Chronic Amphetamine Exposure Research Articles

Amphetamine Self-Administration and Its Extinction Alter the 5-HT1B Receptor Protein Levels in Designated Structures of the Rat Brain

Manipulation of the serotonin (5-HT)1B receptors can modify the behavioral effects of amphetamine including its reinforcing properties. Focus of this study was to examine changes in 5-HT1B receptor protein expression in several brain structures linked to substance drug disorder in different stages of amphetamine addiction—single session of amphetamine self-administration, 20 consecutive days of amphetamine self-administration, and 3 and 14 days of extinction from chronic drug intake. “Yoked” procedure was employed to set apart pharmacological and motivational effects of amphetamine intoxication. Immunohistofluorescence was performed on brain slices containing the following regions: nucleus accumbens (NAc) shell and core, globus pallidum (GP) lateral and ventral, hippocampus (HIP), substantia nigra (SN), and ventral tegmental area (VTA). Single amphetamine session decreased the amount of 5-HT1B receptors in SN, VTA, and HIP in active and yoked rats. On the contrary, 20 days of chronic amphetamine exposure triggered elevation of 5-HT1B receptors exclusively in animals that voluntarily administered the drug in NAc core, GP ventral, and HIP. Furthermore, 14-day (but not 3-day) extinction from amphetamine increased the 5-HT1B receptor expression in ventral and lateral GP, HIP, and SN. This study is the first to demonstrate that exposure to amphetamine and its extinction alter the expression of 5-HT1B receptors in various rat brain regions, and those changes seem to be transient and region specific. Importantly, since increased expression of 5-HT1B receptor after chronic amphetamine self-administration was limited only to active group of animals, we suggest that 5-HT1B receptor is linked to motivational aspect of addiction.

Naltrexone modulates dopamine release following chronic, but not acute amphetamine administration: a translational study

The opioid antagonist naltrexone has been shown to attenuate the subjective effects of amphetamine. However, the mechanisms behind this modulatory effect are currently unknown. We hypothesized that naltrexone would diminish the striatal dopamine release induced by amphetamine, which is considered an important mechanism behind many of its stimulant properties. We used positron emission tomography and the dopamine D2-receptor radioligand [11C]raclopride in healthy subjects to study the dopaminergic effects of an amphetamine injection after pretreatment with naltrexone or placebo. In a rat model, we used microdialysis to study the modulatory effects of naltrexone on dopamine levels after acute and chronic amphetamine exposure. In healthy humans, naltrexone attenuated the subjective effects of amphetamine, confirming our previous results. Amphetamine produced a significant reduction in striatal radioligand binding, indicating increased levels of endogenous dopamine. However, there was no statistically significant effect of naltrexone on dopamine release. The same pattern was observed in rats, where an acute injection of amphetamine caused a significant rise in striatal dopamine levels, with no effect of naltrexone pretreatment. However, in a chronic model, naltrexone significantly attenuated the dopamine release caused by reinstatement of amphetamine. Collectively, these data suggest that the opioid system becomes engaged during the more chronic phase of drug use, evidenced by the modulatory effect of naltrexone on dopamine release following chronic amphetamine administration. The importance of opioid-dopamine interactions in the reinforcing and addictive effects of amphetamine is highlighted by the present findings and may help to facilitate medication development in the field of stimulant dependence.

Amphetamine treatment affects the extra‐hypothalamic vasopressinergic system in a sex‐ and nucleus‐dependent manner

The lateral septum (LS), a brain structure implicated in addictive behaviours, regulates the activation of dopaminergic neurones in the ventral tegmental area. Vasopressinergic projections from the extended amygdala to the LS, which are sexually dimorphic, could be responsible for the vulnerability to addiction in a sex-dependent manner. The present study aimed to investigate the modulatory effects of amphetamine (AMPH) on the expression of vasopressin (AVP) in the vasopressinergic extra-hypothalamic system in sensitised male and female rats. Adult male and female Sprague-Dawley rats underwent an AMPH-locomotor sensitisation protocol. Acute AMPH increased AVP mRNA expression in the medial amygdala (MeA), whereas AMPH-induced sensitisation increased AVP mRNA expression in the bed nucleus of the stria terminalis (BNST) only in females. Interestingly, the increase in AVP expression in BNST was higher in oestrus females compared to dioestrus females and acute AMPH resulted in a decrease in AVP levels in the LS, only in males. Thus, there are complex and region-specific interactions between AMPH and the extra-hypothalamic vasopressinergic system in the brain, underlying possible alterations in different behaviours caused by acute and chronic AMPH exposure.

Chronic amphetamine exposure causes long‐term effects on dopamine uptake

The dopamine transporter (DAT) is a plasma membrane transporter that clears released dopamine from the synaptic cleft; therefore it is an important regulator of extracellular levels of dopamine. DAT is also the main molecular target of many drugs of abuse, most notably the psychostimulants cocaine and amphetamine. Amphetamine (AMPH) is a substrate of DAT and competes for reuptake with dopamine. Several groups have shown that acute AMPH treatments alter the function and the number of DAT on the cell membrane. However, no data is available regarding the long‐term effects of AMPH on DAT activity. Data collected in our laboratory suggest that parental AMPH exposure affects dopamine uptake in C. elegans dopaminergic neurons isolated from progeny. These results led us to hypothesize that reduced dopamine reuptake in subsequent generations, as a result of parental AMPH exposure, is caused by down‐regulation of DAT. In order to overcome the complexity of multicellular organisms and the challenges of primary cell isolation and culture, where very few cells differentiate into dopaminergic neurons, we investigated the long‐term effects caused by AMPH in simplified systems. Initial experiments were carried out in LLC‐PK1 cells, stably transfected with human DAT. Cells were pre‐exposed to AMPH for 15 hours and uptake assays, carried out after cells had doubled, showed a significant decrease in dopamine uptake compared to control (untreated cells). These data suggest that changes caused by AMPH were maintained after one cell cycle. We repeated the same experiment using the human neuroblastoma SH‐SY5Y cells which can be differentiated into authentic dopaminergic neurons. Prior to the uptake assay one group of undifferentiated SH‐SY5Y cells was exposed to AMPH for 15 hours and the other group was treated with control solution. The cells were then allowed to grow and cross generations before they were treated with Retinoic Acid (RA) to induce differentiation. Uptake assays carried out 5 days after RA initiated differentiation revealed a significant decrease in dopamine uptake in AMPH treated groups with respect to control. These results suggest that parental AMPH pretreatment down regulates the expression or activity of DAT. We are currently investigating whether chronic exposure to AMPH in progenitor cells alters DAT expression in daughter cells.Support or Funding InformationCOBRE P20 GM104360

Food consumption and weight gain after cessation of chronic amphetamine administration

Cessation of drug use often coincides with increased food consumption and weight gain in recovering addicts. However, it is not known whether this phenomenon (particularly the weight gain) is uniquely human, or whether it represents a consequence of drug cessation common across species. To address this issue, rats (n = 10/group) were given systemic injections of D-amphetamine (3 mg/kg) or an equal volume of saline vehicle for 9 consecutive days. Beginning 2 days after the final injection, rats were given free access to a highly palatable food mixture (consisting of sugar and butter) along with their standard chow diet, and food consumption and body weight were measured every 48 h for 30 days. Consistent with clinical observations, amphetamine-treated rats showed a greater increase in body weight over the course of the 30 days relative to vehicle-treated rats. Surprisingly, there was no difference in highly palatable food consumption between amphetamine- and vehicle-treated groups, but the amphetamine-treated group consumed significantly more standard chow than the control group. The finding that a history of chronic amphetamine exposure increases food consumption is consistent with previous work in humans showing that withdrawal from drugs of abuse is associated with overeating and weight gain. The current findings may reflect amphetamine-induced sensitization of mechanisms involved in reward motivation, suggesting that weight gain following drug cessation in humans could be due to similar mechanisms.

The atypical dopamine transport inhibitor, JHW 007, prevents amphetamine-induced sensitization and synaptic reorganization within the nucleus accumbens

Benztropine (BZT) analogs, a family of agents with high affinity for the dopamine transporter have been postulated as potential treatments in stimulant abuse due to their ability to attenuate a wide range of effects evoked by psychomotor stimulants such as cocaine and amphetamine (AMPH). Repeating administration of drugs, including stimulants, can result in behavioral sensitization, a progressive increase in their psychomotor activating effects. We examined in mice the sensitizing effects and the neuroplasticity changes elicited by chronic AMPH exposure, and the modulation of these effects by the BZT derivative and atypical dopamine uptake inhibitor, JHW007, a candidate medication for stimulant abuse. The results indicated that JHW007 did not produce sensitized locomotor activity when given alone but prevented the sensitized motor behavior induced by chronic AMPH administration. Morphological analysis of medium spiny neurons of the nucleus accumbens revealed that JHW 007 prevented the neuroadaptations induced by chronic AMPH exposure, including increments in dendritic arborization, lengthening of dendritic processes and increases in spine density. Furthermore, data revealed that AMPH produced an increase in the density of asymmetric, possibly glutamatergic synapses in the nucleus accumbens, an effect that was also blocked by JHW007 pretreatment. The present observations demonstrate that JHW007 is able to prevent not only AMPH-induced behavioral sensitization but also the long-term structural changes induced by chronic AMPH in the nucleus accumbens. Such findings support the development and evaluation of BZT derivatives as possible leads for treatment in stimulant addiction.

Withdrawal effect of chronic amphetamine exposure during adolescence on complex maze performance

National survey data suggest a steady increase in the diagnosis and treatment of mental disorders in children, particularly Attention Deficit/Hyperactivity Disorder (ADHD). As nearly all children diagnosed with ADHD are prescribed stimulant drugs, rationale exists to quantitatively characterize behavioral responses following withdrawal from chronic stimulant dosing. These rodent experiments involved chronic administration of 7.5 mg/kg, s.c. amphetamine to subjects throughout adolescence followed by cognitive tests to gauge learning and performance during the withdrawal stage 7 to 14 days past withdrawal. Tests used a complex Stone 14-unit multiple T-maze, which is a robust paradigm for demonstrating age-related differences in rodent models when behavioral cognitive endpoints are used. Results reveal that amphetamine-treated subjects committed fewer major and retracing errors with increased minor errors and a significantly lower mean completion time. These findings suggest that pharmacotherapy aimed at adolescent-phase treatment of ADHD does not provoke spatial memory deficits at times proximal to drug withdrawal and lends support to amphetamine use in the treatment of ADHD children.

MRI reveals differential effects of amphetamine exposure on neuroglia in vivo

How amphetamine affects the neuroglia in living brains is not well understood. In an effort to elucidate this effect, we investigated neuroglia in response to amphetamine exposure using antisense (AS) or sense (S) phosphorothioate-modified oligodeoxynucleotide (sODN) sequences that correspond to glial fibrillary acidic protein (GFAP) mRNA (AS-gfap or S-gfap, respectively) expression. The control is a random-sequence sODN (Ran). Using cyanine 5.5-superparamagnetic iron oxide nanoparticle (Cy5.5-SPION) labeling and fluorescent microscopy, we demonstrated that living neural progenitor cells (PC-12.1), as well as the cells in fresh brain slices and intact brains of male C57BL6 mice, exhibited universal uptake of all of the sODNs but rapidly excluded all sODN-Ran and most S-gfap. Moreover, transmission electron microscopy revealed electron-dense nanoparticles only in the neuroglia of normal or transgenic mice [B6;DBA-Tg(Fos-tTA, Fos-EGFP*)1MmayTg(tetO-lacZ,tTA*)1Mmay/J] that had been administered AS-gfap or Cy5.5-SPION-gfap. Subtraction R2* maps from mice with acute and chronic amphetamine exposure demonstrated, validated by postmortem immunohistochemistry, a reduction in striatal neuroglia, with gliogenesis in the subventricular zone and the somatosensory cortex in vivo. The sensitivity of our unique gene transcript targeted MRI was illustrated by a positive linear correlation (r(2)=1.0) between in vivo MRI signal changes and GFAP mRNA copy numbers determined by ex vivo quantitative RT-PCR. The study provides direct evidence for targeting neuroglia by antisense DNA-based SPION-gfap that enables in vivo MRI of inaccessible tissue with PCR sensitivity. The results enable us to conclude that amphetamine induces toxicity to neuroglia in vivo, which may cause remodeling or reconnectivity of neuroglia.

Serotonergic neurotransmission in the ventral hippocampus is enhanced by corticosterone and altered by chronic amphetamine treatment

The ventral hippocampus modulates anxiety-like behavior in rats, and serotonergic transmission within the hippocampus facilitates adaptation to stress. Chronic amphetamine treatment results in anxiety-like behavior in rats and reduced monoamine concentrations in the ventral hippocampus. Since reduced hippocampal serotonergic transmission in response to stress is observed in rats that display high anxiety-like behavior, anxiety states in amphetamine-treated rats may be associated with reduced stress-related serotonergic transmission in the hippocampus. Therefore, using in vivo microdialysis in anesthetized rats, we investigated the effect of corticosterone infused locally into the ventral hippocampus on serotonergic transmission, and the effect of chronic amphetamine pretreatment on corticosteroid receptor protein expression and the corticosterone-induced serotonergic response. Extracellular serotonin in the ventral hippocampus was increased by corticosterone in drug naive rats, and this corticosterone-induced serotonin augmentation was blocked by the glucocorticoid receptor antagonist mifepristone. Furthermore, chronic pretreatment with amphetamine abolished the serotonin response to physiologically relevant corticosterone levels and reduced glucocorticoid receptor protein expression. Together, our results suggest that chronic amphetamine exposure reduces serotonergic neurotransmission, in part via alterations to glucocorticoid receptor-facilitation of serotonin release in the rat ventral hippocampus. Reduced serotonergic activity in the ventral hippocampus may contribute to altered stress responses and adaptive coping following repeated drug exposure.

Withdrawal from chronic exposure to amphetamine, but not nicotine, leads to an immediate and enduring deficit in motivated behavior without affecting social interaction in rats

Psychostimulant withdrawal leads to depressive symptoms, such as anhedonia and social dysfunction. We determined the effects of withdrawal from chronic exposure to nicotine (9 mg/kg/day salt, 28 days) or amphetamine (10 mg/kg/day salt, 7 days) on the motivated response for a sucrose reward and on social interaction in rats. Both nicotine and amphetamine exposure increased the motivated response for sucrose. However, only spontaneous amphetamine withdrawal led to an immediate and persistent decrease in motivated behavior, which was not correlated with body weight loss. Social interaction was not affected during withdrawal from either drug. These results indicate that withdrawal from chronic amphetamine exposure leads to an immediate and enduring anhedonic state.

Withdrawal from chronic amphetamine produces persistent anxiety-like behavior but temporally-limited reductions in monoamines and neurogenesis in the adult rat dentate gyrus

Acute amphetamine administration activates monoaminergic pathways and increases systemic corticosterone, both of which influence anxiety states and adult dentate gyrus neurogenesis. Chronic amphetamine increases anxiety states in rats when measured at 24 h and at 2 weeks of withdrawal. However, the effects of chronic amphetamine exposure and withdrawal on long term anxiety-like behavior and adult neurogenesis in the dentate gyrus are unknown. Adult male rats were administered amphetamine (2.5 mg/kg, ip.) daily for two weeks. Anxiety-like behaviors were increased markedly in amphetamine-treated rats following four weeks of withdrawal from amphetamine. Plasma corticosterone level was unaltered by amphetamine treatment or withdrawal. However, norepinephrine and serotonin concentrations were selectively reduced in the dentate gyrus 20 h following amphetamine treatment. This effect did not persist through the four week withdrawal period. In separate experiments, rats received bromodeoxyuridine to label cells in S-phase, prior to or immediately following amphetamine treatment. Newly generated cells were quantified to measure extent of progenitor cell proliferation and neurogenesis following treatment or withdrawal. Progenitor cell proliferation and neurogenesis were not significantly affected by amphetamine exposure when measured 20 h following the last amphetamine treatment. However, neurogenesis in the dentate gyrus was reduced after four weeks of withdrawal when compared to saline-pretreated rats. Overall, our findings indicate that withdrawal from chronic amphetamine leads to persistent anxiety-like behavior which may be maintained by reduced neurogenesis in the dentate gyrus at this protracted withdrawal time point. However, neurogenesis is unaffected at earlier withdrawal time points where anxiety states emerge, suggesting different mechanisms may underlie the emergence of anxiety states during amphetamine withdrawal.

Stability of surface NMDA receptors controls synaptic and behavioral adaptations to amphetamine

Plastic changes in glutamatergic synapses that lead to enduring drug craving and addiction are poorly understood. By focusing on the turnover and trafficking of NMDA receptors, we found that chronic exposure to the psychostimulant amphetamine induces selective downregulation of NMDA receptor NR2B subunits in the confined surface membrane pool of rat striatal neurons at synaptic sites. Remarkably, this downregulation is a long-lived event and results from the destabilization of surface-expressed NR2B due to accelerated ubiquitination and degradation of crucial NR2B-anchoring proteins by the ubiquitin-proteasome system. The biochemical loss of synaptic NR2B further translates to the significant modulation of synaptic plasticity in the form of long-term depression at cortico-accumbal glutamatergic synapses. Behaviorally, genetic disruption of NR2B induces, whereas restoration of NR2B loss prevents, behavioral sensitization to amphetamine. Our data identify NR2B as a key regulator in the remodeling of excitatory synapses and persistent psychomotor plasticity in response to amphetamine.

Long-lasting sensitization of reward-directed behavior by amphetamine

Exposure to psychostimulant drugs of abuse such as amphetamine can result in long-lasting “sensitization” of reward-directed behavior, such that subjects display enhancements in behavior directed by and toward rewards and reward-predictive cues (i.e. “incentive sensitization”). The purpose of these experiments was to determine the degree to which such sensitization resulting from chronic amphetamine exposure influences both appetitive and consummatory food-motivated behavior. Adult male Long-Evans rats received daily i.p. injections of d-amphetamine (2.0 mg/kg) or saline vehicle for five consecutive days. This amphetamine exposure regimen produced lasting sensitization to the acute locomotor stimulant effect of the drug. One month after drug exposure rats were tested for instrumental responding (lever pressing) for food reward under various response schedules. Two months after drug exposure, rats were tested for food consumption in a discriminative Pavlovian context-potentiated eating task, involving pairings of one context with food and another context with no food. Amphetamine exposed rats showed significantly greater instrumental responding for food reward than saline controls, particularly under conditions of high response ratios. In the potentiated eating task, testing under conditions of food satiation revealed that amphetamine exposed rats ate significantly more than saline controls in the food-paired context. These experiments demonstrate that amphetamine exposure can cause enduring increases in both appetitive and consummatory aspects of natural reward-directed behavior. Such long-lasting incentive sensitization could account in part for the propensity for relapse in drug addiction, as well as for reported enhancements in non-drug reward-related behavior.

ΔFosB Mediates Epigenetic Desensitization of the c-fosGene After Chronic Amphetamine Exposure

The molecular mechanisms underlying the transition from recreational drug use to chronic addiction remain poorly understood. One molecule implicated in this process is DeltaFosB, a transcription factor that accumulates in striatum after repeated drug exposure and mediates sensitized behavioral responses to psychostimulants and other drugs of abuse. The downstream transcriptional mechanisms by which DeltaFosB regulates drug-induced behaviors are incompletely understood. We reported previously the chromatin remodeling mechanisms by which DeltaFosB activates the expression of certain genes; however, the mechanisms underlying DeltaFosB-mediated gene repression remain unknown. Here, we identify c-fos, an immediate early gene rapidly induced in striatum after acute psychostimulant exposure, as a novel downstream target that is repressed chronically by DeltaFosB. We show that accumulation of DeltaFosB in striatum after chronic amphetamine treatment desensitizes c-fos mRNA induction to a subsequent drug dose. DeltaFosB desensitizes c-fos expression by recruiting histone deacetylase 1 (HDAC1) to the c-fos gene promoter, which, in turn, deacetylates surrounding histones and attenuates gene activity. Accordingly, local knock-out of HDAC1 in striatum abolishes amphetamine-induced desensitization of the c-fos gene. In concert, chronic amphetamine increases histone H3 methylation on the c-fos promoter, a chromatin modification also known to repress gene activity, as well as expression levels of the H3 histone methyltransferase, KMT1A (lysine methyltransferase 1A, formerly SUV39H1). This study reveals a novel epigenetic pathway through which DeltaFosB mediates distinct transcriptional programs that may ultimately alter behavioral plasticity to chronic amphetamine exposure.

Increased oxidative stress in submitochondrial particles after chronic amphetamine exposure

Previous studies have suggested that reactive oxygen species (ROS) production may play a role in the pathophysiology of many neuropsychiatric disorders, such as bipolar disorder (BD) and schizophrenia (SCZ). In addition, there is an emerging body of data indicating that BD and SCZ may be associated with mitochondrial dysfunction. We studied the effects of acute and chronic d-amphetamine on ROS production in submitochondrial particles of rat brain. Male Wistar rats were divided in two experimental groups: acute and chronic treatment. In the acute treatment, rats received one single IP injection of d-amphetamine (1, 2 or 4 mg/kg) or saline (control group). In the chronic treatment, rats received one daily IP injection of d-amphetamine (1, 2 or 4 mg/kg) or saline for 7 days. Locomotor activity was assessed with the open field task, and thiobarbituric acid reactive substances (TBARS) and superoxide production were measured in submitochondrial particles of the prefrontal cortex and hippocampus. Both acute and chronic amphetamine treatment increased locomotor behavior. Chronic amphetamine exposure induced a 3- to 6-fold increase of TBARS and a 1.5- to 2-fold increase of superoxide production in submitochondrial particles of prefrontal cortex and hippocampus ( P < 0.05). No effects on superoxide or TBARS were observed with acute treatment. These findings suggest that amphetamine-induced mitochondrial ROS generation may be a useful model to investigate the hypothesis of altered brain energy metabolism associated with BD and SCZ. Further studies assessing the effects of mood stabilizers and antipsychotics in preventing mitochondrial oxidative stress are necessary.

Chronic nicotine exposure during adolescence differentially influences calcium‐binding proteins in rat anterior cingulate cortex

We have recently shown that chronic amphetamine exposure selectively up-regulates parvalbumin (PV) calcium-binding proteins in the anterior cingulate cortex (ACC). In this study, we evaluated the effects of chronic nicotine (NIC) exposure on PV, calbindin D28k (CB) and calretinin (CR) calcium-binding protein immunostaining in ACC GABAergic interneurons. Chronic NIC exposure for 3 weeks in adolescent rats, either via drinking water (the oral group) or by twice daily subcutaneous injections (the injection group), resulted in the expression of high levels of CR proteins in the ACC but not in the parietal cortex. Larger increases in the density of CR-immunoreactive (ir) neurons were noted in the NIC-injected rats at 0-day withdrawal (45% increase) compared with the oral group (26% increase). The larger increases in CR-ir neuron density in the NIC-injected rats were also reflected by prominent CR-ir processes across cortical layers. The density of PV-ir neurons was also increased (37%) at 0-day withdrawal but only in the oral NIC group and no changes in CB-ir neuron density were observed in either NIC group. Combined dual-immunofluorescence and confocal microscopy revealed that somatodendritic alpha4 nicotinic acetylcholine receptors colocalized with cortical neurons stained positively for CR, PV or CB. These results suggest that CR- and/or PV-ir-containing GABA interneurons may be involved in channeling the effects of NIC in the ACC, which is closely associated with the ventral basal ganglia circuit that is linked to brain reward function.

L-type Ca2+ channels mediate adaptation of extracellular signal-regulated kinase 1/2 phosphorylation in the ventral tegmental area after chronic amphetamine treatment.

L-type Ca2+ channels (LTCCs) play an important role in chronic psychostimulant-induced behaviors. However, the Ca2+ second messenger pathways activated by LTCCs after acute and recurrent psychostimulant administration that contribute to drug-induced molecular adaptations are poorly understood. Using a chronic amphetamine treatment paradigm in rats, we have examined the role of LTCCs in activating the mitogen-activated protein (MAP) kinase pathway in the ventral tegmental area (VTA), a primary target for the reinforcing properties of psychostimulants. Using immunoblot and immunohistochemical analyses, we find that in chronic saline-treated rats a challenge injection of amphetamine increases phosphorylation of MAP [extracellular signal-regulated kinase 1/2 (ERK1/2)] kinase in the VTA that is independent of LTCCs. However, in chronic amphetamine-treated rats there is no increase in amphetamine-mediated ERK1/2 phosphorylation unless LTCCs are blocked, in which case there is robust phosphorylation in VTA dopamine neurons. Examination of the expression of phosphatases reveals an increase in calcineurin [protein phosphatase 2B (PP2B)] and MAP kinase phosphatase-1 (MKP-1) in the VTA. Using in situ hybridization histochemistry and immunoblot analyses, we further examined the mRNA and protein expression of the LTCC subtypes Ca(v)1.2 and Ca(v)1.3 in VTA dopamine neurons in drug-naive animals and in rats after chronic amphetamine treatment. We found an increase in Ca(v)1.2 mRNA and protein levels, with no change in Ca(v)1.3. Together, our results suggest that one aspect of LTCC-induced changes in second messenger pathways after chronic amphetamine exposure involves activation of the MAP kinase phosphatase pathway by upregulation of Ca(v)1.2 in VTA dopaminergic neurons.

Cholecystokinin modulation of locomotor behavior in rats is sensitized by chronic amphetamine and chronic restraint stress exposure

DA release in the nucleus accumbens (NAcc) is a critical substrate mediating locomotor behavior. Cholecystokinin (CCK) is co-localized with dopamine (DA) in up to 90% of mesolimbic DA neurons. We have previously shown that while CCK A receptor antagonists generally do not affect locomotor behaviors, systemic administration of a CCK A receptor antagonist attenuates amphetamine (AMPH)-induced locomotion in animals previously treated chronically with AMPH, suggesting that chronic stimulant pretreatment may sensitize CCK systems. The present studies examined this issue by testing the effects of CCK A antagonists on AMPH- and novel environment-induced locomotor activity following two manipulations which are known to alter mesolimbic system function: Chronic AMPH administration and chronic restraint stress (RS). Additionally, CCK immunoreactivity in the mesolimbic system following these manipulations was examined using immunohistochemistry. Results indicated that intra-NAcc microinjections of the selective CCK A receptor antagonist PD-140548 attenuated AMPH-induced and novel environment-induced locomotion only in animals which had previously been exposed to chronic AMPH or chronic RS pretreatment. However, chronic AMPH and chronic RS did not produce detectable changes in the number of CCK-immunostained neurons in the ventral tegmental area (VTA) or substantia nigra (SN), or in CCK levels in any of the subregions of the NAcc. Together, these results suggest that the role of endogenous CCK in the modulation of locomotor behaviors is sensitized following chronic psychostimulant or chronic RS exposure. However, this sensitization does not appear to be accompanied by changes in the overall basal levels of CCK or in the number of CCK-positive cells within the mesoaccumbens system.

Cloning, Expression, and Regulation of a Glucocorticoid-Induced Receptor in Rat Brain: Effect of Repetitive Amphetamine

Behavioral sensitization to psychostimulants involves neuroadaptation of stress-responsive systems. We have identified and sequenced a glucocorticoid-induced receptor (GIR) cDNA from rat prefrontal cortex. The full-length GIR cDNA encodes a 422 amino acid protein belonging to G-protein-coupled receptor superfamily. Although the ligand for GIR is still unknown, the dendrogram construction indicates that GIR may belong to peptide receptor subfamily (e.g., substance P receptor), with more distant relationship to subfamilies of glycoprotein hormone receptors (e.g., thyrotropin receptor) and biogenic amine receptors (e.g., dopamine receptor). GIR shares 31-34% amino acid identity to the tachykinin receptors (substance P receptor, neurokinin A receptor, and neurokinin B receptor). GIR mRNA is expressed preferentially in brain, and its neuronal expression is relegated to limbic brain regions, particularly in forebrain. GIR transcript levels are increased significantly and persistently in prefrontal cortex for 7 d after discontinuation of chronic amphetamine exposure. The induction of GIR expression by amphetamine is associated with augmented behavioral activation. These findings suggest that modulation of GIR expression may be involved in behavioral sensitization, and GIR may play a role at the interface between stress and neuroadaptation to psychostimulants.

Chronic Amphetamine Exposure during the Preweanling Period Does Not Affect Avoidance Learning or Novelty-Seeking of Adult Rats

The purpose of the present study was to determine whether exposure to amphetamine during the preweanling period would impact the learning or reward processes of rats tested in adulthood. In three experiments we examined whether amphetamine treatment (0–10 mg/kg per day) on postnatal days 11–17 altered the subsequent performance of adult Sprague–Dawley rats on a step-down passive avoidance, active avoidance, or novelty-seeking task. There was no evidence that postnatal amphetamine exposure affected performance on any of these tasks. These results suggest that the long-term impact of pre- and postnatal psychostimulant exposure differs, because in utero stimulant treatment is known to produce learning deficits and decrease reinforcement efficacy of rats tested in adulthood.