Weeks Of Cocaine Self-administration Research Articles

Chronic oral methylphenidate plus fluoxetine treatment in adolescent rats increases cocaine self-administration

BackgroundDepression and attention deficit hyperactivity disorder are known to be comorbid. Treatment of these commonly coexisting diseases typically involves the combined prescription of methylphenidate (MP), a psychostimulant, and fluoxetine (FLX), a selective serotonin reuptake inhibitor (SSRI). MP and cocaine have similar mechanisms of action and this study examined the effects of chronic treatment of MP combined with FLX on cocaine consumption in rats. MethodsFour groups of rats received access to drinking solutions of water (control), MP (30/60 mg/kg/day), FLX (20 mg/kg/day), or the combination of MP (30/60 mg/kg/day) plus FLX (20 mg/kg/day), during 8 h per day for one month. Following these drug treatments, rats were allowed to self-administer cocaine for 14 days. ResultsOur results showed that, during the first week of cocaine self-administration, the MP-treated rats had significantly greater numbers of active lever presses (plus 127%) and increased consumption of cocaine compared to the control rats. In contrast, during week two of cocaine self-administration, the rats treated with the MP + FLX combination showed significantly more lever presses (plus 198%) and significantly greater cocaine consumption (plus 84%) compared to the water controls. ConclusionChronic oral treatment during adolescence with the combination of MP plus FLX resulted in increased cocaine use after 2 weeks of cocaine self-administration in rats. These novel findings suggest that the combined exposure to these two drugs chronically, during adolescence, may produce increased vulnerability towards cocaine abuse during young adulthood.

Hormonal milieu drives economic demand for cocaine in female rats.

There are substantial sex differences in drug abuse, and a key feature of cocaine addiction is pathologically high motivation for drug. We investigated the role of ovarian hormones on cocaine demand in female rats using a within-session threshold behavioral economics (BE) procedure, which allows us to compare motivation for drug across hormonal states and sex while controlling for differences in dose and intake. This approach quantifies demand elasticity (α) and free consumption (Q0, consumption at null effort) to determine motivation for cocaine. Overall, female rats showed greater motivation for cocaine compared to males. However, this difference was cycle phase-dependent - motivation for cocaine when females were in proestrus was lower compared to the same animals across cycle phases, and overall similar to that of males. Hormonal cycle phase accounted for 70% of the within-subject variance in demand elasticity, obscuring other individual differences in female demand. High serum progesterone (P4; e.g., in proestrus) predicted decreased cocaine motivation (high demand elasticity), whereas serum estradiol (E2) correlated to greater intake at null effort (Q0). However, individual differences were revealed across OVX females, who displayed a range of demand elasticity, as seen in males. E2 replacement in OVX females increased motivation for cocaine, whereas P4 replacement decreased motivation. We also found that as few as 4 weeks of cocaine self-administration accelerated estropause in female rats as young as 12 weeks old. By 13 weeks of self-administration, proestrus epochs were no longer observed, and cocaine demand was potentiated by persistent estrus in all females. Thus, P4 signaling is a key modulator of cocaine demand in females that may underlie previously observed sex differences in addiction phenotypes.

Leptin Protects Against the Development and Expression of Cocaine Addiction-Like Behavior in Heterogeneous Stock Rats.

Cocaine affects food intake, metabolism and bodyweight. It has been hypothesized that feeding hormones like leptin play a role in this process. Preclinical studies have shown a mutually inhibitory relationship between leptin and cocaine, with leptin also decreasing the rewarding effects of cocaine intake. But prior studies have used relatively small sample sizes and did not investigate individual differences in genetically heterogeneous populations. Here, we examined whether the role of individual differences in bodyweight and blood leptin level are associated with high or low vulnerability to addiction-like behaviors using data from 306 heterogeneous stock rats given extended access to intravenous self-administration of cocaine and 120 blood samples from 60 of these animals, that were stored in the Cocaine Biobank. Finally, we tested a separate cohort to evaluate the causal effect of exogenous leptin administration on cocaine seeking. Bodyweight was reduced due to cocaine self-administration in males during withdrawal and abstinence, but was increased in females during abstinence. However, bodyweight was not correlated with addiction-like behavior vulnerability. Blood leptin levels after ∼6 weeks of cocaine self-administration did not correlate with addiction-like behaviors, however, baseline blood leptin levels before any access to cocaine negatively predicted addiction-like behaviors 6 weeks later. Finally, leptin administration in a separate cohort of 59 animals reduced cocaine seeking in acute withdrawal and after 7 weeks of protracted abstinence. These results demonstrate that high blood leptin level before access to cocaine may be a protective factor against the development of cocaine addiction-like behavior and that exogenous leptin reduces the motivation to take and seek cocaine. On the other hand, these results also show that blood leptin level and bodyweight changes in current users are not relevant biomarkers for addiction-like behaviors.

Behavioral economics of cocaine self-administration in male and female rats.

There are sex differences in the development of cocaine addiction. For example, the time that it takes for women from initial use to addiction is significantly shorter than for men. Thus, understanding why females are more vulnerable to cocaine addiction will provide insights into sex differences in the mechanisms underlying cocaine addiction. This study aimed to determine how cocaine demand intensity and elasticity might differ between sexes. In addition, the impact of estrous cycle and cocaine intake on demand was investigated. Male and female rats were trained to self-administer 0.125 mg of cocaine intravenously under a chained schedule in daily 2-h sessions for 2 weeks, and then, the cocaine demand function was determined with a modified within-session threshold procedure. Following the test, the rats began to self-administer a higher dose of cocaine (0.25 mg) to increase the cocaine intake. The demand function was then similarly determined in the same rats after 2 weeks of cocaine self-administration of the higher dose. No sex differences were found in either demand intensity or elasticity. Neither did the level of cocaine intake have an impact on demand. The demand elasticity, but not intensity, was significantly lower during proestrus/estrus compared with diestrus. These data suggest that the faster transition to cocaine addiction in women cannot be explained by sex differences in the demand for cocaine and such a demand may change during different phases of estrus cycle.

Biphasic effect of abstinence duration following cocaine self-administration on spine morphology and plasticity-related proteins in prelimbic cortical neurons projecting to the nucleus accumbens core.

Cocaine self-administration (SA) in rats dysregulates glutamatergic signaling in the prelimbic (PrL) cortex and glutamate release in the nucleus accumbens (NA) core, promoting cocaine seeking. PrL adaptations that affect relapse to drug seeking emerge during the first week of abstinence, switching from an early (2h) hypoglutamatergic state to a later (7 days) hyperglutamatergic state. Different interventions that normalize glutamatergic signaling in PrL cortex at each timepoint are necessary to suppress relapse. We hypothesized that plasticity-related proteins that regulate glutamatergic neurotransmission as well as dendritic spine morphology would be biphasically regulated during these two phases of abstinence in PrL cortical neurons projecting to the NA core (PrL-NA core). A combinatorial viral approach was used to selectively label PrL-NA core neurons with an mCherry fluorescent reporter. Male rats underwent 2weeks of cocaine SA or received yoked-saline infusions and were perfused either 2h or 7 days after the final SA session. Confocal microscopy and 3D reconstruction analyses were performed for Fos and pCREB immunoreactivity (IR) in the nucleus of layer V PrL-NA core neurons and GluA1-IR and GluA2-IR in apical dendritic spines of the same neurons. Here, we show that cocaine SA decreased PrL-NA core spine head diameter, nuclear Fos-IR and pCREB-IR, and GluA1-IR and GluA2-IR in putative mushroom-type spines 2h after the end of cocaine SA, whereas the opposite occurred following 1week of abstinence. Our findings reveal biphasic, abstinence duration-dependent alterations in structural plasticity and relapse-related proteins in the PrL-NA core pathway after cocaine SA.

Inactivation of NMDA Receptors in the Ventral Tegmental Area during Cocaine Self-Administration Prevents GluA1 Upregulation but with Paradoxical Increases in Cocaine-Seeking Behavior.

Cocaine self-administration increases expression of GluA1 subunits in ventral tegmental area (VTA) dopamine neurons, which subsequently enhance the motivation for cocaine. This increase in GluA1 may be dependent on concomitant NMDA receptor (NMDAR) activation during self-administration, similar to cocaine-induced long-term potentiation in the VTA. In this study, we used viral-mediated expression of a dominant-negative GluN1 subunit (HSV-dnGluN1) in VTA neurons to study the effect of transient NMDAR inactivation on the GluA1 increases induced by chronic cocaine self-administration in male rats. We found that dnGluN1 expression in the VTA limited to the 3 weeks of cocaine self-administration prevents the subsequent increase in tissue GluA1 levels when compared with control infusions of HSV-LacZ. Surprisingly, dnGluN1 expression led to an enhancement in the motivation to self-administer cocaine as measured using a progressive ratio reinforcement schedule and to enhanced cocaine seeking measured in extinction/reinstatement tests following an extended 3 week withdrawal period. Despite blocking tissue GluA1 increases in cocaine self-administering animals, the HSV-dnGluN1 treatment resulted in increased membrane levels of GluA1 and GluN2B, along with markedly higher locomotor responses to intra-VTA infusions of AMPA, suggesting a paradoxical increase in VTA AMPA receptor responsiveness. Together, these data suggest that NMDARs mediate cocaine-induced increases in VTA GluA1 expression, but such transient NMDAR inactivation also leads to compensatory scaling of synaptic AMPA receptors that enhance the motivational for cocaine.SIGNIFICANCE STATEMENT Dopamine neurons in the ventral tegmental area (VTA) are critical substrates of drug rewards. Animal models indicate that chronic cocaine use enhances excitatory glutamatergic input to these neurons, making them more susceptible to environmental stimuli that trigger drug craving and relapse. We previously found that self-administration of cocaine increases AMPA glutamate receptors in the VTA, and this effect enhances motivation for cocaine. Here we report that the mechanism for this upregulation involves NMDA receptor activity during cocaine use. While interference with NMDA receptor function blocks AMPA receptor upregulation, it also produces a paradoxical enhancement in membrane AMPA receptor subunits, AMPA responsiveness, and the motivation for cocaine. Thus, pharmacotherapy targeting NMDA receptors may inadvertently produce substantial adverse consequences for cocaine addiction.

D1, but not D2, receptor blockade within the infralimbic and medial orbitofrontal cortex impairs cocaine seeking in a region-specific manner.

Evidence suggests that the infralimbic cortex (IL), a subregion of the ventromedial prefrontal cortex (vmPFC), suppresses cocaine-seeking behavior in a self-administration paradigm, whereas the more anterior vmPFC subregion, the medial orbitofrontal cortex (mOFC), has received very little attention in this regard. Despite the established dopaminergic innervation of the vmPFC, whether dopamine receptor blockade in each subregion alters the reinstatement of cocaine seeking is unclear. To address this issue, male Sprague-Dawley rats underwent 2 weeks of cocaine self-administration, followed by extinction training and reinstatement testing. Immediately prior to each reinstatement test, rats received microinjections of the D1 receptor antagonist SCH 23390, the D2 receptor antagonist sulpiride or their respective vehicles. D1 receptor blockade in the IL reduced cued reinstatement but had no effect on cocaine prime and cue + cocaine-prime reinstatement, whereas D2 receptor blockade in the IL had no effect on reinstatement. For the mOFC, however, D1 receptor blockade reduced cocaine seeking in all reinstatement types, whereas blocking D2 receptors in the mOFC had no effect on any form of cocaine seeking. These findings suggest different roles for D1 receptors in the IL versus the mOFC in regulating cocaine-seeking behavior. Moreover, even as previous work indicates that IL inactivation does not affect reinstatement but, rather, induces cocaine seeking during extinction, the present findings suggest that dopamine receptor activation in the IL is necessary for cocaine seeking under some circumstances.

Abstinence from cocaine-self-administration activates the nELAV/GAP -43 pathway in the hippocampus: A stress-related effect?

We previously demonstrated that nELAV/GAP-43 pathway is pivotal for learning and its hippocampal expression is up-regulated by acute stress following repeated cocaine administration. We therefore hypothesized that abstinence-induced stress may sustain nELAV/GAP-43 pathway during early abstinence following 2 weeks of cocaine self-administration. We found that contingent, but not non-contingent, cocaine exposure selectively increases hippocampal nELAV, but not GAP-43, expression immediately after the last self-administration session, an effect that wanes after 24 h and that comes back 7 days later when nELAV activation becomes associated with increased expression of GAP-43, an effect again observed only in animals self-administering the psychostimulant. Such effect is specific for nELAV since the ubiquitous ELAV/HuR is unchanged. This nELAV profile suggests that its initial transient alteration is perhaps related to the daily administration of cocaine, while the increase in the nELAV/GAP-43 pathway following a week of abstinence may reflect the activation of this cascade as a target of stressful conditions associated with drug-related memories. © 2016 Wiley Periodicals, Inc.

Subchronic administration of atomoxetine causes an enduring reduction in context-induced relapse to cocaine seeking without affecting impulsive decision making.

Previous work has established a robust relationship between impulsivity and addiction, and revealed that impulsive decision making predisposes the vulnerability to cocaine-seeking behavior in rats. An important next step is to assess whether elevated relapse vulnerability can be treated via the reduction of impulsive decision making. Therefore, this study explored whether subchronic atomoxetine treatment can reduce relapse vulnerability by reducing impulsive decision making. Rats were trained in the delayed reward task and were subjected to 3 weeks of cocaine self-administration. Following drug self-administration, animals were divided to different experimental groups and received the noradrenaline transporter inhibitor and attention-deficit/hyperactivity disorder drug atomoxetine or vehicle subchronically for 20 days. On days 1 and 10 after treatment cessation, a context-induced reinstatement test was performed. Throughout the entire experiment, changes in impulsive decision making were continuously monitored. Subchronic treatment with atomoxetine reduced context-induced reinstatement both 1 and 10 days after treatment cessation, only in animals receiving no extinction training. Interestingly, neither subchronic nor acute atomoxetine treatments affected impulsive decision making. Our data indicate that the enduring reduction in relapse sensitivity by atomoxetine occurred independent of a reduction in impulsive decision making. Nonetheless, repeated atomoxetine administration seems a promising pharmacotherapeutical strategy to prevent relapse to cocaine seeking in abstinent drug-dependent subjects.

The role of ventral and dorsal striatum mGluR5 in relapse to cocaine-seeking and extinction learning

Cocaine addiction is a chronic, relapsing disease characterized by an inability to regulate drug-seeking behavior. Here we investigated the role of mGluR5 in the ventral and dorsal striatum in regulating cocaine-seeking following both abstinence and extinction. Animals underwent 2 weeks of cocaine self-administration followed by 3 weeks of home-cage abstinence. Animals were then reintroduced to the operant chamber for a context-induced relapse test, followed by 7-10 days of extinction training. Once responding was extinguished, cue-primed reinstatement test was conducted. Both drug-seeking tests were conducted in the presence of either mGluR5 negative allosteric modulator, MTEP or vehicle infused into either the nucleus accumbens (NA) core or dorsolateral striatum (dSTR). We found that MTEP infused in the NA core attenuated both context-induced relapse following abstinence and cue-primed reinstatement following extinction training. Blocking dSTR mGluR5 had no effect on context- or cue-induced cocaine-seeking. However, the intra-dSTR MTEP infusion on the context-induced relapse test day attenuated extinction learning for 4 days after the infusion. Furthermore, mGluR5 surface expression was reduced and LTD was absent in dSTR slices of animals undergoing 3 weeks of abstinence from cocaine but not sucrose self-administration. LTD was restored by bath application of VU-29, a positive allosteric modulator of mGluR5. Bath application of MTEP prevented the induction of LTD in dSTR slices from sucrose animals. Taken together, this data indicates that dSTR mGluR5 plays an essential role in extinction learning but not cocaine relapse, while NA core mGluR5 modulates drug-seeking following both extinction and abstinence from cocaine self-administration.

Cocaine potentiates excitatory drive in the perifornical/lateral hypothalamus

The hypothalamus is a critical controller of homeostatic responses and plays a fundamental role in reward-seeking behaviour. Recently, hypothalamic neurones in the perifornical/lateral hypothalamic area (PF/LHA) have also been implicated in drug-seeking behaviour through projections to extra-hypothalamic sites such as the ventral tegmental area. For example, a population of neurones that expresses the peptide orexin has been strongly implicated in addiction-relevant behaviours. To date, the effect of addictive drugs on synaptic properties in the hypothalamus remains largely unexplored. Previous studies focusing on the PF/LHA neurones, however, have shown that the orexin system exhibits significant plasticity in response to food or sleep restriction. This neuroadaptive ability suggests that PF/LHA neurones could be highly susceptible to modifications by drug exposure. Here, we sought to determine whether cocaine produces synaptic plasticity in PF/LHA neurones. Whole-cell patch-clamp techniques were used to examine the effects of experimenter-administered (passive) or self-administered (SA) cocaine on glutamatergic synaptic transmission in PF/LHA neurones. These experiments demonstrate that both passive and SA cocaine exposure increases miniature excitatory postsynaptic current (mEPSC) frequency in PF/LHA neurones. In addition, SA cocaine reduced the paired-pulse ratio but the AMPA/NMDA ratio of evoked excitatory inputs was unchanged, indicative of a presynaptic locus for synaptic plasticity. Dual-labelling for orexin and excitatory inputs using the vesicular glutamate transporter (VGLUT2), showed that passive cocaine exposure increased VGLUT2-positive appositions onto orexin neurones. Further, a population of recorded neurones that were filled with neurobiotin and immunolabelled for orexin confirmed that increased excitatory drive occurs in this PF/LHA population. Given the importance of the PF/LHA and the orexin system in modulating drug addiction, we suggest that these cocaine-induced excitatory synapse-remodelling events within the hypothalamus may contribute to persistence in drug-seeking behaviour and relapse.

Changes in the level of calcyon mRNA in the brain of rats exposed to cocaine, self-administered or received passively

The level of mRNA encoding calcyon (measured by in situ hybridization), one of the dopamine receptor interacting proteins, has been examined in the rat brain in the established animal model used to study the mechanisms of cocaine addiction (cocaine self-administration involving a yoked procedure). Two weeks of cocaine self-administration (maintenance) did not affect the level of calcyon mRNA, regardless of the way cocaine was delivered, except for tuberculum olfactorium, where calcyon mRNA was increased after cocaine treatment. In the reinstatement phase of the experiment cocaine alone induced an increase in the calcyon mRNA expression in most of the brain region studied (caudate putamen; tuberculum olfactorium; paraventricular thalamic nucleus; ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus) but only in the yoked saline control group. In other words, these results show that the single dose of cocaine (10 mg/kg) was able to induce an alteration in the level of calcyon mRNA in these rats which never before experienced any cocaine administration. The most significant effects were observed in the ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Interestingly, a similar effect was observed when the reinstatement of cocaine-seeking behaviour was evoked by cue (conditioned stimuli) that indicates that no cocaine was necessary to induce the changes in the level of calcyon mRNA expression. This effect was significant in tuberculum olfactorium, ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Such a result together with the brain areas involved in these effects might suggest the role of calcyon similar to the CART peptides and special vulnerability of calcyon expression rather to acute than chronic stimuli.

Tropomyosin-Related Kinase B in the Mesolimbic Dopamine System: Region-Specific Effects on Cocaine Reward

Previous studies found that brain-derived neurotrophic factor (BDNF) derived from nucleus accumbens (NAc) neurons can mediate persistent behavioral changes that contribute to cocaine addiction. To further investigate BDNF signaling in the mesolimbic dopamine system, we analyzed tropomyosin-related kinase B (TrkB) messenger RNA (mRNA) and protein changes in the NAc and ventral tegmental area (VTA) in rats following 3 weeks of cocaine self-administration. To study the role of BDNF-TrkB activity in the VTA and NAc in cocaine reward, we used localized viral-mediated Cre recombinase expression in floxed BDNF and floxed TrkB mice to knockdown BDNF or TrkB in the VTA and NAc in cocaine place conditioning tests and TrkB in the NAc in cocaine self-administration tests. We found that 3 weeks of active cocaine self-administration significantly increased TrkB protein levels in the NAc shell, while yoked (passive) cocaine exposure produced a similar increase in the VTA. Localized BDNF knockdown in either region reduced cocaine reward in place conditioning, whereas only TrkB knockdown in the NAc reduced cocaine reward. In mice self-administering cocaine, TrkB knockdown in the NAc produced a downward shift in the cocaine self-administration dose-response curve but had no effect on the acquisition of cocaine or sucrose self-administration. Together, these data suggest that BDNF synthesized in either VTA or NAc neurons is important for maintaining sensitivity to cocaine reward but only BDNF activation of TrkB receptors in the NAc mediates this effect. In addition, up-regulation of NAc TrkB with chronic cocaine use could promote the transition to more addicted biological states.

Repeated Cocaine Self-Administration Alters Processing of Cocaine-Related Information in Rat Prefrontal Cortex

One of the core symptoms of cocaine addiction is compulsive drug-seeking behavior. Although the precise neural substrates are unknown, it has been hypothesized that this behavior involves cocaine-induced hypofunction of the prefrontal cortex (PFC) or "hypofrontality." To test this hypothesis, PFC neuronal activity was monitored in rats during approximately 3 weeks of cocaine self-administration (SA). Rats were trained to press a lever to self-administer cocaine in daily 2 h sessions. Responding was reinforced contingent on a modified fixed-ratio 5 schedule of reinforcement. In the first SA session, the overall firing rate and burst rate were significantly decreased after cocaine infusions relative to the period immediately before the session. These effects disappeared after > or = 10 d of drug SA and were replaced by a significant increase in burst duration and firing rate within a burst. Notably, however, the level of basal activity before the first drug infusion of each SA session decreased significantly after multiple weeks of cocaine exposure. Collectively, these data support the view that although repeated sessions of cocaine SA decrease basal PFC activity, increased burst-related firing in response to cocaine infusions suggests that processing of cocaine-related information is enhanced and may contribute to increased control by cocaine over cocaine-seeking behavior.

The role of baclofen during extinction of cocaine self‐administration

Recent studies have focused on the neurobiological mechanism of relapse which plays a critical role in motivational aspect of compulsive drug-taking behavior and intense drug craving. In this respect, extinction procedure provides a measure of the development of motivational properties of drug seeking during the abstinence of drugs. Previous studies using a variety of cocaine self-administration procedures have shown that the GABA(B) agonist, baclofen, attenuates drug-seeking and drug-taking behavior. However, little is known about the role of baclofen on cocaine seeking behavior during extinction phase without the priming injection of cocaine. The purpose of present study was, therefore, to investigate the effect of baclofen on cocaine seeking behavior after cessation of continuing daily cocaine self-administration under a fixed ratio1 (FR1) schedule of reinforcement. Male Sprague-Dawley rats were trained to self-administer cocaine (1 mg/kg, i.v.) during daily 2-h sessions. When stable response was established by rats after 3 weeks of cocaine self-administration, cocaine was replaced by saline. On the first day of extinction, rats were administrated with baclofen (1.25 or 2.5 mg/kg, i.p.) or saline (1 ml/kg, i.p.) 30 min before the extinction session. Our results showed that pretreatment with 2.5 mg/kg baclofen, but not 1.25 mg/kg, significantly suppressed enhanced non-reinforced responding during extinction session. These results suggest that baclofen may be a potentially effective against cocaine addiction by preventing relapse in the abstinence of cocaine. Supported by R12-2003-002-03001-0

Dissociation of Primary and Secondary Reward-Relevant Limbic Nuclei in an Animal Model of Relapse

The neural substrates underlying relapse to drug-seeking behavior after chronic drug abuse may differ from those underlying immediate drug-taking behavior. In a model of relapse to drug-seeking behavior following chronic cocaine self-administration and prolonged extinction, we have previously shown that rats will significantly reinstate lever responding for either primary reward (cocaine) or secondary reward (tone + light stimulus previously paired with cocaine). In the present study, we utilized reversible inactivation of discrete brain nuclei with tetrodotoxin (TTX) in order to examine the neural substrates mediating primary and secondary cocaine reward in rats allowed two weeks of cocaine self-administration. After one week of daily extinction sessions, bilateral inactivation of the basolateral amygdala resulted in significant attenuation of lever pressing for a cocaine-conditioned reward (tone + light). Following three more days of extinction, bilateral TTX inactivation of the basolateral amygdala had no effect on the reinstatement of cocaine self-administration. In contrast, TTX inactivation of the nucleus accumbens produced the exact opposite effects, with significant blockade of primary reward (cocaine alone), but not secondary reward (tone + light). Thus, cocaine-conditioned reward is neuroanatomically dissociated from primary cocaine reward.

Elevated d2 receptor density in 23 schizophrenics

The level of mRNA encoding calcyon (measured by in situ hybridization), one of the dopamine receptor interacting proteins, has been examined in the rat brain in the established animal model used to study the mechanisms of cocaine addiction (cocaine self-administration involving a yoked procedure). Two weeks of cocaine self-administration (maintenance) did not affect the level of calcyon mRNA, regardless of the way cocaine was delivered, except for tuberculum olfactorium, where calcyon mRNA was increased after cocaine treatment. In the reinstatement phase of the experiment cocaine alone induced an increase in the calcyon mRNA expression in most of the brain region studied (caudate putamen; tuberculum olfactorium; paraventricular thalamic nucleus; ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus) but only in the yoked saline control group. In other words, these results show that the single dose of cocaine (10 mg/kg) was able to induce an alteration in the level of calcyon mRNA in these rats which never before experienced any cocaine administration. The most significant effects were observed in the ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Interestingly, a similar effect was observed when the reinstatement of cocaine-seeking behaviour was evoked by cue (conditioned stimuli) that indicates that no cocaine was necessary to induce the changes in the level of calcyon mRNA expression. This effect was significant in tuberculum olfactorium, ventromedial hypothalamic nucleus and paraventricular hypothalamic nucleus. Such a result together with the brain areas involved in these effects might suggest the role of calcyon similar to the CART peptides and special vulnerability of calcyon expression rather to acute than chronic stimuli.