Repeated Cocaine Treatment Research Articles

A Role for Kalirin in the Response of Rat Medium Spiny Neurons to Cocaine

Kalirin-7 (Kal7), the major kalirin isoform in adult brain, plays a key role in the formation of dendritic spines in hippocampal/cortical neurons. Its role in the GABAergic medium spiny neurons (MSNs) of the nucleus accumbens (NAc) and striatum, the areas known to play a key role in the common reward pathway, is not as well understood. Although Kal7 expression in mouse NAc increased in response to cocaine, MSN dendritic spine density did not differ from that for the wild type in Kal7-null mice. Unlike wild-type mice, Kal7-null mice did not respond to cocaine with an increase in MSN dendritic spine density. To explore further the role of Kal7 in cocaine-induced alterations in MSN morphology, we turned to the rat. Based on immunostaining, both Kal7 and Kal12 are expressed at moderate levels in the MSNs of the NAc and striatum. Expression of Kal7 and Kal12 in MSNs of both areas increases after repeated cocaine treatments. Overexpression of Kal7 in cultured MSN neurons increases dendritic spine density, as observed in rats after long-term cocaine administration. Reducing endogenous expression of all major kalirin isoforms in cultured MSN neurons causes a decrease in total dendritic length and dendritic spine density. These data suggest that kalirin is essential for maintaining spine density in NAc MSNs under normal conditions and that Kal7 is an obligatory intermediate in the response of MSNs to repeated exposure to cocaine.

1H NMR-based metabonomics in brain nucleus accumbens and striatum following repeated cocaine treatment in rats

Studies have shown a few cerebral metabolites modified by cocaine in brain regions; however, endogenous metabolic profiling has been lacking. Ex vivo 1H NMR (hydrogen-1 nuclear magnetic resonance) spectroscopy-based metabonomic approach coupled with partial least squares was applied to investigate the changes of cerebral metabolites in nucleus accumbens (NAc) and striatum of rats subjected to cocaine treatment. Our results showed that both single and repeated cocaine treatment can induce significant changes in a couple of cerebral metabolites. The increase of neurotransmitters glutamate and gamma-amino butyric acid (GABA) were observed in NAc and striatum from the rats repeatedly treated with cocaine. Creatine and taurine increased in NAc whereas taurine increased and creatine decreased in striatum after repeated cocaine treatment. Elevation of N-acetylaspartate in NAc and striatum and decrease of lactate in striatum were observed, which may reflect the mitochondria dysregulation caused by cocaine; moreover, alterations of choline, phosphocholine and glycerol in NAc and striatum could be related to membrane disruption. Moreover, groups of rats with and without conditioned place preference (CPP) apparatus are presenting difference in metabolites. Collectively, our results provide the first evidence of metabonomic profiling of NAc and striatum in response to cocaine, exhibiting a regionally-specific alteration patterns. We find that repeated cocaine administration leads to significant metabolite alterations, which are involved in neurotransmitter disturbance, oxidative stress, mitochondria dysregulation and membrane disruption in brain.

Cocaine-induced dendritic remodeling occurs in both D1 and D2 dopamine receptor-expressing neurons in the nucleus accumbens

Repeated exposure to cocaine can induce persistent alterations in the brain's reward system, including increases in the number of dendrites and spine density on medium-sized spiny neurons (MSNs) in the nucleus accumbens (NAc). The structural remodeling of dendrites and spines in the NAc is thought to play a critical role in cocaine addiction. MSNs in the NAc can be classified by expression of either D1 or D2 dopamine receptors, which are localized to the direct and indirect pathway, respectively. It is unknown whether the dendritic changes induced by repeated cocaine treatment occur in MSNs of the direct or indirect pathway. Because the traditional Golgi-Cox impregnation of neurons precludes identifying particular subpopulations of MSNs, we performed dendritic morphology analysis after biocytin-labeling and Golgi-Cox impregnation. We found that the biocytin staining MSNs showed higher dendritic spine density and higher number of dendrites than that in Golgi impregnation group. In addition, we found that the increasing spine density induced by repeated cocaine treatment in female mice was higher than that in male mice. Next we used biocytin staining and dynorphin/D2 receptor colocalization to determine which cell type(s) displayed dendritic changes after repeated cocaine treatment. We found that cocaine-induced changes in dendritic parameters occurred in MSNs of both the direct (D1-expressing) and indirect (D2-expressing) pathways.

Repeated cocaine treatments induce distinct locomotor effects in Crayfish

A repeated injection of cocaine regime is known to induce complex locomotion alterations in both vertebrate and invertebrate models of drug addiction. However, the specific effect of cocaine on behaviorally distinct locomotion and non locomotion parameters is not well known. The present experiments determined whether cocaine has distinct effect on multifarious locomotor activity of crayfish (Orconectes rusticus). Following repeated injections of 2.5 μg/g or 10.0 μg/g dose of cocaine for three days, videotaped recordings of locomotion were analyzed to determine whether repeated injections of cocaine produced distinct effect on multifarious locomotor activity of crayfish. Cocaine decreased immobility in day 1 when compared with saline. Thereafter, cocaine increased immobility in days 2 and 3. Repeated injections of cocaine increased distance traveled, average speed, mobility and decreased lingering episodes. These findings indicate that cocaine has distinct action on movement and non-movement behavioral activities, suggesting that locomotion as a unitary phenomenon comprised of assemblage of multifarious components, which can be manipulated and separated by cocaine in crayfish.

DNA Methylation Regulates Cocaine-Induced Behavioral Sensitization in Mice

The behavioral sensitization produced by repeated cocaine treatment represents the neural adaptations underlying some of the features of addiction in humans. Cocaine administrations induce neural adaptations through regulation of gene expression. Several studies suggest that epigenetic modifications, including DNA methylation, are the critical regulators of gene expression in the adult central nervous system. DNA methylation is catalyzed by DNA methyltransferases (DNMTs) and consequent promoter region hypermethylation is associated with transcriptional silencing. In this study a potential role for DNA methylation in a cocaine-induced behavioral sensitization model in mice was explored. We report that acute cocaine treatment caused an upregulation of DNMT3A and DNMT3B gene expression in the nucleus accumbens (NAc). Using methylated DNA immunoprecipitation, DNA bisulfite modification, and chromatin immunoprecipitation assays, we observed that cocaine treatment resulted in DNA hypermethylation and increased binding of methyl CpG binding protein 2 (MeCP2) at the protein phosphatase-1 catalytic subunit (PP1c) promoter. These changes are associated with transcriptional downregulation of PP1c in NAc. In contrast, acute and repeated cocaine administrations induced hypomethylation and decreased binding of MeCP2 at the fosB promoter, and these are associated with transcriptional upregulation of fosB in NAc. We also found that pharmacological inhibition of DNMT by zebularine treatment decreased cocaine-induced DNA hypermethylation at the PP1c promoter and attenuated PP1c mRNA downregulation in NAc. Finally, zebularine and cocaine co-treatment delayed the development of cocaine-induced behavioral sensitization. Together, these results suggest that dynamic changes of DNA methylation may be an important gene regulation mechanism underlying cocaine-induced behavioral sensitization.

Altered Dopamine Modulation of Inhibition in the Prefrontal Cortex of Cocaine-Sensitized Rats

A functionally hypoactive prefrontal cortex (PFC) is thought to contribute to decreased cognitive inhibitory control over drug-seeking behavior in cocaine addicts. Alterations in PFC dopamine (DA) and γ-aminobutyric acid (GABA) transmission are involved in the development of behavioral sensitization to cocaine, and repeated exposure to cocaine decreases DA D2 receptor (D2R) function in the PFC. We used recordings in PFC slices from adult rats to investigate how repeated cocaine treatment followed by 2 weeks of withdrawal affects DA modulation of GABA transmission and interneuron firing. In agreement with previous results in drug-naïve animals we found that in saline-treated control animals DA (20 μM) modulated evoked inhibitory post-synaptic currents (eIPSCs) in a biphasic, time- and receptor-dependent manner. Activation of D2Rs transiently reduced, whereas D1 receptor activation persistently increased the amplitude of eIPSCs. In cocaine-sensitized animals the D2R-dependent modulation of eIPSCs was abolished and the time course of DA effects was altered. In both saline- and cocaine-treated animals the effects of DA on eIPSCs were paralleled by distinct changes in spontaneous IPSCs (sIPSCs). In cocaine-treated animals the alterations in DA modulation of eIPSCs and sIPSCs correlated with a lack of D2R-specific reduction in action potential-independent GABA release, which might normally oppose D1-dependent increases in GABA transmission. Recordings from interneurons furthermore show that D2R activation can increase current-evoked spike firing in saline, but not in cocaine-treated animals. Altered DA regulation of inhibition during cocaine withdrawal could disturb normal cortical processing and contribute to a hypoactive PFC.

Cocaine administration increases CD4/CD8 lymphocyte ratio in peripheral blood despite lymphopenia and elevated corticosterone

The CD4/CD8 lymphocyte ratio in peripheral blood is used in the diagnosis of HIV infection, autoimmune disorders or susceptibility to infections. The present experiment aimed to evaluate the lymphocyte subsets, their distribution and CD4/CD8 ratio in blood after repeated, intravenous administration of cocaine. Adult male Wistar rats received three daily, in 30 min intervals, intravenous infusions of cocaine hydrochloride (5 mg/kg) or saline for 14 consecutive days. After each infusion the locomotor-activating effects of cocaine were assessed. Blood samples were collected 30 min after the last daily infusion on the 1st, 7th and 14th day of treatment. Total leukocyte numbers, percentages of leukocyte subpopulations, and T, B, NK, T CD4 +, and T CD8 + lymphocyte subsets, IFN-γ, and plasma corticosterone were determined. Repeated cocaine treatment resulted in an increase in neutrophil numbers and a significant decrease in total leukocyte and lymphocyte numbers involving a significant reduction in numbers of T, B, and NK lymphocyte subsets. T CD4 + and T CD8 + lymphocyte numbers were reduced but with a considerably smaller decrease in T CD4 + number. Cocaine treatment altered proportions between the lymphocyte subsets by decreasing the percentages of T CD8 +, B, and NK cells but increasing a percentage of T CD4 + cells. Destabilization in proportions between T CD4 + and T CD8 + was manifested as an elevated CD4 /CD8 ratio that occurred despite increased plasma corticosterone and the lymphocytopenia. Cocaine did not affect the concentration of IFN-γ. The results suggest that although cocaine induced lymphopenia, it did not suppress the overall immune activity in terms of the CD4 /CD8 ratio.

Dietary restriction mitigates cocaine‐induced alterations of olfactory bulb cellular plasticity and gene expression, and behavior

Because the olfactory system plays a major role in food consumption, and because 'food addiction' and associated morbidities have reached epidemic proportions, we tested the hypothesis that dietary energy restriction can modify adverse effects of cocaine on behavior and olfactory cellular and molecular plasticity. Mice maintained on an alternate day fasting (ADF) diet exhibited increased baseline locomotion and increased cocaine-sensitized locomotion during cocaine conditioning, despite no change in cocaine conditioned place preference, compared with mice fed ad libitum. Levels of dopamine and its metabolites in the olfactory bulb (OB) were suppressed in mice on the ADF diet compared with mice on the control diet, independent of acute or chronic cocaine treatment. The expression of several enzymes involved in dopamine metabolism including tyrosine hydroxylase, monoamine oxidases A and B, and catechol-O-methyltransferase were significantly reduced in OBs of mice on the ADF diet. Both acute and chronic administration of cocaine suppressed the production of new OB cells, and this effect of cocaine was attenuated in mice on the ADF diet. Cocaine administration to mice on the control diet resulted in up-regulation of OB genes involved in mitochondrial energy metabolism, synaptic plasticity, cellular stress responses, and calcium- and cAMP-mediated signaling, whereas multiple olfactory receptor genes were down-regulated by cocaine treatment. ADF abolished many of the effects of cocaine on OB gene expression. Our findings reveal that dietary energy intake modifies the neural substrates underlying some of the behavioral and physiological responses to repeated cocaine treatment, and also suggest novel roles for the olfactory system in addiction. The data further suggest that modification of dietary energy intake could provide a novel potential approach to addiction treatments.

Orphanin FQ/Nociceptin Not Only Blocks but Also Reverses Behavioral Adaptive Changes Induced by Repeated Cocaine in Mice

Orphanin FQ/nociceptin (OFQ/N), the endogenous ligand of the opioid receptor-like (ORL1) receptor, blocks cocaine sensitization in rats. In this study, we tested whether OFQ/N would block sensitization to the motor stimulatory and conditioned rewarding actions of cocaine in mice. We also examined whether OFQ/N, given to cocaine-sensitized mice, would reverse the sensitized response and whether it would prevent the amplified sensitized response induced by a second cocaine-sensitizing regimen in sensitized mice. ORL1 knockout and wild-type mice were treated with saline or OFQ/N before saline or cocaine on Days 1-3 and tested for sensitization on Day 8. Additionally, wild-type mice were treated similarly but tested for the conditioned rewarding action of cocaine, in which mice were tested for place preference before and after single conditioning with cocaine. Furthermore, mice were rendered sensitized, treated with saline or OFQ/N before saline or cocaine on Days 13-15, and received cocaine on Day 20 to test whether OFQ/N would reverse sensitization or block the amplified sensitized response induced by a second cocaine-sensitizing regimen in sensitized mice. OFQ/N blocked cocaine-induced psychomotor sensitization in wild-type but not knockout mice. It also blocked sensitization to the conditioned rewarding action of cocaine and reversed a preexisting locomotor sensitized response. Furthermore, OFQ/N prevented the amplified sensitized response that developed following a second cocaine sensitizing regimen given to sensitized mice. These results illustrate that OFQ/N not only blocks but also reverses maladaptive behavioral changes induced by repeated cocaine treatment in mice.

Identification of a Deubiquitinating Enzyme as a Novel AGS3-Interacting Protein

Activator of G protein Signaling 3 (AGS3) is a receptor-independent G protein activator that has been implicated in multiple biological events such as brain development, neuroplasticity and addiction, cardiac function, Golgi structure/function, macroautophagy and metabolism. However, how AGS3 is regulated is little known. We demonstrate here that AGS3 interacts with a ubiquitin specific protease USP9x, and this interaction is at least partially mediated through the C-terminal G protein regulatory domain of AGS3. Knockdown of USP9x causes a moderate reduction in the level of AGS3. In contrast, overexpression of either USP9x or its deubiquitinating domain UCH increases the amount of AGS3, whereas expression of the mutant UCH domain that lacks deubiquitinating activity does not have the same effect. As previously observed in AGS3 knockdown cells, the localization of several marker proteins of the late Golgi compartments is disturbed in cells depleted of USP9x. Taken together, our study suggests that USP9x can modulate the level of a subpopulation of AGS3, and this modulation plays a role in regulating the structure of the late Golgi compartments. Finally, we have found that levels of AGS3 and USP9x are co-regulated in the prefrontal cortex of rats withdrawn from repeated cocaine treatment. In conjunction with the above data, this observation indicates a potential role of USP9X in the regulation of the AGS3 level during cocaine-induced neuroplasticity.

Dopamine transporter down‐regulation following repeated cocaine: implications for 3,4‐methylenedioxymethamphetamine‐induced acute effects and long‐term neurotoxicity in mice

3,4-Methylenedioxymethamphetamine (MDMA) and cocaine are two widely abused psychostimulant drugs targeting the dopamine transporter (DAT). DAT availability regulates dopamine neurotransmission and uptake of MDMA-derived neurotoxic metabolites. We aimed to determine the effect of cocaine pre-exposure on the acute and long-term effects of MDMA in mice. Mice received a course of cocaine (20 mg*kg(-1), x2 for 3 days) followed by MDMA (20 mg*kg(-1), x2, 3 h apart). Locomotor activity, extracellular dopamine levels and dopaminergic neurotoxicity were determined. Furthermore, following the course of cocaine, DAT density in striatal plasma membrane and endosome fractions was measured. Four days after the course of cocaine, challenge with MDMA attenuated the MDMA-induced striatal dopaminergic neurotoxicity. Co-administration of the protein kinase C (PKC) inhibitor NPC 15437 prevented cocaine protection. At the same time, after the course of cocaine, DAT density was reduced in the plasma membrane and increased in the endosome fraction, and this effect was prevented by NPC 15437. The course of cocaine potentiated the MDMA-induced increase in extracellular dopamine and locomotor activity, following challenge 4 days later, compared with those pretreated with saline. Repeated cocaine treatment followed by withdrawal protected against MDMA-induced dopaminergic neurotoxicity by internalizing DAT via a mechanism which may involve PKC. Furthermore, repeated cocaine followed by withdrawal induced behavioural and neurochemical sensitization to MDMA, measures which could be indicative of increased rewarding effects of MDMA.

Anxiety-like responses in adolescent rats following a 10–11-day withdrawal period from repeated cocaine administration

The present study was conducted to determine whether increases in anxiety-like behavior in adolescent rats repeatedly treated with cocaine would be observed following a withdrawal period of an intermediate length (i.e., more than 1 day but less than 2 weeks). Accordingly, rats were injected with either 10 or 20 mg/kg cocaine or with saline for 7–8 consecutive days followed by a 10–11-day drug-free recovery period at which time assessment in a zero maze was instituted. All animals were assessed in the zero maze twice with the two sessions being separated by 24 h of non-testing. The results indicated that all three groups spent equivalent amounts of time in the open portions of the maze on the initial test day. Group differences, however, emerged on the second test day with animals in both cocaine-treated groups spending significantly less time in the open portions of the maze relative to saline-injected control subjects. This was due to control animals increasing the amount of time spent in the open areas of the maze from the first to the second day of testing, an effect not observed in either of the two cocaine-treated groups. These data provide evidence that an anxiogenic-like response produced by repeated cocaine treatment in adolescent rats emerges after a relatively short withdrawal period.

Influence of progesterone on GAD65 and GAD67 mRNA expression in the dorsolateral striatum and prefrontal cortex of female rats repeatedly treated with cocaine

Female rats are intensely affected by cocaine, with estrogen probably playing an important role in this effect. Progesterone modulates the GABA system and attenuates the effects of cocaine; however, there is no information about its relevance in changing GABA synthesis pathways after cocaine administration to female rats. Our objective was to investigate the influence of progesterone on the effects of repeated cocaine administration on the isoenzymes of glutamic acid decarboxylase (GAD(65) and GAD(67)) mRNA in brain areas involved in the addiction circuitry. Ovariectomized, intact and progesterone replacement-treated female rats received saline or cocaine (30 mg/kg, ip) acutely or repeatedly. GAD isoenzyme mRNA levels were determined in the dorsolateral striatum (dSTR) and prefrontal cortex (PFC) by RT-PCR, showing that repeated, but not acute, cocaine decreased GADs/beta-actin mRNA ratio in the dSTR irrespective of the hormonal condition (GAD(65): P < 0.001; and GAD(67): P = 0.004). In the PFC, repeated cocaine decreased GAD(65) and increased GAD(67) mRNA ratio (P < 0.05). Progesterone replacement decreased both GAD isoenzymes mRNA ratio after acute cocaine in the PFC (P < 0.001) and repeated cocaine treatment reversed this decrease (P < 0.001). These results suggest that cocaine does not immediately affect GAD mRNA expression, while repeated cocaine decreases both GAD(65) and GAD(67) mRNA in the dSTR of female rats, independently of their hormonal conditions. In the PFC, repeated cocaine increases the expression of GAD isoenzymes, which were decreased due to progesterone replacement.

The involvement of type IV phosphodiesterases in cocaine-induced sensitization and subsequent pERK expression in the mouse nucleus accumbens

Cocaine exposure produces sensitization that is partly mediated by cyclic adenosine monophosphate (cAMP) pathways within the nucleus accumbens (NAc). Type IV phosphodiesterases (PDE4s) break down cAMP and are required for cocaine-induced conditioned place preference. Whether PDE4 disruption attenuates induction of behavioral sensitization to cocaine and subsequent NAc expression of phosphorylated extracellular signal-regulated kinase (ERK), which is involved in cocaine-induced sensitization, is unknown. The objective of this study was to determine whether inhibition of PDE4s prevents cocaine-induced locomotor sensitization and if reduced behavioral sensitization is accompanied by decreased expression of phosphorylated ERK (pERK) within the NAc. Mice were administered the PDE4 inhibitor, rolipram, or vehicle before or after five daily injections of cocaine or saline, and activity was monitored on days 1 and 5. After nine drug-free days, locomotor sensitization was tested. Some subjects were sacrificed following testing for behavioral sensitization to measure pERK expression in the NAc. PDE4 inhibition, during the induction of sensitization, reduced behavioral sensitization only if rolipram (1.0 mg/kg) was administered before cocaine. Re-exposure to the cocaine-paired environment following a 9-day drug-free period enhanced pERK expression in the NAc core and shell. Rolipram did not alter pERK induction despite blocking behavioral sensitization. Rolipram given during, but not following, cocaine treatment prevents development of locomotor sensitization to cocaine but does not affect subsequent pERK activation induced by exposure to a cocaine-paired context or following a cocaine challenge. Although PDE4 inhibition during the induction of sensitization blocks the locomotor component of sensitization, other long-term changes induced by repeated cocaine treatment remain.

Long‐lasting dysregulation of gene expression in corticostriatal circuits after repeated cocaine treatment in adult rats: effects on zif 268 and homer 1a

Human imaging studies show that psychostimulants such as cocaine produce functional changes in several areas of cortex and striatum. These may reflect neuronal changes related to addiction. We employed gene markers (zif 268 and homer 1a) that offer a high anatomical resolution to map cocaine-induced changes in 22 cortical areas and 23 functionally related striatal sectors, in order to determine the corticostriatal circuits altered by repeated cocaine exposure (25 mg/kg, 5 days). Effects were investigated 1 day and 21 days after repeated treatment to assess their longevity. Repeated cocaine treatment increased basal expression of zif 268 predominantly in sensorimotor areas of the cortex. This effect endured for 3 weeks in some areas. These changes were accompanied by attenuated gene induction by a cocaine challenge. In the insular cortex, the cocaine challenge produced a decrease in zif 268 expression after the 21-day, but not 1-day, withdrawal period. In the striatum, cocaine also affected mostly sensorimotor sectors. Repeated cocaine resulted in blunted inducibility of both zif 268 and homer 1a, changes that were still very robust 3 weeks later. Thus, our findings demonstrate that cocaine produces robust and long-lasting changes in gene regulation predominantly in sensorimotor corticostriatal circuits. These neuronal changes were associated with behavioral stereotypies, which are thought to reflect dysfunction in sensorimotor corticostriatal circuits. Future studies will have to elucidate the role of such neuronal changes in psychostimulant addiction.

Aquaporin 4 knockout resists negative regulation of neural cell proliferation by cocaine in mouse hippocampus

Our previous study revealed that aquaporin 4 (AQP4) knockout attenuated locomotor activity in cocaine exposure mice and reduced the extracellular dopamine levels in the nucleus accumbens, suggesting that AQP4 might participate in cocaine addiction. The aim of the present study was to investigate the impact of AQP4 on cell proliferation of dentate gyrus in the mouse hippocampus after repeated cocaine treatment and withdrawal. The immunohistochemistry results showed that repeated cocaine administration significantly decreased cellular proliferation in the subgranular zone, which was followed by a rebound increase after 2-wk withdrawal and a return to normal level after 3-wk withdrawal. AQP4 knockout resisted cocaine-induced reductions of neural cell proliferation. Further studies through immunohistochemistry and immunoblot analysis showed that AQP4 knockout sustained the levels of glial fibrillary acidic protein in the hippocampus, and suppressed the enhancement of extracellular signal-regulated kinase phosphorylation induced by repeated cocaine administration. Notably, AQP4 knockout increased protein kinase C activity examined by substrate protein phosphorylation method, which was not affected by cocaine administration or withdrawal. We also found that repeated cocaine administration could elevate the expression of AQP4 in wild-type mice. In conclusion, it is reported for the first time that AQP4 knockout resisted cocaine-mediated inhibition of neural cell proliferation via up-regulating PKC-mediated signal transduction, suggesting that AQP4 might regulate neurogenesis during drug addiction. Our findings may have helpful implications in the cell biology of neurogenesis.

Chronic cocaine enhances release of neuroprotective amino acid taurine: a microdialysis study

AbstractCocaine inhibits high-affinity neurotransmitter uptake at the presynaptic nerve terminals to increase synaptic levels of dopamine, norepinephrine and serotonin^1^. This increase of synaptic dopamine may cause neurotoxicity^2,3^. At least two different mechanisms have been proposed for the development of dopamine-related neurotoxicity: 1) dopamine produces a free radical that may induce cell toxicity^2,3^ and 2) dopamine reduces glutamate transport at its presynaptic sites to increase synaptic levels of this amino acid^4^ and augments glutamate transmission by activating dopamine D1 receptors in different areas of the brain^5-7^. Increase in glutamatergic transmission mediated by the activation on N-methyl dextro-aspartate (NMDA) receptors has been shown to cause excitotoxicity and neuro-degeneration^8^. Others and we have reported protection against different psychotropic drug-induced neurotoxicity that may be achieved by prior or simultaneous administration of various pharmacological agents. For example, repeated treatment of rats with haloperidol induced neuronal damage that is ameliorated by prior administration of either GM1 ganglioside^9^ or the endogenous amino acid, taurine^10^. Similarly, chronic gestational cocaine exposure causes neurotoxicity that could be prevented by co-administration of clozapine^11^. To our knowledge, there is no information if chronic cocaine would enhance release of endogenous protective agents that may oppose the over activation of glutamatergic system. Here we show that repeated cocaine treatment increased synaptic levels of the neuroprotective amino acid taurine that opposes the excessive excitatory actions of the glutamatergic system in the rat brain. Thus, mammalian brain has an auto-protective mechanism to counter excitotoxicity and taurine or its synthetic derivative may be useful in the management and treatment of cocaine addiction and its neurotoxic effect.

Cell Surface AMPA Receptors in the Rat Nucleus Accumbens Increase during Cocaine Withdrawal But Internalize after Cocaine Challenge in Association with Altered Activation of Mitogen-Activated Protein Kinases

Although some studies report increased responsiveness of nucleus accumbens (NAc) AMPA receptors (AMPARs) after withdrawal from repeated cocaine treatment, others report decreased responsiveness after withdrawal plus cocaine challenge. Here we examine this apparent contradiction by quantifying cell surface and intracellular AMPAR subunits in the NAc before and after a challenge injection in behaviorally sensitized rats. Because MAPKs (mitogen-activated protein kinases) regulate AMPAR trafficking and are implicated in addiction, we also evaluated phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38. Glutamate receptor 1 (GluR1) and GluR2 surface/intracellular (S/I) ratios were increased after 14 d of withdrawal in sensitized rats but were decreased 24 h after challenge with cocaine (which elicited a sensitized locomotor response) or saline (which elicited conditioned locomotion). These findings suggested redistribution of GluR1/2-containing receptors, a possibility supported by immunoprecipitation experiments indicating that most AMPARs in the NAc are GluR1/2 or GluR2/3, with few homomeric GluR1 or GluR1/3 receptors. In sensitized rats, ERK phosphorylation in the NAc increased during withdrawal and normalized after cocaine challenge. JNK phosphorylation also increased after withdrawal, but after cocaine challenge, it was inversely related to GluR1 and GluR2 S/I ratios. After saline challenge, p38 phosphorylation was increased. In summary, surface expression of GluR1/2-containing AMPARs increased in the NAc of sensitized rats, but AMPARs internalized after a single reexposure to cocaine or cocaine-related cues. ERK phosphorylation paralleled AMPAR surface expression. Although JNK results were complex, JNK and p38 may be involved in AMPAR internalization after cocaine or saline challenge, respectively.

Interactions among ovarian hormones and time of testing on behavioral sensitization and cocaine self-administration

Ovarian hormones play a role in the use of drugs of abuse in women. In female rats estradiol has been shown to enhance acquisition of cocaine self-administration and behavioral sensitization induced by repeated cocaine treatment. Experiments were conducted to determine the effects of estradiol and/or progesterone on cocaine self-administration and behavioral sensitization to cocaine (10 mg/kg; in animals with unilateral 6-hydroxydopamine lesions). Five groups of ovariectomized females were tested: (1) oil vehicle; (2) estradiol (E); (3) progesterone (P); (4) estradiol and progesterone given concurrently (EPC); (5) estradiol and progesterone given sequentially (EPS: 3 days of estradiol, 1 day progesterone, 1 day oil). All animals were tested during the dark phase of the light:dark cycle at ZT1600 and ZT2000–2100. Behavioral sensitization results: there was substantial conditioned turning throughout the habituation periods, and all animals exhibited behavioral sensitization with repeated cocaine treatment. Multivariate analysis indicated a significant effect of hormone treatment, time of day and day of testing. When individual groups were compared, however, only at ZT1600 did the E-treated and the EPS-treated animals show a trend ( p < 0.06) for greater behavioral sensitization to cocaine relative to the oil-treated animals. Self-administration results: all groups showed rapid acquisition of cocaine self-administration at 0.3 mg/kg/infusion, so we did not see an effect of ovarian hormones on acquisition, or a difference between groups tested at ZT1600 versus ZT2100 ( p < 0.05). There was, however, enhanced total intake of cocaine at 0.75 mg/kg/infusion in the E and the EPS groups. Concurrent administration of progesterone with estradiol counteracted the effect of estradiol on cocaine intake at 0.75 mg/kg/infusion, while progesterone alone did not enhance cocaine self-administration.

Long‐lasting up‐regulation of orexin receptor type 2 protein levels in the rat nucleus accumbens after chronic cocaine administration

Hypothalamic orexin (hypocretin) neurons project to the key structures of the limbic system and orexin receptors, both orexin receptor type 1 (OXR1) and type 2 (OXR2), are expressed in most limbic regions. Emerging evidence suggests that orexin is among important neurotransmitters that regulate addictive properties of drugs of abuse. In this study, we examined the effect of psychostimulant cocaine on orexin receptor protein abundance in the rat limbic system in vivo. Intermittent administration of cocaine (20 mg/kg, i.p., once daily for 5 days) caused a typical behavioral sensitization response to a challenge cocaine injection at a 14-day withdrawal period. Repeated cocaine administration at the same withdrawal time also increased OXR2 protein levels in the nucleus accumbens while repeated cocaine had no effect on OXR1 and orexin neuropeptide (both orexin-A and orexin-B) levels in this region. In contrast to the nucleus accumbens, OXR2 levels in the frontal cortex, the ventral tegmental area, the hippocampus, and the dorsal striatum (caudate putamen) were not altered by cocaine. Remarkably, the up-regulated OXR2 levels in the nucleus accumbens showed a long-lasting nature as it persisted up to 60 days after the discontinuation of repeated cocaine treatments. In contrast to chronic cocaine administration, an acute cocaine injection was insufficient to modify levels of any orexin receptor and peptide. Our data identify the up-regulation of OXR2 in the nucleus accumbens as an enduring molecular event that is correlated well with behavioral plasticity in response to chronic psychostimulant administration. This OXR2 up-regulation may reflect a key adaptation of limbic orexinergic transmission to chronic drug exposure and may thus be critical for the expression of motor plasticity.