5-HT2A Receptor Knockout Mice Show Sex-Dependent Differences following Acute Noribogaine Administration.

Noribogaine (noribo) is the primary metabolite from ibogaine, an atypical psychedelic alkaloid isolated from the root bark of the African shrub Tabernanthe iboga. The main objective of this study was to test the hypothesis that molecular, electrophysiological, and behavioral responses of noribo are mediated by the 5-HT2A receptor (5-HT2AR) in mice. In that regard, we used male and female, 5-HT2AR knockout (KO) and wild type (WT) mice injected with a single noribo dose (10 or 40 mg/kg; i.p.). After 30 min., locomotor activity was recorded followed by mRNA measurements by qPCR (immediate early genes; IEG, glutamate receptors, and 5-HT2AR levels) and electrophysiology recordings of layer V pyramidal neurons from the medial prefrontal cortex. Noribo 40 decreased locomotion in male, but not female WT. Sex and genotype differences were observed for IEG and glutamate receptor expression. Expression of 5-HT2AR mRNA increased in the mPFC of WT mice following Noribo 10 (males) or Noribo 40 (females). Patch-clamp recordings showed that Noribo 40 reduced the NMDA-mediated postsynaptic current density in mPFC pyramidal neurons only in male WT mice, but no effects were found for either KO males or females. Our results highlight that noribo produces sexually dimorphic effects while the genetic removal of 5HT2AR blunted noribo-mediated responses to NMDA synaptic transmission.

Gamma oscillations in the pedunculopontine nucleus are regulated by F-actin: neuroepigenetic implications.

The pedunculopontine nucleus (PPN) is part of the reticular activating system (RAS) in charge of arousal and rapid eye movement sleep. The presence of high-frequency membrane oscillations in the gamma-band range in the PPN has been extensively demonstrated both in vivo and in vitro. Our group previously described histone deacetylation (HDAC) inhibition in vitro induced protein changes in F-actin cytoskeleton and intracellular Ca2+ concentration regulation proteins in the PPN. Here, we present evidence that supports the presence of a fine balance between HDAC function and calcium calmodulin kinase II-F-actin interactions in the PPN. We modified F-actin polymerization in vitro by using jasplakinolide (1 µM, a promoter of F-actin stabilization), or latrunculin-B (1 µM, an inhibitor of actin polymerization). Our results showed that shifting the balance in either direction significantly reduced PPN gamma oscillation as well as voltage-dependent calcium currents.

HDAC superfamily promoters acetylation is differentially regulated by modafinil and methamphetamine in the mouse medial prefrontal cortex.

Dysregulation of histone deacetylases (HDAC) has been proposed as a potential contributor to aberrant transcriptional profiles that can lead to changes in cognitive functions. It is known that METH negatively impacts the prefrontal cortex (PFC) leading to cognitive decline and addiction whereas modafinil enhances cognition and has a low abuse liability. We investigated if modafinil (90 mg/kg) and methamphetmine (METH) (1 mg/kg) may differentially influence the acetylation status of histones 3 and 4 (H3ac and H4ac) at proximal promoters of class I, II, III, and IV HDACs. We found that METH produced broader acetylation effects in comparison with modafinil in the medial PFC. For single dose, METH affected H4ac by increasing its acetylation at class I Hdac1 and class IIb Hdac10, decreasing it at class IIa Hdac4 and Hdac5. Modafinil increased H3ac and decreased H4ac of Hdac7. For mRNA, single-dose METH increased Hdac4 and modafinil increased Hdac7 expression. For repeated treatments (4 d after daily injections over 7 d), we found specific effects only for METH. We found that METH increased H4ac in class IIa Hdac4 and Hdac5 and decreased H3/H4ac at class I Hdac1, Hdac2, and Hdac8. At the mRNA level, repeated METH increased Hdac4 and decreased Hdac2. Class III and IV HDACs were only responsive to repeated treatments, where METH affected the H3/H4ac status of Sirt2, Sirt3, Sirt7, and Hdac11. Our results suggest that HDAC targets linked to the effects of modafinil and METH may be related to the cognitive-enhancing vs cognitive-impairing effects of these psychostimulants.

Bottom-up gamma and bipolar disorder, clinical and neuroepigenetic implications.

OBJECTIVES: This limited review examines the role of the reticular activating system (RAS), especially the pedunculopontine nucleus (PPN), one site of origin of bottom-up gamma, in the symptoms of bipolar disorder (BD). METHODS: The expression of neuronal calcium sensor protein 1 (NCS-1) in the brains of BD patients is increased. It has recently been found that all PPN neurons manifest intrinsic membrane beta/gamma frequency oscillations mediated by high threshold calcium channels, suggesting that it is one source of bottom-up gamma. This review specifically addresses the involvement of these channels in the manifestation of BD. RESULTS: Excess NCS-1 was found to dampen gamma band oscillations in PPN neurons. Lithium, a first line treatment for BD, was found to decrease the effects of NCS-1 on gamma band oscillations in PPN neurons. Moreover, gamma band oscillations appear to epigenetically modulate gene transcription in PPN neurons, providing a new direction for research in BD. CONCLUSIONS: This is an area needing much additional research, especially since the dysregulation of calcium channels may help explain many of the disorders of arousal in, elicit unwanted neuroepigenetic modulation in, and point to novel therapeutic avenues for, BD.

Cocaine alters the mouse testicular epigenome with direct impact on histone acetylation and DNA methylation marks.

RESEARCH QUESTION: Recent evidence suggests that cocaine administration in animal models can trigger non-genetic inheritance of addiction traits from father to offspring, affecting development and behaviour. Is chronic cocaine intake involved in alterations of epigenetic homeostasis in the testis? DESIGN: Epigenetic marks and mediators in testis and isolated germ cells of adult mice treated with cocaine (10 mg/kg) or vehicle (sterile saline solution) were evaluated in an intermittent binge protocol: three intraperitoneal injections, 1 h apart, one day on/off for 13 days, collecting tissue 24 h after the last binge administration (day 14). RESULTS: It was shown that chronic cocaine intake in mice disrupts testicular epigenetic homeostasis, increasing global methylated cytosine levels in DNA from germ cells and sperm. Cocaine also increased testicular and germ cell acetylated histone 3 and 4 and decreased expression of histone deacetylases HDAC1/2. Immunolocalization studies showed that HDAC1/2 and acetylated histone 3 and 4 proteins localize to meiotic germ cells. Analysis of mRNA expression in isolated germ cells shows decreased levels of Hdac1/2/8, Dnmt3b and Tet1 and increased levels of Dnmt3a gene expression after cocaine treatment. CONCLUSIONS: Cocaine intake is associated with testicular toxicity and significant reproductive function impairment. The results presented here broaden the basic knowledge of the impact of addictive stimulants on testicular pathophysiology, fertility and male reproductive health and imply that altered epigenetic homeostasis by cocaine may have potential consequences on future generations.

Modulation of GABA release from the thalamic reticular nucleus by cocaine and caffeine: role of serotonin receptors.

Serotonin receptors are targets of drug therapies for a variety of neuropsychiatric and neurodegenerative disorders. Cocaine inhibits the re-uptake of serotonin (5-HT), dopamine, and noradrenaline, whereas caffeine blocks adenosine receptors and opens ryanodine receptors in the endoplasmic reticulum. We studied how 5-HT and adenosine affected spontaneous GABAergic transmission from thalamic reticular nucleus. We combined whole-cell patch clamp recordings of miniature inhibitory post-synaptic currents (mIPSCs) in ventrobasal thalamic neurons during local (puff) application of 5-HT in wild type (WT) or knockout mice lacking 5-HT2A receptors (5-HT2A -/-). Inhibition of mIPSCs frequency by low (10 µM) and high (100 µM) 5-HT concentrations was observed in ventrobasal neurons from 5-HT2A -/- mice. In WT mice, only 100 µM 5-HT significantly reduced mIPSCs frequency. In 5-HT2A -/- mice, NAN-190, a specific 5-HT1A antagonist, prevented the 100 µM 5-HT inhibition while blocking H-currents that prolonged inhibition during post-puff periods. The inhibitory effects of 100 µM 5-HT were enhanced in cocaine binge-treated 5-HT2A -/- mice. Caffeine binge treatment did not affect 5-HT-mediated inhibition. Our findings suggest that both 5-HT1A and 5-HT2A receptors are present in pre-synaptic thalamic reticular nucleus terminals. Serotonergic-mediated inhibition of GABA release could underlie aberrant thalamocortical physiology described after repetitive consumption of cocaine. Our findings suggest that both 5-HT1A , 5-HT2A and A1 receptors are present in pre-synaptic TRN terminals. 5-HT1A and A1 receptors would down-regulate adenylate cyclase, whereas 5-HT1A would also increase the probability of the opening of G-protein-activated inwardly rectifying K(+) channels (GIRK). Sustained opening of GIRK channels would hyperpolarize pre-synaptic terminals activating H-currents, resulting in less GABA release. 5-HT2A -would activate PLC and IP3 , increasing intracellular [Ca(2+) ] and thus facilitating GABA release.

Cognitive enhancers versus addictive psychostimulants: The good and bad side of dopamine on prefrontal cortical circuits.

In this review we describe how highly addictive psychostimulants such as cocaine and methamphetamine actions might underlie hypoexcitabilty in frontal cortical areas observed in clinical and preclinical models of psychostimulant abuse. We discuss new mechanisms that describe how increments on synaptic dopamine release are linked to reduce calcium influx in both pre and postsynaptic compartments on medial PFC networks, therefore modulating synaptic integration and information. Sustained DA neuromodulation by addictive psychostimulants can "lock" frontal cortical networks in deficient states. On the other hand, other psychostimulants such as modafinil and methylphenidate are considered pharmacological neuroenhancement agents that are popular among healthy people seeking neuroenhancement. More clinical and preclinical research is needed to further clarify mechanisms of actions and physiological effects of cognitive enhancers which show an opposite pattern compared to chronic effect of addictive psychostimulants: they appear to increase cortical excitability. In conclusion, studies summarized here suggest that there is frontal cortex hypoactivity and deficient inhibitory control in drug-addicted individuals. Thus, additional research on physiological effects of cognitive enhancers like modafinil and methylphenidate seems necessary in order to expand current knowledge on mechanisms behind their therapeutic role in the treatment of addiction and other neuropsychiatric disorders.

Methamphetamine blunts Ca(2+) currents and excitatory synaptic transmission through D1/5 receptor-mediated mechanisms in the mouse medial prefrontal cortex.

Psychostimulant addiction is associated with dysfunctions in frontal cortex. Previous data demonstrated that repeated exposure to methamphetamine (METH) can alter prefrontal cortex (PFC)-dependent functions. Here, we show that withdrawal from repetitive non-contingent METH administration (7 days, 1 mg/kg) depressed voltage-dependent calcium currents (ICa ) and increased hyperpolarization-activated cation current (IH ) amplitude and the paired-pulse ratio of evoked excitatory postsynaptic currents (EPSCs) in deep-layer pyramidal mPFC neurons. Most of these effects were blocked by systemic co-administration of the D1/D5 receptor antagonist SCH23390 (0.5 and 0.05 mg/kg). In vitro METH (i.e. bath-applied to slices from naïve-treated animals) was able to emulate its systemic effects on ICa and evoked EPSCs paired-pulse ratio. We also provide evidence of altered mRNA expression of (1) voltage-gated calcium channels P/Q-type Cacna1a (Cav 2.1), N-type Cacna1b (Cav 2.2), T-type Cav 3.1 Cacna1g, Cav 3.2 Cacna1h, Cav 3.3 Cacna1i and the auxiliary subunit Cacna2d1 (α2δ1); (2) hyperpolarization-activated cyclic nucleotide-gated channels Hcn1 and Hcn2; and (3) glutamate receptors subunits AMPA-type Gria1, NMDA-type Grin1 and metabotropic Grm1 in the mouse mPFC after repeated METH treatment. Moreover, we show that some of these changes in mRNA expression were sensitive D1/5 receptor blockade. Altogether, these altered mechanisms affecting synaptic physiology and transcriptional regulation may underlie PFC functional alterations that could lead to PFC impairments observed in METH-addicted individuals.

Neuropathology of substance use disorders.

Addictions to licit and illicit drugs are chronic relapsing brain disorders that affect circuits that regulate reward, motivation, memory, and decision-making. Drug-induced pathological changes in these brain regions are associated with characteristic enduring behaviors that continue despite adverse biopsychosocial consequences. Repeated exposure to these substances leads to egocentric behaviors that focus on obtaining the drug by any means and on taking the drug under adverse psychosocial and medical conditions. Addiction also includes craving for the substances and, in some cases, involvement in risky behaviors that can cause death. These patterns of behaviors are associated with specific cognitive disturbances and neuroimaging evidence for brain dysfunctions in a diverse population of drug addicts. Postmortem studies have also revealed significant biochemical and/or structural abnormalities in some addicted individuals. The present review provides a summary of the evidence that has accumulated over the past few years to implicate brain dysfunctions in the varied manifestations of drug addiction. We thus review data on cerebrovascular alterations, brain structural abnormalities, and postmortem studies of patients who abuse cannabis, cocaine, amphetamines, heroin, and "bath salts". We also discuss potential molecular, biochemical, and cellular bases for the varied clinical presentations of these patients. Elucidation of the biological bases of addiction will help to develop better therapeutic approaches to these patient populations.

Stress, sex, and addiction: potential roles of corticotropin-releasing factor, oxytocin, and arginine-vasopressin.

Stress sensitivity and sex are predictive factors for the development of neuropsychiatric disorders. Life stresses are not only risk factors for the development of addiction but also are triggers for relapse to drug use. Therefore, it is imperative to elucidate the molecular mechanisms underlying the interactions between stress and drug abuse, as an understanding of this may help in the development of novel and more effective therapeutic approaches to block the clinical manifestations of drug addiction. The development and clinical course of addiction-related disorders do appear to involve neuroadaptations within neurocircuitries that modulate stress responses and are influenced by several neuropeptides. These include corticotropin-releasing factor, the prototypic member of this class, as well as oxytocin and arginine-vasopressin that play important roles in affiliative behaviors. Interestingly, these peptides function to balance emotional behavior, with sexual dimorphism in the oxytocin/arginine-vasopressin systems, a fact that might play an important role in the differential responses of women and men to stressful stimuli and the specific sex-based prevalence of certain addictive disorders. Thus, this review aims to summarize (i) the contribution of sex differences to the function of dopamine systems, and (ii) the behavioral, neurochemical, and anatomical changes in brain stress systems.

Differential effects of methylphenidate and cocaine on GABA transmission in sensory thalamic nuclei.

Methylphenidate (MPH) is widely used to treat children and adolescents diagnosed with attention deficit/hyperactivity disorder. Although MPH shares mechanistic similarities to cocaine, its effects on GABAergic transmission in sensory thalamic nuclei are unknown. Our objective was to compare cocaine and MPH effects on GABAergic projections between thalamic reticular and ventrobasal (VB) nuclei. Mice (P18-30) were subjected to binge-like cocaine and MPH acute and sub-chronic administrations. Cocaine and MPH enhanced hyperlocomotion, although sub-chronic cocaine-mediated effects were stronger than MPH effects. Cocaine and MPH sub-chronic administration altered paired-pulse and spontaneous GABAergic input differently. The effects of cocaine on evoked paired-pulse GABA-mediated currents changed from depression to facilitation with the duration of the protocols used, while MPH induced a constant increase throughout the administration protocols. Thalamic reticular nucleus GAD67 and VB Ca(V) 3.1 protein levels were measured using western blot to better understand their link to increased GABA release. Both proteins were increased by sub-chronic administration of cocaine. MPH showed effects on GABAergic transmission that seems less disruptive than cocaine. Unique effects of cocaine on postsynaptic VB calcium currents might explain deleterious cocaine effects on sensory thalamic nuclei. These results suggest that cocaine and MPH produced distinct presynaptic alterations on GABAergic transmission.

Moderate and severe perinatal asphyxia induces differential effects on cocaine sensitization in adult rats.

Perinatal asphyxia (PA) increases the likelihood of suffering from dopamine-related disorders, such as ADHD and schizophrenia. Since dopaminergic transmission plays a major role in cocaine sensitization, the purpose of this study was to determine whether PA could be associated with altered behavioral sensitization to cocaine. To this end, adult rats born vaginally (CTL), by caesarean section (C+), or by C+ with 15 min (PA15, moderate PA) or 19 min (PA19, severe PA) of global anoxia were repeatedly administered with cocaine (i.p., 15 mg/kg) and then challenged with cocaine (i.p., 15 mg/kg) after a 5-day withdrawal period. In addition, c-Fos, FosB/ΔFosB, DAT, and TH expression were assessed in dorsal (CPu) and ventral (NAcc) striatum. Results indicated that PA15 rats exhibited an increased locomotor sensitization to cocaine, while PA19 rats displayed an abnormal acquisition of locomotor sensitization and did not express a sensitized response to cocaine. c-Fos expression in NAcc, but not in CPu, was associated with these alterations in cocaine sensitization. FosB/ΔFosB expression was increased in all groups and regions after repeated cocaine administration, although it reached lower expression levels in PA19 rats. In CTL, C+, and PA15, but not in PA19 rats, the expression of TH in NAcc was reduced in groups repeatedly treated with cocaine, independently of the challenge test. Furthermore, this reduction was more pronounced in PA15 rats. DAT expression remained unaltered in all groups and regions studied. These results suggest that moderate PA may increase the vulnerability to drug abuse and in particular to cocaine addiction.

Attenuated methamphetamine induced neurotoxicity by modafinil administration in mice.

Methamphetamine (METH) is a highly addictive drug that might induce neurotoxicity. Clinical trials have reported that modafinil, a wake-promoting agent used to treat sleep disorders, may have some efficacy for the treatment of psychostimulant addiction. In this study we tested possible neuroprotective effects of modafinil after toxic METH administration in mice. We evaluated the effect of modafinil (two injections of either 90 or 180 mg/kg) and METH binge (3 × 7 mg/kg i.p. injections, 3-h apart) coadministration on DA striatal content, TH immunoreactivity in striatal areas and spontaneous locomotor activity. We also investigated acute locomotor activity and stereotypy profile in mice treated with a single METH dose (2 and 7 mg/kg) pretreated with modafinil (90 and 180 mg/kg). We found that mice treated with a METH binge showed a marked decrease in DA and dopaminergic metabolites as well as lower levels of TH immunoreactivity in the dorsal striatum. Pretreatment with modafinil (both 90 and 180 mg/kg) attenuated these effects but did not prevent METH induced decrease in locomotion. We also found that groups that received the combination of both modafinil and single dose METH showed a decrease in total distance traveled in an open field compared with METH groups. We observed an increment in the time mice expended doing stereotypic movements (continuous sniffing) in the group that received the combination of both METH and modafinil (i.e., decreasing locomotion). Our results suggest a possible protective role of modafinil against METH acute striatal toxicity.

Histone Deacetylases and Immediate Early Genes: Key Players in Psychostimulant-Induced Neuronal Plasticity.

IEGs play a critical functional role of in molecular, cellular, and behavioral alterations induced by psychostimulants. IEGs appear to have specific chromatin structures that may contribute to the rapid activation of their transcription. HDAC enzymes regulate reversible acetylation of lysine residues of histones and non-histone proteins. Dysregulation of HDACs has been proposed to modulate the establishment and maintenance of aberrant transcriptional programs and behaviors associated with cognitive dysfunctions and drug addiction. In this mini-review we focus our attention on recent discoveries concerning networks of protein-protein interactions for the two classes of HDAC protein family members that are highly expressed in neurons, class I and IIa HDACs. Because dynamic histone acetylation appears to be critical to IEG expression in the brain, we discuss the role of these epigenetic regulators on IEG expression induced by cocaine and methamphetamine intake.

Simultaneous administration of cocaine and caffeine dysregulates HCN and T-type channels.

RATIONALE: The abuse of psychostimulants has adverse consequences on the physiology of the central nervous system. In Argentina, and other South American countries, coca paste or "PACO" (cocaine and caffeine are its major components) is massively consumed with deleterious clinical consequences for the health and well-being of the general population. A scant number of studies have addressed the consequences of stimulant combination of cocaine and caffeine on the physiology of the somatosensory thalamocortical (ThCo) system. OBJECTIVES: Our aim was to study ion conductances that have important implications regulating sleep-wake states 24-h after an acute or chronic binge-like administration of a cocaine and caffeine mixture following previously analyzed pasta base samples ("PACO"-like binge") using mice. METHODS: We randomly injected (i.p.) male C57BL/6JFcen mice with a binge-like psychostimulants regimen during either 1 day (acute) or 1 day on/1 day off during 13 days for a total of 7 binges (chronic). Single-cell patch-clamp recordings of VB neurons were performed in thalamocortical slices 24 h after the last psychostimulant injection. We also recorded EEG/EMG from mice 24 h after being systemically treated with chronic administration of cocaine + caffeine versus saline, vehicle. RESULTS: Our results showed notorious changes in the intrinsic properties of the VB nucleus neurons that persist after 24-h of either acute or chronic binge administrations of combined cocaine and caffeine ("PACO"-like binge). Functional dysregulation of HCN (hyperpolarization-activated cyclic nucleotide-gated) and T-type VGC (voltage-gated calcium) channels was described 24-h after acute/chronic "PACO"-like administrations. Furthermore, intracellular basal [Ca2+] disturbances resulted a key factor that modulated the availability and the activation of T-type channels, altering T-type "window currents." As a result, all these changes ultimately shaped the low-threshold spikes (LTS)-associated Ca2+ transients, regulated the membrane excitability, and altered sleep-wake transitions. CONCLUSION: Our results suggest that deleterious consequences of stimulants cocaine and caffeine combination on the thalamocortical physiology as a whole might be related to potential neurotoxic effects of soaring intracellular [Ca2+].

IMT504 Provides Analgesia by Modulating Cell Infiltrate and Inflammatory Milieu in a Chronic Pain Model.

IMT504 is a non-CPG, non-coding synthetic oligodeoxinucleotide (ODN) with immunomodulatory properties and a novel inhibitory role in pain transmission, exerting long-lasting analgesic effects upon multiple systemic administrations. However, its mechanisms of anti-nociceptive action are still poorly understood. In the present study in male adult rats undergoing complete Freund's adjuvant-induced hindpaw inflammation, we focused in the analysis of the immunomodulatory role of IMT504 over the cellular infiltrate, the impact on the inflammatory milieu, and the correlation with its anti-allodynic role. By means of behavioral analysis, we determined that a single subcutaneous administration of 6 mg/kg of IMT504 is sufficient to exert a 6-week-long full reversal of mechanical and cold allodynia, compromising neither acute pain perception nor locomotor activity. Importantly, we found that the anti-nociceptive effects of systemic IMT504, plus quick reductions in hindpaw edema, were associated with a modulatory action upon cellular infiltrate of B-cells, macrophages and CD8+ T-cells populations. Accordingly, we observed a profound downregulation of several inflammatory leukocyte adhesion proteins, chemokines and cytokines, as well as of ß-endorphin and an increase in the anti-inflammatory cytokine, interleukin-10. Altogether, we demonstrate that at least part of the anti-nociceptive actions of IMT504 relate to the modulation of the peripheral immune system at the site of injury, favoring a switch from pro- to anti-inflammatory conditions, and provide further support to its use against chronic inflammatory pain. Graphical abstract GA short description - IMT504 systemic Administration. Systemic administration of the non-CpG ODN IMT504 results in a 6-week long blockade of pain-like behavior in association with anti-inflammatory responses at the site of injury. These include modulation of lymphoid and myeloid populations plus downregulated expression levels of multiple pro-inflammatory cytokines and ß-endorphin. Nocifensive responses and locomotion remain unaltered.

Molecular changes in the nucleus accumbens and prefrontal cortex associated with the locomotor sensitization induced by coca paste seized samples.

RATIONALE: In previous studies, we have demonstrated that seized samples of a smokable form of cocaine, also known as coca paste (CP), induced behavioral sensitization in rats. Interestingly, this effect was accelerated and enhanced when the samples were adulterated with caffeine. While the cocaine phenomenon is associated with persistent functional and structural alterations in the prefrontal cortex (PFC) and nucleus accumbens (NAc), the molecular mechanisms underlying the CP sensitization and the influence of caffeine remains still unknown. OBJECTIVE: We examined the gene expression in NAc and mPFC after the expression caffeine-adulterated and non-adulterated CP locomotor sensitization. METHODS: The locomotor sensitization was established in C57BL/6 mice, repeatedly treated with a CP-seized sample adulterated with caffeine (CP-2) and a non-adulterated one (CP-1). We then assessed the mRNA expression of receptor subunits of the dopaminergic and glutamatergic systems in the medial PFC (mPFC) and NAc. Other molecular markers (e.g., adenosinergic, endocannabinoid receptor subunits, and synaptic plasticity-associated genes) were also analyzed. RESULTS: Only CP-2-treated mice expressed locomotor sensitization. This phenomenon was associated with increased Drd1a, Gria1, Cnr1, and Syn mRNA expression levels in the NAc. Drd3 mRNA expression levels were only significantly increased in mPFC of CP-2-treated group. CONCLUSIONS: Our results demonstrated that caffeine actively collaborates in the induction of the molecular changes underlying CP sensitization. The present study provides new knowledge on the impact of active adulterants to understand the early dependence induced by CP consumption.

Differential effects of HDAC inhibitors on PPN oscillatory activity in vivo.

The pedunculopontine nucleus (PPN) has long been known to be part of the reticular activating system (RAS) in charge of arousal and REM sleep. We previously showed that in vitro exposure to a HDAC Class I and II mixed inhibitor (TSA), or a specific HDAC class IIa inhibitor (MC 1568), decreased PPN gamma oscillations. Given the lack of information on systemic in vivo treatments on neuronal synaptic properties, the present study was designed to investigate the systemic effect of HDAC inhibitors (HDACi) on PPN rhythmicity. Rat pups were injected (acute, single dose) with TSA (4 or 20 mg/kg), MC1568 (4 or 20 mg/kg), or MS275 (20 or 100 mg/kg). Our results show that MC1568 (20 mg/kg) reduced mean frequency of PPN oscillations at gamma band, while increasing mean input resistance (Rm) of PPN neurons. For TSA (4 and 20 mg/kg), we observed reduced mean frequency of oscillations at gamma band and increased mean Rm of PPN neurons. Systemic administration of 20 mg/kg MC1568 and TSA effects on Rm were washed out after 60 min of in vitro incubation of PPN slices, suggesting an underlying functional recovery of PPN calcium-mediated gamma band oscillations over time. In addition, at a lower dose, 4 mg/kg, MC1568 and TSA induced higher mean amplitude gamma oscillations. Blocking HDAC class I might not have deleterious effects on gamma activity, but, more importantly, the inhibition of HDAC class I (at 100 mg/kg) increased gamma band oscillations. In conclusion, the present results in vivo validate our previous findings in vitro and further expand information on the effects of HDAC inhibition on PPN rhythmicity. PPN neurons require normal activity of HDAC class IIa in order to oscillate at gamma band.

Acute Regulation of the Arousal-Enhancing Drugs Caffeine and Modafinil on Class IIa HDACs In Vivo and In Vitro: Focus on HDAC7.

Psychostimulant drugs, such as modafinil and caffeine, induce transcriptional alterations through the dysregulation of epigenetic mechanisms. We have previously demonstrated that acute modafinil administration is accompanied by multiple changes in the expression of histone deacetylases (HDACs) within the mouse medial prefrontal cortex (mPFC). Herein, we compared alterations in class IIa HDACs in the mouse mPFC and dorsal striatum (DS) after a single exposure to each psychostimulant. We treated male C57BL/6 mice with modafinil (90 mg/kg, i.p.), caffeine (10 mg/kg, i.p.), or vehicle and evaluated locomotor activity. Following, we examined hdac4, hdac5, and hdac7 mRNA expression using qRT-PCR and HDAC7, pHDAC7, and pHDACs4/5/7 using Western blot. Last, we explored generalized effects in N2a cell line using modafinil (100 µM and 1 mM) or caffeine (80 µM and 800 µM). Our results indicate that modafinil had greater effects on locomotor activity compared with caffeine. qRT-PCR experiments revealed that modafinil decreased hdac5 and hdac7 mRNA expression in the DS, while caffeine had no effects. In the mPFC, modafinil increased hdac7 mRNA expression, with no effects observed for caffeine. Western blot revealed that within the DS, modafinil induced increases in HDAC7, pHDAC7, and pHDACs4/5/7 protein expression, while, in the mPFC, caffeine induced decreases in HDAC7, pHDAC7, and pHDACs4/5/7 protein levels. In vitro studies revealed that modafinil increased hdac4, hdac5, and hdac7 mRNA levels in N2a, while caffeine only increased hdac5 at a higher dose. These findings support the notion that modafinil and caffeine exert distinct regulation of class IIa HDAC family members and that these transcriptional and translational consequences are region-specific.

Expression of immediate early genes in brain reward circuitries: Differential regulation by psychostimulant and opioid drugs.

Although some of the clinical manifestations of substance use disorders might be superficially similar, it is highly likely that different classes of abused drugs including opioids (heroin, morphine, and oxycodone, other opioids) and psychostimulants (cocaine and amphetamines) cause different neuroadaptations in various brain regions dependent in the distribution and concentration of their biochemical sites of actions. In fact, different molecular networks are indeed impacted by acute and chronic administration of addictive substances. Some of the genes whose expression is influenced by the administration of these substances are immediate-early genes (IEGs). IEGs include classes of low expression genes that can become very highly induced within seconds or minutes of activation by endogenous or exogenous stimuli. These IEGs might play important roles in activating target genes that regulate adaptations implicated in the behavioral manifestations diagnosed as addiction. Therefore, the purpose of this review is to provide an overview of recent data on the effects of psychostimulants and opioids on IEG expression in the brain. The review documents some contrasting effects of these classes of drugs on gene expression and indicates that further studies are necessary to identify the specific effects of each drug class when trying to predict clinical responses to therapeutic agents.