDNF, and mesencephalic tyrosine hydroxylase mRNA levels were only increased in WT mice. These results indicate that BDNF modifies the locomotor responses of mice to acute amphetamine and differentially regulates amphetamine-induced gene expression.     

“Sex, drugs and the brain”: The interaction between drugs of abuse and sexual behavior in the female rat

Preclinical and clinical research investigating female sexual motivation has lagged behind research on male sexual function. The present review summarizes recent advances in our understanding of the specific roles of various brain areas, as well as our understanding of the role of dopaminergic neurotransmission in sexual motivation of the female rat. A number of behavioral paradigms that can be used to thoroughly evaluate sexual behavior in the female rat are first discussed. Although traditional assessment of the reflexive, lordosis posture has been useful in understanding the neuroanatomical and neurochemical systems that contribute to copulatory behavior, the additional behavioral paradigms described in this review have helped us expand our understanding of appetitive and consumatory behavioral patterns that better assess sexual motivation - the equivalent of “desire” in humans. A summary of numerous lesion studies indicates that different areas of the brain, including forebrain and midbrain structures, work together to produce the complex repertoire of female sexual behavior. In addition, by investigating the effects of commonly addictive drugs, we are beginning to elucidate the role of dopaminergic neurotransmission in female sexual motivation. Consequently, research in this area may contribute to meaningful advances in the treatment of human female sexual dysfunction.     

Enhanced dopamine function in DISC1‐L100P mutant mice: implications for schizophrenia

Significant advances have been made in understanding the role of disrupted‐in‐schizophrenia‐1 (DISC1) in the brain and accumulating findings suggest the possible implication of DISC1 in the regulation of dopamine (DA) function. A mutation in the second exon of DISC1 at L100P leads to the development of schizophrenia‐related behavior in mutant mice (DISC1‐L100P). We investigated here the role of DA in the expression of schizophrenia‐related endophenotypes in the DISC1‐L100P genetic mouse model. The mutated DISC1 resulted in facilitation of the psychostimulant effect of amphetamine in DISC1‐L100P mutant mice assessed in the open field and prepulse inhibition (PPI) tests. Biochemical studies detected a 2.1‐fold increase in the proportion of striatal D receptors without significant changes in DA release in vivo in the striatum of DISC1‐L100P mutants in response to the low dose of amphetamine. The D₂ receptor antagonist haloperidol reversed the hyperactivity, PPI and latent inhibition (LI) deficits and blocked the psychostimulant effect of amphetamine in DISC1‐L100P mutants. Taken together, our findings show the role of DISC1 in D₂‐related pathophysiological mechanism of schizophrenia, linking DISC1 with well‐established DA hypothesis of schizophrenia.     

Iron Deficiency Alters the Day‐Night Variation in Monoamine Levels in Mice

Monoamine metabolism in the central nervous system is altered by dietary iron deficiency, with a stronger effect seen during the active than rest span of the circadian cycle. In this report, we examined changes in intracellular and extracellular monoamine levels, synthetic enzymes, transporter and receptor densities, and responses to amphetamine‐induced dopamine (DA) efflux in iron‐deficient and iron‐sufficient mice. Extracellular striatal DA levels were 15–20% higher in all groups during the active dark phase compared to the inactive light phase, with correspondingly lower dopamine transporter (DAT) and higher tyrosine hydroxylase levels. Iron deficiency decreased DAT density by 20% and 28% in the light and dark phases, respectively, and elevated the DOPAC/DA ratio only in the dark, indicating that iron deficiency does interact with the normal diurnal cues for cyclicity. Enhanced DA efflux after amphetamine stimulation indicates no limitation on monoamine synthesis and release and is consistent with altered synaptic efficacy and perhaps recycling of DA in iron deficiency. These experimental findings provide new evidence that brain iron insufficiency does have a differential effect on the DA system at different biological times of the day and night and may be causally related to the phasic motor symptoms observed in Restless Legs Syndrome.     

OPRM1 gene variants modulate amphetamine-induced euphoria in humans

The μ-opioid receptor is involved in the rewarding effects of not only opioids like morphine but also psychostimulants like amphetamine. This study aimed to investigate associations between subjective response to amphetamine and genetic polymorphisms and haplotypes in the μ-opioid receptor including the exonic variant rs1799971 (Asp40Asn). One hundred and sixty-two Caucasian volunteers participated in three sessions receiving either placebo or d-amphetamine (10 and 20 mg). Associations between levels of self-reported Euphoria, Energy and Stimulation [Addiction Research Center Inventory 49-item questionnaire (ARCI-49)] after d-amphetamine ingestion and polymorphisms in OPRM1 were investigated. The intronic single nucleotide polymorphisms (SNPs) rs510769 and rs2281617 were associated with significantly higher ratings of Euphoria, Energy and Stimulation after 10 mg amphetamine. Feelings of Euphoria, Energy and Stimulation were also found to be associated with a two-SNP haplotype formed with rs1799971 and rs510769 and a three-SNP haplotype formed with rs1918760, rs2281617 and rs1998220. These results support the hypothesis that genetic variability in the μ-opioid receptor gene influences the subjective effects of amphetamine and may suggest new strategies for prevention and treatment of psychostimulant abuse.     

Differential Effects of δ- and μ-Opioid Receptor Antagonists on the Amphetamine-Induced Increase in Extracellular Dopamine in Striatum and Nucleus Accumbens

Abstract: The specific opioid receptor antagonist naloxone attenuates the behavioral and neurochemical effects of amphetamine. Furthermore, the amphetamine-induced increase in locomotor activity is attenuated by intracisternally administered naltrindole, a selective δ-opioid receptor antagonist, but not by the irreversible μ-opioid receptor antagonist β-funaltrexamine. Therefore, this research was designed to determine if naltrindole would attenuate the neurochemical response to amphetamine as it did the behavioral response. In vivo microdialysis was used to monitor the change in extracellular concentrations of dopamine in awake rats. Naltrindole (3.0, 10, or 30 µg) or vehicle was given 15 min before and β-funaltrexamine (10 µg) or vehicle 24 h before the start of cumulative dosing, intracisternally in a 10-µl volume, while the rats were lightly anesthetized with methoxyflurane. Cumulative doses of subcutaneous d-amphetamine (0.0, 0.1, 0.4, 1.6, and 6.4 mg/kg) followed pretreatment injections at 30-min intervals. Dialysate samples were collected every 10 min from either the striatum or nucleus accumbens and analyzed for dopamine content by HPLC. Amphetamine dose-dependently increased dopamine content in both the striatum and nucleus accumbens, as reported previously. Naltrindole (3.0, 10, and 30 µg) significantly reduced the dopamine response to amphetamine in the striatum. In contrast, 30 µg of naltrindole did not modify the dopamine response to amphetamine in the nucleus accumbens. On the other hand, β-funaltrexamine (10 µg) had no effect in the striatum but significantly attenuated the amphetamine-induced increase in extracellular dopamine content in the nucleus accumbens. These data suggest that δ-opioid receptors play a relatively larger role than μ-opioid receptors in mediating the amphetamine-induced increase in extracellular dopamine content in the striatum, whereas μ-opioid receptors play a larger role in mediating these effects in the nucleus accumbens.     

Acute and Repeated Systemic Amphetamine Administration: Effects on Extracellular Glutamate, Aspartate, and Serine Levels in Rat Ventral Tegmental Area and Nucleus Accumbens

Abstract: Recent work indicates an important role for excitatory amino acids in behavioral sensitization to amphetamine. We therefore examined, using in vivo microdialysis in awake rats, the effects of amphetamine on efflux of glutamate, aspartate, and serine in the ventral tegmental area and nucleus accumbens, brain regions important for the initiation and expression of amphetamine sensitization, respectively. Water-pretreated and amphetamine-pretreated rats were compared to determine if sensitization altered such effects. In both brain regions, Ca²⁺-dependent efflux of glutamate accounted for ∼20% of basal glutamate efflux. A challenge injection of water or 2.5 mg/kg of amphetamine did not significantly alter glutamate, aspartate, or serine efflux in the ventral tegmental area or nucleus accumbens of water- or amphetamine-pretreated rats. However, 5 mg/kg of amphetamine produced a gradual increase in glutamate efflux in both regions that did not reverse, was observed in both water- and amphetamine-pretreated rats, and was prevented by haloperidol. Although increased glutamate efflux occurred with too great a delay to mediate acute behavioral responses to amphetamine, it is possible that repeated augmentation of glutamate efflux during repeated amphetamine administration results in compensatory changes in levels of excitatory amino acid receptors in the ventral tegmental area and nucleus accumbens that contribute to development or expression of amphetamine sensitization.     

Analogs of SR-141716A (Rimonabant) alter d-amphetamine-evoked [3H] dopamine overflow from preloaded striatal slices and amphetamine-induced hyperactivity

The CB(1) cannabinoid receptor antagonist SR-141716A (Rimonabant) markedly diminishes the behavioral effects of opiates and nicotine and has been an important tool to ascertain the role of cannabinoid receptors in drug addiction. The present goal was to determine the less-explored interaction of SR-141716A and d-amphetamine in neurochemical and behavioral assays. Additionally, the effect of the substituents and substitution patterns on the phenyl ring located at the 5 position of SR-141716A (4-chlorophenyl), and of the CB(1)/CB(2) cannabinoid receptor agonist WIN-55,212-2, was determined. SR-141716A, AM-251 (4-iodophenyl) and NIDA-41020 (4-methoxyphenyl) did not alter amphetamine-evoked [(3)H]overflow from rat striatal slices preloaded with [(3)H]dopamine. MRI-8273-30-1 (4-fluorophenyl; 0.1-10 microM) attenuated amphetamine (3 microM)-evoked [(3)H]overflow, and MRI-8273-59 (3,4-dichlorphenyl; 0.01-10 microM) augmented amphetamine (0.3-3 microM)-evoked [(3)H]overflow. WIN-55,212-2 was without effect. In a locomotor activity experiment, SR-141716A and MRI-8273-30-1 did not alter amphetamine-induced hyperactivity. However, MRI-8273-59 (1-3 mg/kg) dose-dependently attenuated amphetamine (1 mg/kg)-induced hyperactivity. The present results suggest that SR-141716A is less efficacious to alter amphetamine effects than its reported efficacy to diminish the effects of opiates and nicotine. Modification of the 5-phenyl position of SR-141716A affords compounds that do interact with amphetamine in vitro and in vivo.