USP7/Maged1-mediated H2A monoubiquitination in the paraventricular thalamus: an epigenetic mechanism involved in cocaine use disorder.

The risk of developing drug addiction is strongly influenced by the epigenetic landscape and chromatin remodeling. While histone modifications such as methylation and acetylation have been studied in the ventral tegmental area and nucleus accumbens (NAc), the role of H2A monoubiquitination remains unknown. Our investigations, initially focused on the scaffold protein melanoma-associated antigen D1 (Maged1), reveal that H2A monoubiquitination in the paraventricular thalamus (PVT) significantly contributes to cocaine-adaptive behaviors and transcriptional repression induced by cocaine. Chronic cocaine use increases H2A monoubiquitination, regulated by Maged1 and its partner USP7. Accordingly, Maged1 specific inactivation in thalamic Vglut2 neurons, or USP7 inhibition, blocks cocaine-evoked H2A monoubiquitination and cocaine locomotor sensitization. Additionally, genetic variations in MAGED1 and USP7 are linked to altered susceptibility to cocaine addiction and cocaine-associated symptoms in humans. These findings unveil an epigenetic modification in a non-canonical reward pathway of the brain and a potent marker of epigenetic risk factors for drug addiction in humans.

Activation of a hypothalamus-habenula circuit by mechanical stimulation inhibits cocaine addiction-like behaviors.

BACKGROUND: Mechanoreceptor activation modulates GABA neuron firing and dopamine (DA) release in the mesolimbic DA system, an area implicated in reward and substance abuse. The lateral habenula (LHb), the lateral hypothalamus (LH), and the mesolimbic DA system are not only reciprocally connected, but also involved in drug reward. We explored the effects of mechanical stimulation (MS) on cocaine addiction-like behaviors and the role of the LH-LHb circuit in the MS effects. MS was performed over ulnar nerve and the effects were evaluated by using drug seeking behaviors, optogenetics, chemogenetics, electrophysiology and immunohistochemistry. RESULTS: Mechanical stimulation attenuated locomotor activity in a nerve-dependent manner and 50-kHz ultrasonic vocalizations (USVs) and DA release in nucleus accumbens (NAc) following cocaine injection. The MS effects were ablated by electrolytic lesion or optogenetic inhibition of LHb. Optogenetic activation of LHb suppressed cocaine-enhanced 50 kHz USVs and locomotion. MS reversed cocaine suppression of neuronal activity of LHb. MS also inhibited cocaine-primed reinstatement of drug-seeking behavior, which was blocked by chemogenetic inhibition of an LH-LHb circuit. CONCLUSION: These findings suggest that peripheral mechanical stimulation activates LH-LHb pathways to attenuate cocaine-induced psychomotor responses and seeking behaviors.

The orexin (hypocretin) neuropeptide system is a target for novel therapeutics to treat cocaine use disorder with alcohol coabuse.

An estimated 50-90% of individuals with cocaine use disorder (CUD) also report using alcohol. Cocaine users report coabusing alcohol to 'self-medicate' against the negative emotional side effects of the cocaine 'crash', including the onset of anxiety. Thus, pharmaceutical strategies to treat CUD would ideally reduce the motivational properties of cocaine, alcohol, and their combination, as well as reduce the onset of anxiety during drug withdrawal. The hypothalamic orexin (hypocretin) neuropeptide system offers a promising target, as orexin neurons are critically involved in activating behavioral and physiological states to respond to both positive and negative motivators. Here, we seek to describe studies demonstrating efficacy of orexin receptor antagonists in reducing cocaine, alcohol- and stress-related behaviors, but note that these studies have largely focused on each of these phenomena in isolation. For orexin-based compounds to be viable in the clinical setting, we argue that it is imperative that their efficacy be tested in animal models that account for polysubstance use patterns. To begin to examine this, we present new data showing that rats' preferred level of cocaine intake is significantly increased following chronic homecage access to alcohol. We also report that cocaine intake and motivation are reduced by a selective orexin-1 receptor antagonist when rats have a history of cocaine + alcohol, but not a limited history of cocaine alone. In light of these proof-of-principle data, we outline what we believe to be the key priorities going forward with respect to further examining the orexin system in models of polysubstance use. This article is part of the special issue on Neurocircuitry Modulating Drug and Alcohol Abuse.

Electro-acupuncture alleviates adolescent cocaine exposure-enhanced anxiety-like behaviors in adult mice by attenuating the activities of PV interneurons in PrL.

We recently found that adolescent cocaine exposure (ACE) resulted in an enhancement of the γ-aminobutyric acid (GABA) neurotransmitter system in the prelimbic cortex (PrL) of adult mice. Here, we aim to further investigate the role of GABAergic transmission, especially parvalbumin (PV) interneurons within PrL in the development of ACE-induced anxiety-like behavior, and to assess whether and how electro-acupuncture (EA) therapeutically manage the ACE-induced abnormal behaviors in adulthood. ACE mice exhibited the enhanced anxiety-like behaviors in their adulthood, accompanied by increased GABAergic transmission and PV interneurons in PrL. Chemogenetic blocking PV interneurons in PrL alleviated ACE-enhanced anxiety-like behaviors in mice. Importantly, 37-day EA treatments (mixture of 2 Hz/100 Hz, 1 mA, 30 minutes once a day) at the acupoints of Yintang (GV29) and Baihui (GV20) also alleviated ACE-induced anxiety-like behaviors, and rescued ACE-impaired GABAergic neurotransmitter system and PV interneurons in PrL. In parallel, EA treatments further suppressed the activities of pyramidal neurons in PrL, suggesting that EA treatments seem to perform it beneficial effects on the ACE-induced abnormal emotional behaviors by "calming down" the whole PrL. Collectively, these findings revealed that hyper-function of GABAergic transmission, especially mediating by PV interneurons in PrL may be key etiology underlying ACE-induced anxiety-like behaviors. At least by normalizing the function of GABAergic and PV interneurons, EA may represent a promising therapeutic strategy for managing adolescent substance use-related emotional disorders.

Xie2-64, a novel CB2 receptor inverse agonist, reduces cocaine abuse-related behaviors in rodents.

Cocaine abuse remains a public health threat around the world. There are no pharmacological treatments approved for cocaine use disorder. Cannabis has received growing attention as a treatment for many conditions, including addiction. Most cannabis-based medication development has focused on cannabinoid CB1 receptor (CB1R) antagonists (and also inverse agonists) such as rimonabant, but clinical trials with rimonabant have failed due to its significant side-effects. Here we sought to determine whether a novel and selective CB2R inverse agonist, Xie2-64, has similar therapeutic potential for cocaine use disorder. Computational modeling indicated that Xie2-64 binds to CB2R in a way similar to SR144528, another well-characterized but less selective CB2R antagonist/inverse agonist, suggesting that Xie2-64 may also have CB2R antagonist profiles. Unexpectedly, systemic administration of Xie2-64 or SR144528 dose-dependently inhibited intravenous cocaine self-administration and shifted cocaine dose-response curves downward in rats and wild-type, but not in CB2R-knockout, mice. Xie2-64 also dose-dependently attenuated cocaine-enhanced brain-stimulation reward maintained by optical stimulation of ventral tegmental area dopamine (DA) neurons in DAT-Cre mice, while Xie2-64 or SR144528 alone inhibited optical brain-stimulation reward. In vivo microdialysis revealed that systemic or local administration of Xie2-64 into the nucleus accumbens reduced extracellular dopamine levels in a dose-dependent manner in rats. Together, these results suggest that Xie2-64 has significant anti-cocaine reward effects likely through a dopamine-dependent mechanism, and therefore, deserves further study as a new pharmacotherapy for cocaine use disorder.

Clinical Potential of an Enzyme-based Novel Therapy for Cocaine Overdose.

It is a grand challenge to develop a truly effective medication for treatment of cocaine overdose. The current available, practical emergence treatment for cocaine overdose includes administration of a benzodiazepine anticonvulsant agent (e.g. diazepam) and/or physical cooling with an aim to relieve the symptoms. The inherent difficulties of antagonizing physiological effects of drugs in the central nervous system have led to exploring protein-based pharmacokinetic approaches using biologics like vaccines, monoclonal antibodies, and enzymes. However, none of the pharmacokinetic agents has demonstrated convincing preclinical evidence of clinical potential for drug overdose treatment without a question mark on the timing used in the animal models. Here we report the use of animal models, including locomotor activity, protection, and rescue experiments in rats, of drug toxicity treatment with clinically relevant timing for the first time. It has been demonstrated that an efficient cocaine-metabolizing enzyme developed in our previous studies can rapidly reverse the cocaine toxicity whenever the enzyme is given to a living rat, demonstrating promising clinical potential of an enzyme-based novel therapy for cocaine overdose as a successful example in comparison with the commonly used diazepam.

Functional consequences of cocaine expectation: findings in a non-human primate model of cocaine self-administration.

Exposure to stimuli and environments associated with drug use is considered one of the most important contributors to relapse among substance abusers. Neuroimaging studies have identified neural circuits underlying these responses in cocaine-dependent subjects. But these studies are often difficult to interpret because of the heterogeneity of the participants, substances abused, and differences in drug histories and social variables. Therefore, the goal of this study was to assess the functional effects of exposure to cocaine-associated stimuli in a non-human primate model of cocaine self-administration, providing precise control over these variables, with the 2-[(14) C]deoxyglucose method. Rhesus monkeys self-administered 0.3 mg/kg/injection cocaine (n = 4) under a fixed-interval 3-minute (FI 3-min) schedule of reinforcement (30 injections/session) for 100 sessions. Control animals (n = 4) underwent identical schedules of food reinforcement. Sessions were then discontinued for 30 days, after which time, monkeys were exposed to cocaine- or food-paired cues, and the 2-[(14) C]deoxyglucose experiment was conducted. The presentation of the cocaine-paired cues resulted in significant increases in functional activity within highly restricted circuits that included portions of the pre-commissural striatum, medial prefrontal cortex, rostral temporal cortex and limbic thalamus when compared with control animals presented with the food-paired cues. The presentation of cocaine-associated cues increased brain functional activity in contrast to the decreases observed after cocaine consumption. Furthermore, the topography of brain circuits engaged by the expectation of cocaine is similar to the distribution of effects during the earliest phases of cocaine self-administration, prior to the onset of neuroadaptations that accompany chronic cocaine exposure.

Synthesis and characterization of a dual kappa-delta opioid receptor agonist analgesic blocking cocaine reward behavior.

3-Iodobenzoyl naltrexamine (IBNtxA) is a potent analgesic belonging to the pharmacologically diverse 6ß-amidoepoxymorphinan group of opioids. We present the synthesis and pharmacological evaluation of five analogs of IBNtxA. The scaffold of IBNtxA was modified by removing the 14-hydroxy group, incorporating a 7,8 double bond and various N-17 alkyl substituents. The structural modifications resulted in analogs with picomolar affinities for opioid receptors. The lead compound (MP1104) was found to exhibit approximately 15-fold greater antinociceptive potency (ED50 = 0.33 mg/kg) compared with morphine, mediated through the activation of kappa- and delta-opioid receptors. Despite its kappa agonism, this lead derivative did not cause place aversion or preference in mice in a place-conditioning assay, even at doses 3 times the analgesic ED50. However, pretreatment with the lead compound prevented the reward behavior associated with cocaine in a conditioned place preference assay. Together, these results suggest the promise of dual acting kappa- and delta-opioid receptor agonists as analgesics and treatments for cocaine addiction.

Persistent increases in rat hypothalamic POMC gene expression following chronic withdrawal from chronic "binge" pattern escalating-dose, but not steady-dose, cocaine.

Recent research suggests an involvement of pro-opiomelanocortin (POMC) gene products (e.g., beta-endorphin) in modulating cocaine-induced reward and addiction-like behaviors in rodents. In this study, we investigated whether chronic "binge" cocaine and its withdrawal altered POMC gene expression in the brain of rats. Male Fischer rats were treated with two different chronic (14-day) "binge" pattern cocaine administration regimens (three injections at 1-h intervals, i.p.): steady-dose (45mg/kg/day) and escalating-dose (90mg/kg on the last day). Although there was no POMC mRNA alteration after chronic steady-dose cocaine, a significant decrease in POMC mRNA levels in the hypothalamus was found after chronic escalating-dose cocaine. In contrast, after acute (1-day) withdrawal from chronic "binge" escalating-dose regimen, but not steady-dose regimen, there were increased hypothalamic POMC mRNA levels that persisted into 14days of protracted withdrawal. To study the role of the endogenous opioid systems in the cocaine withdrawal effects, we administered a single naloxone injection (1mg/kg) that caused elevated POMC mRNA levels observed 24h later in cocaine naïve rats, but it did not lead to further increases in cocaine-withdrawn rats. Our results suggest that during withdrawal from chronic "binge" escalating-dose cocaine: (1) there was a persistent increase in hypothalamic POMC gene expression; and (2) hyposensitivity of the POMC gene expression to naloxone indicates altered opioidergic tone at or above the hypothalamic level.

Pharmacology and anti-addiction effects of the novel κ opioid receptor agonist Mesyl Sal B, a potent and long-acting analogue of salvinorin A.

BACKGROUND AND PURPOSE: Acute activation of κ opioid (KOP) receptors results in anticocaine-like effects, but adverse effects, such as dysphoria, aversion, sedation and depression, limit their clinical development. Salvinorin A, isolated from the plant Salvia divinorum, and its semi-synthetic analogues have been shown to have potent KOP receptor agonist activity and may induce a unique response with similar anticocaine addiction effects as the classic KOP receptor agonists, but with a different side effect profile. EXPERIMENTAL APPROACH: We evaluated the duration of effects of Mesyl Sal B in vivo utilizing antinociception assays and screened for cocaine-prime induced cocaine-seeking behaviour in self-administering rats to predict anti-addiction effects. Cellular transporter uptake assays and in vitro voltammetry were used to assess modulation of dopamine transporter (DAT) function and to investigate transporter trafficking and kinase signalling pathways modulated by KOP receptor agonists. KEY RESULTS: Mesyl Sal B had a longer duration of action than SalA, had anti-addiction properties and increased DAT function in vitro in a KOP receptor-dependent and Pertussis toxin-sensitive manner. These effects on DAT function required ERK1/2 activation. We identified differences between Mesyl Sal B and SalA, with Mesyl Sal B increasing the Vmax of dopamine uptake without altering cell-surface expression of DAT. CONCLUSIONS AND IMPLICATIONS: SalA analogues, such as Mesyl Sal B, have potential for development as anticocaine agents. Further tests are warranted to elucidate the mechanisms by which the novel salvinorin-based neoclerodane diterpene KOP receptor ligands produce both anti-addiction and adverse side effects. LINKED ARTICLES: This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2.

Determination of cocaine/crack biomarkers in colostrum by LC-MS following protein precipitation.

Drug abuse by nursing mothers is an ongoing concern because it may cause many adverse effects to the newborns. The development of analytical methods to analyze drugs of abuse in colostrum (first milk produced after birth) has a huge importance, because it enables the monitoring and the correct follow-up to users and newborns. A liquid chromatography mass spectrometry (LC-MS) method was developed and validated for the determination of cocaine and smoked cocaine (crack) biomarkers in colostrum. Cocaine (COC) and its major metabolite benzoylecgonine (BZE), the pyrolytic products anhydroecgonine methyl ester (AEME) and anhydroecgonine (AEC) were analyzed after a simple protein precipitation procedure using atropine (ATP) as internal standard (IS). Applying a chemometric approach study, all peaks were chromatographically separated at isocratic condition with a Kinetex HILIC column for polar compounds, at 30°C in 12min. One ion was detected for the quantification and three ions for confirmation of each analyte. The method was linear for all analytes in the concentration range of 5-300ng/mL with correlation coefficients (r) between 0.9983 and 0.9996. The lower limit of quantification (LLOQ) was 5ng/mL with acceptable validation parameters. Matrix effect was assessed by post-extraction addition approach and showed good results, demonstrating that protein precipitation cleaning procedure is fast, reliable and demand small quantities of organic solvent. The LC-MS method is fast and cheap compared to other equipments and was also successfully applied to assess real samples of colostrum from nursing mothers who were suspect of cocaine/crack abuse.

Salvinorin A analogs and other κ-opioid receptor compounds as treatments for cocaine abuse.

Acute activation of kappa-opioid receptors produces anti-addictive effects by regulating dopamine levels in the brain. Unfortunately, classic kappa-opioid agonists have undesired side effects such as sedation, aversion, and depression, which restrict their clinical use. Salvinorin A (Sal A), a novel kappa-opioid receptor agonist extracted from the plant Salvia divinorum, has been identified as a potential therapy for drug abuse and addiction. Here, we review the preclinical effects of Sal A in comparison with traditional kappa-opioid agonists and several new analogs. Sal A retains the anti-addictive properties of traditional kappa-opioid receptor agonists with several improvements including reduced side effects. However, the rapid metabolism of Sal A makes it undesirable for clinical development. In an effort to improve the pharmacokinetics and tolerability of this compound, kappa-opioid receptor agonists based on the structure of Sal A have been synthesized. While work in this field is still in progress, several analogs with improved pharmacokinetic profiles have been shown to have anti-addictive effects. While in its infancy, it is clear that these compounds hold promise for the future development of anti-addictive therapeutics.

Hypothalamic proteoglycan syndecan-3 is a novel cocaine addiction resilience factor.

Proteoglycans like syndecan-3 have complex signaling roles in addition to their function as structural components of the extracellular matrix. Here, we show that syndecan-3 in the lateral hypothalamus has an unexpected new role in limiting compulsive cocaine intake. In particular, we observe that syndecan-3 null mice self-administer greater amounts of cocaine than wild-type mice. This effect can be rescued by re-expression of syndecan-3 in the lateral hypothalamus with an adeno-associated viral vector. Adeno-associated viral vector delivery of syndecan-3 to the lateral hypothalamus also reduces motivation for cocaine in normal mice. Syndecan-3 limits cocaine intake by modulating the effects of glial-cell-line-derived neurotrophic factor, which uses syndecan-3 as an alternative receptor. Our findings indicate syndecan-3-dependent signaling as a novel therapeutic target for the treatment of cocaine addiction.

Guanfacine effects on stress, drug craving and prefrontal activation in cocaine dependent individuals: preliminary findings.

Cocaine dependence is associated with increased stress and drug cue-induced craving and physiological arousal but decreased prefrontal activity to emotional and cognitive challenge. As these changes are associated with relapse risk, we investigated the effects of α2 receptor agonist guanfacine on these processes. Twenty-nine early abstinent treatment-seeking cocaine dependent individuals were randomly assigned to either daily placebo or guanfacine (up to 3 mg) for four weeks. In a laboratory experiment, all patients were exposed to three 10-min guided imagery conditions (stress/stress, drug cue/drug cue, stress/drug cue), one per day, consecutively in a random, counterbalanced order. Subjective craving, anxiety and arousal as well as cardiovascular output were assessed repeatedly. Brain response to stress, drug cue and relaxing imagery was also assessed during a functional magnetic resonance (fMRI) imaging session. In the current study, guanfacine was found to be safe and well-tolerated. Lower basal heart rate and blood pressure was observed in the guanfacine versus placebo group. Guanfacine lowered stress and cue-induced nicotine craving and cue-induced cocaine craving, anxiety and arousal. The guanfacine group also showed increased medial and lateral prefrontal activity following stress and drug cue exposure compared with placebo. Data suggest further exploration of guanfacine is warranted in terms of its potential for reducing stress-induced and cue-induced drug craving and arousal.

Orexin-1 receptor signalling within the ventral tegmental area, but not the paraventricular thalamus, is critical to regulating cue-induced reinstatement of cocaine-seeking.

Orexinergic signalling is critical to drug relapse-like behaviour; however, the CNS sites(s) of action remain unknown. Two candidate brain regions are the paraventricular thalamus (PVT) and ventral tegmental area (VTA). We assessed the effect of intra-PVT or -VTA administration of the orexin-1 receptor (OrxR1) antagonist SB-334867 on discriminative cue-induced cocaine-seeking. Animals received either PVT- or VTA-directed SB-334867 (0, 3 or 6 µg; 0, 1 or 3 µg, respectively) prior to reinstatement testing elicited by presenting cocaine-paired stimuli (S+). The effect of VTA-directed injections of SB-334867 (0 or 3 µg) on locomotor activity was also assessed. Intra-VTA, but not -PVT, SB-334867 dose-dependently attenuated S+-induced reinstatement (3 µg dose, p<0.01). Intra-VTA SB-334867 had no effect on locomotor activity. We conclude that OrxR1 signalling within the VTA, but not the PVT, mediates cue-induced cocaine-seeking behaviour. We hypothesize that blockade of VTA OrxR1 signalling may reduce nucleus accumbens dopamine in response to drug cue presentation.

Acoustic startle reduction in cocaine dependence persists for 1 year of abstinence.

RATIONALE: Chronic cocaine use results in long-lasting neurochemical changes that persist beyond the acute withdrawal period. Previous work from our group reported a profound reduction in the acoustic startle response (ASR) in chronic cocaine-dependent subjects in early abstinence compared to healthy controls that may be related to long-lasting neuroadaptations following withdrawal from chronic cocaine use. OBJECTIVES: This study aims to investigate the persistence and time course of the decrements in the ASR of cocaine-dependent subjects during prolonged abstinence. METHODS: Seventy-six cocaine-dependent (COC) subjects and 30 controls (CONT) were tested, the former after a period of heavy cocaine dependence. COC subjects were retested sequentially for 1 year of abstinence or until relapse. ASR testing was conducted at 3-dB levels and the eye-blink component of the startle response was quantified with electromyographic recording of the orbicularis oculi muscle. RESULTS: While there was no difference in startle magnitude between CONT and COC in early abstinence, by day 40 of abstinence COC subjects exhibited a statistically significant decline (p = 0.0057) in ASR magnitude as compared with CONT and this decrement persisted for up to 1 year of abstinence (p = 0.0165). In addition, startle latency was slower in COC subjects as compared with CONT at all stages of abstinence. CONCLUSIONS: These results replicate and expand upon the earlier finding that chronic cocaine use impairs the ASR in a manner that persists beyond the acute withdrawal period. This phenomenon may represent a biological measure of long-term neural changes accompanying cocaine dependence and subsequent withdrawal.

Cocaine- and amphetamine-regulated transcript (CART) signaling within the paraventricular thalamus modulates cocaine-seeking behaviour.

BACKGROUND: Cocaine- and amphetamine-regulated transcript (CART) has been demonstrated to play a role in regulating the rewarding and reinforcing effects of various drugs of abuse. A recent study demonstrated that i.c.v. administration of CART negatively modulates reinstatement of alcohol seeking, however, the site(s) of action remains unclear. We investigated the paraventricular thalamus (PVT) as a potential site of relapse-relevant CART signaling, as this region is known to receive dense innervation from CART-containing hypothalamic cells and to project to a number of regions known to be involved in mediating reinstatement, including the nucleus accumbens (NAC), medial prefrontal cortex (mPFC) and basolateral amygdala (BLA). METHODOLOGY/PRINCIPAL FINDINGS: Male rats were trained to self-administer cocaine before being extinguished to a set criterion. One day following extinction, animals received intra-PVT infusions of saline, tetrodotoxin (TTX; 2.5 ng), CART (0.625 µg or 2.5 µg) or no injection, followed by a cocaine prime (10 mg/kg, i.p.). Animals were then tested under extinction conditions for one hour. Treatment with either TTX or CART resulted in a significant attenuation of drug-seeking behaviour following cocaine-prime, with the 2.5 µg dose of CART having the greatest effect. This effect was specific to the PVT region, as misplaced injections of both TTX and CART resulted in responding that was identical to controls. CONCLUSIONS/SIGNIFICANCE: We show for the first time that CART signaling within the PVT acts to inhibit drug-primed reinstatement of cocaine seeking behaviour, presumably by negatively modulating PVT efferents that are important for drug seeking, including the NAC, mPFC and BLA. In this way, we identify a possible target for future pharmacological interventions designed to suppress drug seeking.

Mitochondrial impairment contributes to cocaine-induced cardiac dysfunction: Prevention by the targeted antioxidant MitoQ.

The goal of this study was to assess mitochondrial function and ROS production in an experimental model of cocaine-induced cardiac dysfunction. We hypothesized that cocaine abuse may lead to altered mitochondrial function that in turn may cause left ventricular dysfunction. Seven days of cocaine administration to rats led to an increased oxygen consumption detected in cardiac fibers, specifically through complex I and complex III. ROS levels were increased, specifically in interfibrillar mitochondria. In parallel there was a decrease in ATP synthesis, whereas no difference was observed in subsarcolemmal mitochondria. This uncoupling effect on oxidative phosphorylation was not detectable after short-term exposure to cocaine, suggesting that these mitochondrial abnormalities were a late rather than a primary event in the pathological response to cocaine. MitoQ, a mitochondrial-targeted antioxidant, was shown to completely prevent these mitochondrial abnormalities as well as cardiac dysfunction characterized here by a diastolic dysfunction studied with a conductance catheter to obtain pressure-volume data. Taken together, these results extend previous studies and demonstrate that cocaine-induced cardiac dysfunction may be due to a mitochondrial defect.

Functional activation of monoamine transporters by luteolin and apigenin isolated from the fruit of Perilla frutescens (L.) Britt.

Monoamine transporters playing major roles in regulating normal and abnormal synaptic activity are associated with various neuropsychological disorders. In spite of the discovery of a series of structurally different monoamine transporter antagonists for the therapy approach, no practical pharmaceutical can act as a transporter activator. Here, we isolated luteolin and apigenin from the fruit of Perilla frutescens (L.) Britt by using an activity-guided extraction technique, and proved that the two compounds possess actions of enhancing monoamine uptake either upon monoamine-transporter transgenic Chinese hamster ovary (CHO) cells or upon wild dopaminergic cell lines, with higher specificity for dopamine (DA) uptake than for norepinephrine (NE)- and serotonin (5HT)-uptake, as well as with more potency and greater efficacy for luteolin than for apigenin. Further, in the transgenic cells, the principal NE/DA uptake activation by luteolin was significantly prevented by respective transporter inhibitor, and the transmitter-uptake-enhancing action was independent of its ligands, which is in support of the compounds as monoamine transporter activators. Furthermore, luteolin evoked a marked disinhibition of cocaine-targeted effect in CHO cells overexpressing dopamine transporter. Thus, luteolin and apigenin function as monoamine transporter activators, which would improve several hypermonoaminergic neuropsychological disorders, especially cocaine dependence, through up-regulating monoamine transporter activity.

Acupuncture attenuates cocaine-induced expression of behavioral sensitization in rats: possible involvement of the dopaminergic system in the ventral tegmental area.

Acupuncture is widely used for the treatment of many functional disorders, such as substance abuse, and has the suppressive effect on the central nervous system. Many studies have suggested that behavioral sensitization by repeated injections of cocaine produce an increase in locomotor activity and an increase in the expression of tyrosine hydroxylase (TH), in the central dopaminergic system. In order to investigate the effects of acupuncture on the repeated cocaine-induced neuronal and behavioral sensitization alternations, we examined the influence of acupuncture on the repeated cocaine-induced locomotor activity and the expression of TH in the brain using immunohistochemistry. Male SD rats were given repeated injections of cocaine hydrochloride (15 mg/kg, i.p. for 10 consecutive days) followed by one challenge injection on the 4th day after the last daily injection. Cocaine challenge produced a large increase in the locomotor activity and the expression of TH in the ventral tegmental area (VTA). Treatment with acupuncture bilaterally at the Shenman (HT7) points for 1 min significantly inhibited the increase of locomotor activity as well as the TH expression in the VTA. Our data demonstrated that the inhibitory effects of acupuncture on cocaine-induced expression of behavioral sensitization were closely associated with the reduction of dopamine (DA) biosynthesis and the postsynaptic neuronal activity. These results provide evidence that acupuncture may be effective for inhibiting the behavioral effects of cocaine by possible modulation of the central dopaminergic system.