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Activation of arc, a Putative "Effector" Immediate Early Gene, by Cocaine in Rat Brain
Journal of Neurochemistry
Abstract
As immediate early genes (IEGs) are thought to play a critical role in mediating stimulus-induced neuronal plasticity, several laboratories have characterized the IEG response induced by cocaine to help define the changes in gene expression that may underlie its long-lasting behavioral effects. Although activation of several transcription factor IEGs has been described, little is known about which "effector" IEGs, if any, are also induced. In the present study, we have examined whether cocaine administration affects expression of a recently identified "effector" IEG, referred to as arc (activity-regulated, cytoskeleton-associated). This IEG encodes a protein with homology to spectrin that appears to be associated with the actin cytoskeleton. Using in situ hybridization, we have found that systemic cocaine administration elicits a robust, transient rise in arc mRNA levels in striatum, which is suppressed by D1 dopamine receptor blockade, reserpine treatment, or striatal 6-hydroxydopamine lesions. D2 receptor antagonist triggered arc expression when administered alone. Immunohistochemical studies indicated that Arc protein induced by cocaine is expressed in neuronal cell bodies and dendrites. As Arc appears to be component of the neuronal cytoskeleton, it may be involved in structural alterations underlying neuronal plasticity triggered by cocaine.
... Several studies have demonstrated an increase in Arc mRNA in the striatum and various other brain areas in response to amphetamine and cocaine (Tan et al., 2000;Fosnaugh et al., 1995;Fumagalli et al., 2006;Kodama et al., 1998). Arc is increased in the rat striatum within 15 min of cocaine administration and returns to basal levels by 2 h. ...
... Interestingly, DA D2 antagonists such as haloperidol, spiperone and eticlopride also induce Arc in the striatum. Collectively, these findings demonstrate a role for DA D1 antagonists in inducing Arc in response to cocaine (Fosnaugh et al., 1995;Fumagalli et al., 2006). Similarly, within 1 h of a single methamphetamine (METH) intraperitoneal injection, Arc significantly increases in the parietal, orbital and medial prefrontal cortex (mPFC) and to a lesser degree in medium-sized striatal neurons and the hippocampus, returning to basal levels by 6 h. ...
... The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. (Fosnaugh et al., 1995;Fumagalli et al., 2006) FTY720 Fingolimod functional sphingosine 1-phosphate receptor antagonist decreases Arc in migratory dendritic cells Ufer et al. (2016) interleukin 6 (IL-6) neuroinflammatory agent increases Arc in dorsal root ganglion neurons Barragan-Iglesias et al. ...
Activity-regulated cytoskeleton-associated protein (Arc) is a brain-enriched immediate early gene that regulates important mechanisms implicated in learning and memory. Arc levels are controlled through a balance of induction and degradation in an activity-dependent manner. Arc further undergoes multiple post-translational modifications (PTMs) that regulate its stability, localization and function. Recent studies demonstrate that these features of Arc can be pharmacologically manipulated. In this review, we discuss some of these compounds, with an emphasis on drugs of abuse and psychotropic drugs. We also discuss inflammatory states which have also been shown to regulate Arc.
... Recent studies have conveyed to the role of immediate early genes (IEGs) in mediating stimulusinduced neuronal plasticity (Fosnaugh et al., 1995;Goelet et al., 1986;Morgan and Curran, 1989;Sheng and Greenberg, 1990;Worley et al., 1990). IEGs are a subset of genes that is induced by neural activity (e.g., via NMDA receptor activation and neuronal calcium influx) during behavioral and sensory experience (Heroux et al., 2018;Minatohara et al., 2015). ...
... Actin cytoskeleton plays an important role in regulating neuronal responsiveness (Fosnaugh et al., 1995;Johnson and Byerly, 1993;Rosenmund and Westbrook, 1993), and drugs of abuse, such as cocaine, appear to elicit changes in the cellular cytoskeleton (Nestler, 1993). More specifically, drugs of abuse affect the pattern of neurofilament expression and the rate of axonal transport (Beitner- Johnson et al., 1992;Nestler, 1993). ...
... Human neuroimaging studies found correlation of those cognitive dysfunctions and hypofunction of the frontal area in cocaine abusers (Goldstein and Volkow, 2002). In preclinical studies, it has been demonstrated that Arc expression increases after systemic injections of a numerous of psychostimulants, including methamphetamine (Kodama et al., 1998;Yamagata et al., 2000), methylphenidate (Chase et al., 2007), amphetamine (Gonzalez-Nicolini and McGinty, 2002;Klebaur et al., 2002) and cocaine (Fosnaugh et al., 1995;Fumagalli et al., 2006;Zavala et al., 2008) in several brain regions. Assessments of Arc mRNA expression following cue-induced reinstatement of rats subjected to repeated cocaine self-administration showed increased mRNA levels in ACC, Prl, IFL, OFC, BLA, CPu and NAc core in the cocaine-treated group compared to control yokedsaline group, indicating that these brain regions are responsive for cocaine-associated cues (Zavala et al., 2008). ...
Cocaine addiction is a chronic mental illness affecting a small subgroup of cocaine users (approx. 18%). Little efforts have been taken so far to understand individual differences in the vulnerability to cocaine addiction; i.e. it is unclear why some users become addicted whereas the majority of them are able to maintain the control over drug-taking and -seeking. Most preclinical studies have not considered the possibility to study individual differences in addiction vulnerability and furthermore may explain the high failure rates of drugs that reach Phase III-IV of clinical trials. Hence, a translational animal model is fundamental to produce meaningful results to the clinic. In this thesis, a DSM-IV/5-based preclinical model of cocaine addiction was used to identify pre-existing vulnerability differences and the changes produced by pathological state of addiction. The so-called 0/3crit animal model consists of training rats for intravenous cocaine self-administration for at least 45 sessions. Thereafter, three main DSM-based addiction behavioral criteria were tested, (1) motivation to take the drug, (2) persistence to seek the drug, and (3) persistence of self-administration despite negative consequences (i.e., application of an electric foot-shock). For each of the three criteria, a score of either 1 or 0 was given to animals performing above or below 60th percentile of the population distribution, respectively. Animals positive for all criteria (3crit) were classified as addicted-like, whereas animals negative for all criteria (0crit) were classified as non-addicted-like/resilient rats. This thesis had three aims: (i) Neuroimaging studies in cocaine addicts revealed various structural and functional changes but findings are inconclusive and this may be due to pre-existing features in the function or structure in brains of vulnerable subjects and different drug intake patterns. Based on this, longitudinal studies, particularly in controlled animal models, are warranted. Here a longitudinal translational multimodal neuroimaging study was performed using the 0/3crit animal model of cocaine addiction to investigate pre-existing differences and changes in brains of cocaine addicted-like and non-addicted-like rats. (ii) Conditioned cocaine cues can trigger craving and relapse. Pavlovian conditioned stimuli can also impact ongoing instrumental behavior, even if the instrumental behavior is acquired independently of Pavlovian conditioning. This process is called Pavlovian-to-Instrumental transfer (PIT). A hypothesis in drug addiction posits that Pavlovian conditioned cues can bias instrumental behavior towards drug seeking and intake and that a more pronounced PIT response occurs in vulnerable subjects. The second aim was therefore to test this hypothesis and to ask whether 3crit rats show a more pronounced PIT response than 0crit rats. (iii) Pharmacological treatments in cocaine addiction are still lacking. In previous preclinical studies different glutamate receptor antagonists and GABAB agonists such as baclofen, have been tested but either failed in clinical studies or reported severe side effects in pharmacovigilance. Here, different pharmacological approaches were taken: First, the novel GABAB positive allosteric modulator CMPPE was tested in comparison to baclofen in order to produce lesser side effects. Second the NR3A subunit was selectively targeted as an alternative to NMDAR antagonists. Finally, melatonin was tested for normalizing altered circadian rhythmicity in cocaine addiction. The longitudinal MRI results revealed structural changes in the brains of 3crit and 0crit rats. Grey matter (GM) volume was found to have increased in prelimbic (Prl) and cingulate (Cg) cortices, nucleus accumbens (NAc), caudate putamen (CPu), substantia nigra, and ventral and globus pallidum in the 3crit group, whereas 0crit rats showed no changes in GM volume, except for the CPu, compared to control. Diffusion tensor imaging (DTI) analysis revealed a higher fractional anisotropy in the zona incerta in the 0crit rats compared to 3crit rats. PET results showed higher activity of mPFC and right CPu in the 0crit group compared to controls. Arc expression was significantly reduced in the infralimbic (IFL) cortex in the 3crit group. Salience of conditioned-cues was found similar in 0crit and 3crit rats in the PIT paradigm. Positive allosteric modulator of GABAB receptors and melatonin abolished cue-elicited cocaine-seeking behavior. In summary, cocaine addiction produced structural changes in brain regions central for motivation and drug rewarding effects in 3crit rats, whereas the addiction resilient rats showed increased volume in brain regions involved in habit behavior as well as an increased in microstructural integrity in a brain area that regulates adaptive behavior. Functional assessments indicated the relevance of the mPFC (Prl and IFL) activity for both controlled or compulsive drug-seeking and –taking. These results agree with clinical studies, where mPFC function negatively correlates with impulsive behavior in psychostimulant abusers as well as changes were found in brain regions, such as Prl, Cg, CPu, and NAc, that are known to play a role in drug addiction. Moreover, other factors, such as structural and functional changes, instead of Pavlovian or instrumental conditioning may lead to addiction behaviors because salience to conditioned stimuli and learning ability were similar between the groups. And finally, positive allosteric modulator of GABAB receptors and melatonin appears to be promising candidates for medication development in cocaine addiction. The 0/3crit model of cocaine addiction has excellent face validity and can be used to study the underpinning mechanisms that lead to compulsive drug use.
... Besides electrical activity, Arc expression in adult brain is influenced by a variety of neuromodulators in a brain regionspecific manner (Fosnaugh et al. 1995;Pei et al. 2000;Sanders et al. 2008;Gil-Bea et al. 2011;Soule et al. 2012;de Bartolomeis et al. 2015). The frontal cortex receives prominent dopaminergic innervation (Verney et al. 1982;Kalsbeek et al. 1988) and activation of dopaminergic signaling through D1-type receptors (D1R), but not D2-type receptors (D2R), can enhance Arc transcription in adult brain (Fosnaugh et al. 1995). ...
... Besides electrical activity, Arc expression in adult brain is influenced by a variety of neuromodulators in a brain regionspecific manner (Fosnaugh et al. 1995;Pei et al. 2000;Sanders et al. 2008;Gil-Bea et al. 2011;Soule et al. 2012;de Bartolomeis et al. 2015). The frontal cortex receives prominent dopaminergic innervation (Verney et al. 1982;Kalsbeek et al. 1988) and activation of dopaminergic signaling through D1-type receptors (D1R), but not D2-type receptors (D2R), can enhance Arc transcription in adult brain (Fosnaugh et al. 1995). However, since dopaminergic signaling is not required for Arc expression induced by electrical activation in neuronal cultures, dopamine is generally thought to augment rather than to induce Arc expression (Shepherd et al. 2006;Bloomer et al. 2008). ...
... A previous study in adult rats suggested that D1R-specific antagonist SCH23390, but not D2R antagonist, might inhibit psychostimulant-induced Arc mRNA expression (Fosnaugh et al. 1995). In addition, D1R is expressed in neonatal frontal cortical neurons (Leslie et al. 1991). ...
The activity-regulated gene Arc/Arg3.1 encodes a postsynaptic protein crucially involved in glutamatergic synaptic plasticity. Genetic mutations in Arc pathway and altered Arc expression in human frontal cortex have been associated with schizophrenia. Although Arc expression has been reported to vary with age, what mechanisms regulate Arc mRNA levels in frontal cortex during postnatal development remains unclear. Using quantitative mRNA analysis of mouse frontal
cortical tissues, we mapped the developmental profiles of Arc expression and found that its mRNA levels are sharply amplified near the end of the second postnatal week, when mouse pups
open their eyes for the first time after birth. Surprisingly, electrical stimulation of the frontal cortex before eye-opening
is not sufficient to drive the amplification of Arc mRNA. Instead, this amplification needs both electrical stimulation and dopamine D1-type receptor (D1R) activation. Furthermore,
visual stimuli-driven amplification of Arc mRNA is also dependent on D1R activation and dopamine neurons located in the ventral midbrain. These results indicate that
dopamine is required to drive activity-dependent amplification of Arc mRNA in the developing postnatal frontal cortex and suggest that joint electrical and dopaminergic activation is essential
to establish the normal expression pattern of a schizophrenia-associated gene during frontal cortical development.
... This early transcription is characterized by the induction of immediate early genes (IEGs), which encode either transcription factors, including c-Fos, and Zif268, regulating a second wave of genes, or effector proteins acting directly on cellular functions and homeostasis. Activity-regulated cytoskeleton-associated protein (Arc), which belongs to this second category of IEGs, is rapidly induced by cocaine in cortical and striatal regions and cocaine-associated stimuli (3)(4)(5)(6)(7)(8)(9). However, the role of striatal Arc in the development of neuronal and behavioral adaptations to cocaine is not known. ...
... With regard to signaling cascades driving Arc expression, a D1R antagonist was previously shown to abolish Arc induction by acute cocaine (3). Herein, we confirm this observation and further demonstrate that Arc induction also strictly depends on NMDAR stimulation. ...
... In the striatum, Arc induction is associated with early phase of learning as well as inversion phase during reversal learning, suggesting a role in striatal plasticity underlying learning acquisition (48,49). In the context of addiction, Arc is increased by cocaine (3)(4)(5) or upon re-exposure to a context previously associated with the Nuclear Accumulation of Arc and Responses to Cocaine drug (6,7). In rats trained to self-administer cocaine, the local inhibition of Arc in the dorsolateral part of the striatum did not affect cocaine seeking but blocked extinction of this behavior (9), a result in agreement with the increased Arc expression during reversal learning and changes of rules (49). ...
Addiction relies on persistent alterations of neuronal properties, which depends on gene regulation. Activity-regulated cytoskeleton-associated protein (Arc) is an immediate early gene that modulates neuronal plasticity underlying learning and memory. Its role in cocaine-induced neuronal and behavioral adaptations remains elusive.
Acute cocaine-treated mice were used for Q-RT-PCR, immunocytochemistry and confocal imaging from striatum. Live imaging and transfection assays for Arc overexpression were performed from primary cultures. Molecular and behavioral adaptations to cocaine were studied from _Arc_-deficient mice and their wild-type littermates.
_Arc_ mRNA and proteins are rapidly induced in the striatum after acute cocaine administration, via an ERK-dependent _de novo_ protein synthesis. Although detected in dendrites, Arc accumulates in the nucleus in active zones of transcription where it colocalizes with phosphorylated histone-H3 (pH3), an important component of nucleosomal response. _In vitro,_ Arc overexpression down-regulates pH3 without modifying ERK phosphorylation in the nucleus_. In vivo_, _Arc_-deficient mice display decreased heterochromatin domains, a high RNA-Pol II activity and enhanced c-Fos expression. These mice presented an exacerbated psychomotor sensitization and conditioned place preference induced by low doses of cocaine.
Cocaine induces the rapid induction of Arc and its nuclear accumulation in striatal neurons. Locally, it alters the nucleosomal response, and acts as a brake on chromatin remodeling and gene regulation. These original observations posit Arc as a major homeostatic modulator of molecular and behavioral responses to cocaine. Thus, modulating Arc levels may provide promising therapeutic approaches in drug addiction.
... Interestingly, we also found reduced expression of the integral glutamatergic protein PSD-95 and the cytoskeletal protein Arc/Arg3.1, known to be sensitive to cocaine exposure (Fosnaugh et al., 1995;Fumagalli et al., 2006;Fumagalli, Franchi, et al., 2009). The peculiarity of these findings derives from the evidence that such changes were restricted to the postsynaptic density, with no effects in the homogenate, implying that cocaine does not affect the synthesis of these glutamate determinants but, rather, their synaptic retention, suggesting a functional effect of these cocaine-induced glutamatergic modifications. ...
... Structural remodelling occurs also at longer points of drug withdrawal indicating the enduring consequence set in motion by developmental cocaine exposure. Repeated exposure to cocaine during adolescence altered the expression and the mechanisms of synthesis of Arc/Arg3.1 , an effector, immediate early, gene critical for cytoskeletal plasticity and known to be engaged in structural synaptic plasticity (Bramham et al., 2008) as well as in the action of cocaine (Fosnaugh et al., 1995;Fumagalli, Franchi, et al., 2009;Hearing et al., 2008). Arc/Arg3.1 expression was significantly increased in the mPFC of rats at adulthood, through the alteration of finely tuned mechanisms that, under physiological conditions, regulate its synthesis involving, for instance, the ubiquitin-protein ligase E3A and the metabotropic glutamate mGlu 5 receptor . ...
Drug addiction is a devastating disorder with a huge economic and social burden for modern society. Although an individual may slip into drug abuse throughout his/her life, adolescents are at higher risk, but, so far, only a few studies have attempted to elucidate the underlying cellular and molecular bases of such vulnerability. Indeed, preclinical evidence indicates that psychostimulants and adolescence interact and contribute to promoting a dysfunctional brain. In this review, we have focused our attention primarily on changes in neuroplasticity brought about by cocaine, taking into account that there is much less evidence from exposure to cocaine in adolescence, compared with that from adults. This review clearly shows that exposure to cocaine during adolescence, acute or chronic, as well as contingent or non‐contingent, confers a vulnerable endophenotype, primarily, by causing changes in neuroplasticity. Given the close relationship between drug abuse and psychiatric disorders, we also discuss the translational implications providing an interpretative framework for clinical studies involving addictive as well as affective or psychotic behaviours.
LINKED ARTICLES
This article is part of a themed issue on New discoveries and perspectives in mental and pain disorders. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.17/issuetoc
... For a more thorough description of Arc induction and function, please refer to other reviews in this journal issue. The induction of Arc mRNA is known to be transient, as it dissipates within hours upon learning and administration of drugs of abuse [18][19][20][21][22][23][24][25]. Although early studies focused predominantly on Arc mRNA expression, the development of viable Arc-specific antibodies allowed researchers to correlate mRNA induction patterns to Arc protein levels [18,[26][27][28]. ...
... As expected, the rise and fall of Arc mRNA was strongly correlated with an increase and decrease in Arc protein. Following diverse behaviors, Arc protein is elevated and then rapidly declines [19,26,[29][30][31][32]. These findings indicated that a coupling of Arc induction and turnover are related to behaviors in a variety of neural circuits, and suggested that disruption of these dynamics likely results in perturbation of normal brain functions, particularly learning. ...
The activity-regulated cytoskeleton-associated protein (Arc) is a neuron-expressed activity regulated immediate early gene (IEG) product that is essential for memory consolidation and serves as a direct readout for neural activation during learning. Arc contributes to diverse forms of synaptic plasticity mediated by the trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. Notably, Arc protein expression abruptly increases and then rapidly decreases following augmented network activity. A large body of work has focused on Arc transcription and translation. Far fewer studies have explored the relevance of Arc protein stability and turnover. Here, we review recent findings on the mechanisms controlling Arc degradation and discuss its contributions to AMPA receptor trafficking and synaptic plasticity.
... Historically, Arc gene was first cloned from adult brain tissue according to its strong induction in response to electrical seizure activity (Link et al., 1995;Lyford et al., 1995). In addition to electrical activity, Arc expression in adult brain is also affected by several neuromodulators in a regional specific manner (Fosnaugh et al., 1995;Pei et al., 2000;Sanders et al., 2008;Soulé et al., 2012). The frontal cortex receives extensive DA innervation (Verney et al., 1982;Kalsbeek et al., 1988) and activation of dopamine D1-type receptors (D1R) can enhance Arc transcription in adult brain (Fosnaugh et al., 1995). ...
... In addition to electrical activity, Arc expression in adult brain is also affected by several neuromodulators in a regional specific manner (Fosnaugh et al., 1995;Pei et al., 2000;Sanders et al., 2008;Soulé et al., 2012). The frontal cortex receives extensive DA innervation (Verney et al., 1982;Kalsbeek et al., 1988) and activation of dopamine D1-type receptors (D1R) can enhance Arc transcription in adult brain (Fosnaugh et al., 1995). However, some in vitro studies have suggested that dopamine's role in Arc induction is not essential. ...
Mental functions involve coordinated activities of specific neuronal ensembles that are embedded in complex brain circuits. Aberrant neuronal ensemble dynamics is thought to form the neurobiological basis of mental disorders. A major challenge in mental health research is to identify these cellular ensembles and determine what molecular mechanisms constrain their emergence and consolidation during development and learning. Here, we provide a perspective based on recent studies that use activity-dependent gene Arc/Arg3.1 as a cellular marker to identify neuronal ensembles and a molecular probe to modulate circuit functions. These studies have demonstrated that the transcription of Arc is activated in selective groups of frontal cortical neurons in response to specific behavioral tasks. Arc expression regulates the persistent firing of individual neurons and predicts the consolidation of neuronal ensembles during repeated learning. Therefore, the Arc pathway represents a prototypical example of activity-dependent genetic feedback regulation of neuronal ensembles. The activation of this pathway in the frontal cortex starts during early postnatal development and requires dopaminergic (DA) input. Conversely, genetic disruption of Arc leads to a hypoactive mesofrontal dopamine circuit and its related cognitive deficit. This mutual interaction suggests an auto-regulatory mechanism to amplify the impact of neuromodulators and activity-regulated genes during postnatal development. Such a mechanism may contribute to the association of mutations in dopamine and Arc pathways with neurodevelopmental psychiatric disorders. As the mesofrontal dopamine circuit shows extensive activity-dependent developmental plasticity, activity-guided modulation of DA projections or Arc ensembles during development may help to repair circuit deficits related to neuropsychiatric disorders.
... Firstly, chronic administration of other inhibitors of monoamine reuptake or metabolism increase Arc expression . Secondly, Arc expression in cortical and other brain regions increases in response to the administration of direct and indirect 5-HT agonists (Pei et al., 2000;Castro et al., 2003;Tordera et al., 2003), and other evidence also suggests an stimulatory influence of noradrenaline and dopamine (Berke et al., 1998;Fosnaugh et al., 1995;Kodama et al., 1998;Cirelli and Tononi, 2000). Finally, S33005 increases extracellular levels of 5-HT in the striatum, cortex and hippocampus, noradrenaline in the cortex and hippocampus, and dopamine in the striatum (Millan et al., 2001a,b). ...
... Though a role of noradrenaline in the induction of Arc by S33005 (and venlafaxine) cannot be discounted, the potential importance of 5-HT is underlined by the increase in Arc expression in the striatum, a structure virtually devoid of noradrenaline and in which S33005 elevates extracellular levels of 5-HT but neither noradrenaline nor dopamine (Millan et al., 2001a). Thus, the mechanism by which S33005 increases striatal Arc expression may be different from that underlying the effects of other agents such as amphetamine and cocaine (Fosnaugh et al., 1995;Kodama et al., 1998;Berke et al., 1998). However, a role of dopaminergic mechanisms in the effects of S33005 and venlafaxine should not be excluded inasmuch as their chronic administration is accompanied by a modest elevation in dopamine D 2 receptor expression in this structure (Ainsworth et al., 1998;Sharp and Millan, unpublished observations). ...
Recent data show that corticolimbic expression of the effector immediate early gene Arc is up-regulated by standard antidepressant drugs. Here, we tested the effect upon Arc expression of a novel antidepressant and selective 5-hydroxytryptamine/noradrenaline reuptake inhibitor (SNRI), (−)1-(1-dimethylaminomethyl) 5-methoxybenzocyclobutan-1-yl) cyclohexanol (S33005). Arc mRNA abundance in frontal, cingulate, orbital and parietal cortices, hippocampus (CA1 pyramidal layer) and striatum was elevated in rats treated daily for 14 but not 7 days with 10 mg/kg i.p. S33005 compared to saline. Fourteen but not 7 days treatment with 10 mg/kg i.p. venlafaxine, the prototypical SNRI, also elevated Arc mRNA, but its effects were not as pronounced and detected in fewer regions, compared to S33005. Neither S33005 nor venlafaxine altered Arc mRNA after acute injection nor altered brain derived neurotrophic factor mRNA after repeated administration. These data demonstrate that sustained treatment with SNRIs increases Arc expression in corticolimbic regions, and underpin previous neurochemical and behavioural evidence that S33005 is efficacious in models predictive of antidepressant action.
... Our results showed an overexpression of Arc mRNA levels in both the DS and the VS 2 h after administration of NEPD, NEP, 4-MeAP, and pentylone. Therefore, the observed up-regulation of this IEG may induce prolonged neuroadaptations and cause alterations in the cell structure affecting the morphology of neurons (Fosnaugh et al., 1995;Fumagalli et al., 2006). Regarding other synthetic cathinones, it has been demonstrated that acute MDPV and α-PVP administration induces an up-regulation of Arc mRNA levels in the striatum (Giannotti et al., 2017;Wojcieszak et al., 2019). ...
Several new synthetic cathinones, which mimic the effect of classical psychostimulants such as cocaine or MDMA, have appeared in the global illicit drug market in the last decades. In fact, the illicit drug market is continually evolving by constantly adding small modifications to the common chemical structure of synthetic cathinones. Thus, the aim of this study was to investigate the in vitro and in vivo structure–activity relationship (SAR) of six novel synthetic cathinones currently popular as recreational drugs, pentedrone, pentylone, N-ethyl-pentedrone (NEPD), N-ethyl-pentylone (NEP), 4-methyl-pentedrone (4-MPD), and 4-methyl-ethylaminopentedrone (4-MeAP), which structurally differ in the absence or presence of different aromatic substituents and in their amino terminal group. Human embryonic kidney (HEK293) cells expressing the human isoforms of SERT and DAT were used for the uptake inhibition and release assays. Moreover, Swiss CD-1 mice were used to investigate the psychostimulant effect, rewarding properties (3, 10, and 30 mg/kg, i.p.), and the induction of immediate-early genes (IEGs), such as Arc and c-fos in the dorsal striatum (DS) and ventral striatum (VS) as well as bdnf in the medial prefrontal cortex (mPFC), of the test compounds. Our results demonstrated that all tested synthetic cathinones are potent dopamine (DA) uptake inhibitors, especially the N-ethyl analogs, while the ring-substituted cathinones tested showed higher potency as SERT inhibitors than their no ring-substituted analogs. Moreover, unlike NEP, the remaining test compounds showed clear “hybrid” properties, acting as DAT blockers but SERT substrates. Regarding the locomotion, NEP and NEPD were more efficacious (10 mg/kg) than their N-methyl analogs, which correlates with their higher potency inhibiting the DAT and an overexpression of Arc levels in the DS and VS. Furthermore, all compounds tested induced an increase in c-fos expression in the DS, except for 4-MPD, the least effective compound in inducing hyperlocomotion. Moreover, NEP induced an up-regulation of bdnf in the mPFC that correlates with its 5-HTergic properties. Finally, the present study demonstrated for the first time that NEP, 4-MPD, and 4-MeAP induce reward in mice. Altogether, this study provides valuable information about the mechanism of action and psychostimulant and rewarding properties as well as changes in the expression of IEGs related to addiction induced by novel second-generation synthetic cathinones.
... Fos is induced in the nucleus accumbens (NAc), dorsal striatum and prefrontal cortex (PFC) following treatment with cocaine, amphetamine or methamphetamine (Crombag et al., 2002;Graybiel et al., 1990;Hope et al., 1992;Moratalla et al., 1993;Norman et al., 1993;Robertson, Paul, Moratalla, & Graybiel, 1991;Rosen et al., 1994;Torres et al., 2015;Young et al., 1991). Like Fos, Arc expression is increased in the same three brain regions after both acute and chronic cocaine (Fosnaugh et al., 1995;Freeman et al., 2010;Fumagalli et al., 2006;Salery et al., 2017;Tan et al., 2000;Yuferov et al., 2003), methamphetamine (Fujiyama et al., 2003;Kodama et al., 1998) and amphetamine (Tan et al., 2000). Arc expression is also increased in the CA1 region of the hippocampus after chronic treatment with psychostimulants (Fumagalli et al., 2006;Kodama et al., 1998). ...
Paul Greengard's name is and will remain profoundly associated with Neuroscience, with brain signaling and chemical transmission, with Parkinson's and Alzheimer's diseases, with fundamental discoveries and solving paradoxes, but much less perhaps with drug discovery. This should not be mistaken as disdain. Paul in fact did contemplate developing therapeutic avenues to actually treat brain diseases much more than it is known, perhaps during his entire career, and certainly over the last two decades. As a matter of fact, he did more than contemplate it, he directly and indirectly contributed in the development of treatments for neurological diseases and disorders. Paul's impact on fundamental aspects of the brain has been so gargantuan that any other aspect of Paul's life will have difficulty to shine. It is precisely this less known aspect of Paul's career that will be covered in this review. We will discover how Paul very early on moved away from biophysics to avoid working on nuclear weapons and instead started his career in the pharmacological spheres of a large pharmaceutical company.
... Arc is considered an effector immediate early gene, a reference to its direct, activity-induced influence on cellular function, namely, removing glutamatergic AMPA receptors from synapses (Chowdhury et al., 2006), and its expression (mRNA and protein) is induced at early time points following cocaine exposure (Fosnaugh et al., 1995;Samaha et al., 2004). Arc's potential role in mediating long-term molecular and behavioral responses to cocaine has also been widely reported (Freeman et al., 2010;Hearing, Miller, et al., 2008;Salery et al., 2017;Zavala et al., 2008). ...
The fragile X mental retardation protein (FMRP), an RNA‐binding protein, regulates cocaine‐induced neuronal plasticity and is critical for the normal development of drug‐induced locomotor sensitization, as well as reward‐related learning in the conditioned place preference assay. However, it is unknown whether FMRP impacts behaviors that are used to more closely model substance use disorders. Utilizing an intravenous cocaine self‐administration (IVSA) assay in Fmr1 knockout (KO) and wild type (WT) littermate mice, we find that, despite normal acquisition and extinction learning, Fmr1 KO mice fail to make a normal upward shift in responding during dose‐response testing. Later, when given access to the original acquisition dose under increasing schedules of reinforcement (FR1, FR3, FR5), Fmr1 KO mice earn significantly fewer cocaine infusions than WT mice. Importantly, similar deficits are not present in operant conditioning using a palatable food reinforcer, indicating that our results do not represent broad learning or reward‐related deficits in Fmr1 KO mice. Additionally, we find an FMRP target, the activity‐regulated cytoskeleton‐associated protein (Arc), to be significantly reduced in synaptic cellular fractions prepared from the nucleus accumbens of Fmr1 KO, compared to WT, mice following operant tasks reinforced with cocaine, but not food. Overall, our findings suggest that FMRP facilitates adjustments in drug self‐administration behavior that generally serve to preserve reinforcement level, and combined with our similar IVSA findings in Arc KO mice may implicate Arc, along with FMRP, in behavioral shifts that occur in drug taking when drug availability is altered.
... Fos is induced in the nucleus accumbens (NAc), dorsal striatum and prefrontal cortex (PFC) following treatment with cocaine, amphetamine or methamphetamine (Crombag et al., 2002;Graybiel et al., 1990;Hope et al., 1992;Moratalla et al., 1993;Norman et al., 1993;Robertson, Paul, Moratalla, & Graybiel, 1991;Rosen et al., 1994;Torres et al., 2015;Young et al., 1991). Like Fos, Arc expression is increased in the same three brain regions after both acute and chronic cocaine (Fosnaugh et al., 1995;Freeman et al., 2010;Fumagalli et al., 2006;Salery et al., 2017;Tan et al., 2000;Yuferov et al., 2003), methamphetamine (Fujiyama et al., 2003;Kodama et al., 1998) and amphetamine (Tan et al., 2000). Arc expression is also increased in the CA1 region of the hippocampus after chronic treatment with psychostimulants (Fumagalli et al., 2006;Kodama et al., 1998). ...
Beyond their rapid rewarding effects, drugs of abuse can durably alter an individual's response to their environment as illustrated by the compulsive drug seeking and risk of relapse triggered by drug-associated stimuli. The persistence of these associations even long after cessation of drug use demonstrates the enduring mark left by drugs on brain reward circuits. However, within these circuits, neuronal populations are differently affected by drug exposure and growing evidence indicates that relatively small subsets of neurons might be involved in the encoding and expression of drug-mediated associations. The identification of sparse neuronal populations recruited in response to drug exposure has benefited greatly from the study of immediate early genes (IEGs) whose induction is critical in initiating plasticity programs in recently activated neurons. In particular, the development of technologies to manipulate IEG-expressing cells has been fundamental to implicate broadly distributed neuronal ensembles coincidently activated by either drugs or drug-associated stimuli and to then causally establish their involvement in drug responses. In this review, we summarize the literature regarding IEG regulation in different learning paradigms and addiction models to highlight their role as a marker of activity and plasticity. As the exploration of neuronal ensembles in addiction improves our understanding of drug-associated memory encoding, it also raises several questions regarding the cellular and molecular characteristics of these discrete neuronal populations as they become incorporated in drug-associated neuronal ensembles. We review recent efforts towards this goal and discuss how they will offer a more comprehensive understanding of addiction pathophysiology.
... Arc is considered an effector immediate early gene, a reference to its direct, activity-induced influence on cellular function, namely removing glutamatergic AMPA receptors from synapses (Chowdhury et al., 2006), and its expression (mRNA and protein) is induced by cocaine exposure (Fosnaugh et al., 1995;Samaha et al., 2004). Arc's potential role in mediating long term molecular and behavioral responses to cocaine has also been widely reported (Hearing et al., 2008;Zavala et al., 2008;Freeman et al., 2010;Salery et al., 2017). ...
The fragile X mental retardation protein (FMRP), an RNA-binding protein, regulates cocaine-induced neuronal plasticity and is critical for the normal development of drug-induced locomotor sensitization, as well as reward-related learning in the conditioned place preference assay. However, it is unknown whether FMRP impacts behaviors that are used to more closely model substance use disorders. Utilizing an intravenous cocaine self-administration (IVSA) assay in Fmr1 knockout (KO) and wild type (WT) littermate mice, we find that, despite normal acquisition and extinction learning, Fmr1 KO mice fail to make a normal upward shift in responding during dose-response testing. Later, with access to the original acquisition dose under increasing schedules of reinforcement (FR1, FR3, FR5), Fmr1 KO mice earn significantly fewer cocaine infusions than WT mice. Importantly, operant conditioning with a palatable food reinforcer does not show similar deficits, indicating that our results do not stem from broad learning or reward-related deficits in Fmr1 KO mice. Additionally, we find an FMRP target, the activity-regulated cytoskeleton-associated protein (Arc), to be significantly reduced in Fmr1 KO mouse synaptic fractions from the nucleus accumbens following cocaine IVSA. Overall, our findings suggest that FMRP facilitates adjustments in drug self-administration behavior that generally serve to preserve reinforcement level, and combined with our similar IVSA findings in Arc KO mice, suggest Arc as a target of FMRP to investigate in behavioral shifts that occur when drug availability is altered.
... This is composed of 595 transcripts upregulated and 374 downregu-lated by cocaine; only 59 genes were found to be in common in these 2 analyses ( Figure S2C). Among all differentially expressed transcripts identified in cocaine-treated mice, we extracted 60 highly significant genes (padj < 0.05), which represent the bona fide signature of cocaine-induced genomic response (Figure 2A) (Chandra and Lobo, 2017;Fosnaugh et al., 1995;Gonzales et al., 2019;Hope et al., 1992;McClung and Nestler, 2003). ...
Cocaine drastically elevates dopamine (DA) levels in the striatum, a brain region that is critical to the psychomotor and rewarding properties of the drug. DA signaling regulates intrastriatal circuits connecting medium spiny neurons (MSNs) with afferent fibers and interneurons. While the cocaine-mediated increase in DA signaling on MSNs is well documented, that on cholinergic interneurons (ChIs) has been more difficult to assess. Using combined pharmacological, chemogenetic, and cell-specific ablation approaches, we reveal that the D2R-dependent inhibition of acetylcholine (ACh) signaling is fundamental to cocaine-induced changes in behavior and the striatal genomic response. We show that the D2R-dependent control of striatal ChIs enables the motor, sensitized, and reinforcing properties of cocaine. This study highlights the importance of the DA- and D2R-mediated inhibitory control of ChIs activity in the normal functioning of striatal networks.
... Interestingly, in line with our previous observations in the limbic system of Roman rats (Sanna et al., 2015b(Sanna et al., , 2017b, 2019 and references therein), the differences in the markers' expression in the VTA, mPFC and Acb among DAT KO, HET and WT rats parallel the different sexual patterns and the differential changes found during sexual activity among the three rat lines in the activity of the mesolimbic dopamine and frontocortical glutamic acid neurons, which interconnect these brain areas, with the higher difference in extracellular dopamine and glutamic acid levels in the Acb shell dialysate found between DAT KO and WT rats. Since DAT KO rats lack one of the most important regulators of dopamine function at synaptic level (Kuhar et al., 1990), and thus bear permanent alterations in the processes of neuronal plasticity and tropism driven by it (Fosnaugh et al., 1995;Fasano et al., 2013;Pitchers et al., 2013;Collo et al., 2014), and DAT KO rats are also those that show the majority of differences compared to WT rats in the expression of the markers investigated in the VTA, mPFC and Acb, it is then likely that (i) these differences are due to the absence of DAT, which leads to a reduced/altered neuronal activation and synaptic function and plasticity in the above brain areas of DAT KO compared to HET and WT rats, and (ii) a causal relationship may exist among the distinct tissue expression profiles of these markers in DAT KO, HET and WT rats and the differences in behavioral sexual patterns, copulatory parameters, and dopamine and glutamic acid release in the Acb shell. Accordingly, and in line with previous studies in mice and rats with the DAT KO genotype (Fumagalli et al., 2003;Yao et al., 2004;Leo et al., 2018b), we found a lower expression of most of the markers investigated mainly in the mPFC of DAT KO vs. WT rats. ...
Central dopamine plays a key role in sexual behavior. Recently, a Dopamine Transporter knockout (DAT KO) rat has been developed, which displays several behavioral dysfunctions that have been related to increased extracellular dopamine levels and altered dopamine turnover secondary to DAT gene silencing. This prompted us to characterize the sexual behavior of these DAT KO rats and their heterozygote (HET) and wild type (WT) counterparts in classical copulatory tests with a sexually receptive female rat and to verify if and how the acquisition of sexual experience changes along five copulatory tests in these rat lines. Extracellular dopamine and glutamic acid concentrations were also measured in the dialysate obtained by intracerebral microdialysis from the nucleus accumbens (Acb) shell of DAT KO, HET and WT rats, which underwent five copulatory tests, when put in the presence of an inaccessible sexually receptive female rat and when copulation was allowed. Markers of neurotropism (BDNF, trkB), neural activation (Δ-FosB), functional (Arc and PSA-NCAM) and structural synaptic plasticity (synaptophysin, syntaxin-3, PSD-95) were also measured in the ventral tegmental area (VTA), Acb (shell and core) and medial prefrontal cortex (mPFC) by Western Blot assays. The results indicate that the sexual behavior of DAT KO vs. HET and WT rats shows peculiar differences, mainly due to a more rapid acquisition of stable sexual activity levels and to higher levels of sexual motivation and activity. These differences occurred with differential changes in dopamine and glutamic acid concentrations in Acb dialysates during sexual behavior, with lower increases of dopamine and glutamic acid in DAT KO vs. WT and HET rats, and a lower expression of the markers investigated, mainly in the mPFC, in DAT KO vs. WT rats. Together these findings confirm a key role of dopamine in sexual behavior and provide evidence that the permanently high levels of dopamine triggered by DAT gene silencing cause alterations in both the frontocortical glutamatergic neurons projecting to the Acb and VTA and in the mesolimbic dopaminergic neurons, leading to specific brain regional changes in trophic support and neuroplastic processes, which may have a role in the sexual behavior differences found among the three rat genotypes.
... IEGs are induced immediately after neuronal activation and participate in diverse functions. Arc, one of the effector IEGs, is directly involved in cellular changes at locations such as the cytoskeleton or receptors [22], while Zif268, a regulatory transcription factor, is responsible for inducing the transcription of late-response genes [23]. Most IEGs are located and synthesized in the cell soma, although the transcripts of Arc are transported to the dendrites and translated there in accordance with n-methyl-d-aspartate (NMDA) receptor activation [24,25]. ...
When vestibular function is lost, vestibular compensation works for the reacquisition of body balance. For the study of vestibular dysfunction and vestibular compensation, surgical or chemical labyrinthectomy has been performed in various animal species. In the present study, we performed chemical labyrinthectomy using arsanilic acid in mice and investigated the time course of vestibular compensation through behavioral observations and histological studies. The surgical procedures required only paracentesis and storage of 50 µL of p-arsanilic acid sodium salt solution in the tympanic cavity for 5 min. From behavioral observations, vestibular functions were worst at 2 days and recovered by 7 days after surgery. Spontaneous nystagmus appeared at 1 day after surgery with arsanilic acid and disappeared by 2 days. Histological studies revealed specific damage to the vestibular endorgans. In the ipsilateral spinal vestibular nucleus, the medial vestibular nucleus, and the contralateral prepositus hypoglossal nucleus, a substantial number of c-Fos-immunoreactive cells appeared by 1 day after surgery with arsanilic acid, with a maximum increase in number by 2 days and complete disappearance by 7 days. Taken together, these findings indicate that chemical labyrinthectomy with arsanilic acid and the subsequent observation of vestibular compensation is a useful strategy for elucidation of the molecular mechanisms underlying vestibular pathophysiologies.
... Psychostimulant exposure also affects IEG encoding effector proteins such as Arc, which is induced in the striatum downstream of D 1 R and ERK activation (121)(122)(123). Owing to its peculiar dendritic expression, the role of Arc in neuronal plasticity has been extensively described at synapses (124,125). ...
Addiction is characterized by a compulsive pattern of drug seeking and consumption and a high risk of relapse after withdrawal that are thought to result from persistent adaptations within brain reward circuits. Drugs of abuse increase dopamine (DA) concentration in these brain areas, including the striatum, which shapes an abnormal memory trace of drug consumption that virtually highjacks reward processing. Long-term neuronal adaptations of gamma-aminobutyric acidergic striatal projection neurons (SPNs) evoked by drugs of abuse are critical for the development of addiction. These neurons form two mostly segregated populations, depending on the DA receptor they express and their output projections, constituting the so-called direct (D1 receptor) and indirect (D2 receptor) SPN pathways. Both SPN subtypes receive converging glutamate inputs from limbic and cortical regions, encoding contextual and emotional information, together with DA, which mediates reward prediction and incentive values. DA differentially modulates the efficacy of glutamate synapses onto direct and indirect SPN pathways by recruiting distinct striatal signaling pathways, epigenetic and genetic responses likely involved in the transition from casual drug use to addiction. Herein we focus on recent studies that have assessed psychostimulant-induced alterations in a cell-type-specific manner, from remodeling of input projections to the characterization of specific molecular events in each SPN subtype and their impact on long-lasting behavioral adaptations. We discuss recent evidence revealing the complex and concerted action of both SPN populations on drug-induced behavioral responses, as these studies can contribute to the design of future strategies to alleviate specific behavioral components of addiction.
... We observed a reduced expression of Arc at both mRNA and protein levels in Gnal+/− rats, as well as a tendency to decreased ERK phosphorylation. Gene expression and post transcriptional regulation of Arc is controlled by several receptor signaling pathways including NMDA- (Steward and Worley, 2001;Bloomer et al., 2008), brain-derived neurotrophic factor- (Ying et al., 2002), insulin- (Kremerskothen et al., 2002), serotonin (Pei et al., 2000)-Adreno-(McIntyre, 2005 and dopamine and adenosine signaling pathways (Fosnaugh et al., 1995;Kull et al., 2000). ...
Dystonia is a neurological movement disorder characterized by sustained or intermittent involuntary muscle contractions. Loss-of-function mutations in the GNAL gene have been identified to be the cause of "isolated" dystonia DYT25. The GNAL gene encodes for the guanine nucleotide-binding protein G(olf) subunit alpha (Gαolf), which is mainly expressed in the olfactory bulb and the striatum and functions as a modulator during neurotransmission coupling with D1R and A2AR. Previously, heterozygous Gαolf -deficient mice (Gnal+/-) have been generated and showed a mild phenotype at basal condition. In contrast, homozygous deletion of Gnal in mice (Gnal-/-) resulted in a significantly reduced survival rate. In this study, using the CRISPR-Cas9 system we generated and characterized heterozygous Gnal knockout rats (Gnal+/-) with a 13 base pair deletion in the first exon of the rat Gnal splicing variant 2, a major isoform in both human and rat striatum. Gnal+/- rats showed early-onset phenotypes associated with impaired dopamine transmission, including reduction in locomotor activity, deficits in rotarod performance and an abnormal motor skill learning ability. At cellular and molecular level, we found down-regulated Arc expression, increased cell surface distribution of AMPA receptors, and the loss of D2R-dependent corticostriatal long-term depression (LTD) in Gnal+/- rats. Based on the evidence that D2R activity is normally inhibited by adenosine A2ARs, co-localized on the same population of striatal neurons, we show that blockade of A2ARs restores physiological LTD. This animal model may be a valuable tool for investigating Gαolf function and finding a suitable treatment for dystonia associated with deficient dopamine transmission.
... Arc/Arg3.1 expression is induced robustly following a number of experiences involving learning (Montag-Sallaz and Montag, 2003), including fear learning (Gouty-Colomer et al., 2016) and exposure to novel environments (Ons et al., 2004;Guzowski et al., 1999). Its expression is also induced in multiple brain regions, such as hippocampus, cortex and/or striatum, following exposure to psychostimulants (Fosnaugh et al., 1995) and other drugs of abuse, as well as following re-exposure to drug-paired contexts (Hearing et al., 2008a;Hearing et al., 2010a;Hearing et al., 2008b;Lv et al., 2015), drug self-administration training (Fumagalli et al., 2009) and reinstatement of drug seeking (Zavala et al., 2008;Ziolkowska et al., 2011;Kuntz et al., 2008;Fanous et al., 2012). However, while critical roles for Arc/Arg3.1 in fear conditioning (Ploski et al., 2008) and extinction (Onoue et al., https://doi.org/10.1016/j.pbb.2019.172818 ...
The activity-regulated cytoskeleton-associated protein (Arc, also known as Arg3.1), an immediate early gene and synaptic regulator, is upregulated following a single cocaine exposure. However, there is not much known regarding Arc/Arg3.1's potential contribution to addiction-relevant behaviors. Despite known learning and memory deficits in contextual fear and water-maze reversal learning tasks, we find that mice lacking Arc/Arg3.1 perform conditioned place preference and operant conditioning involving positive reinforcers (food and cocaine) with little-to-no impairment. Specifically, Arc/Arg3.1 KO mice show a mild impairment early in the acquisition of cocaine intravenous self-administration (IVSA). However, following normal saline-extinction, WT mice show a classic inverted-U dose-response function, while Arc/Arg3.1 KO mice fail to adjust their intake across multiple doses. Importantly, Arc/Arg3.1 KO and WT mice behave comparably on an increasing cost task (FR1-FR3; acquisition dose), providing evidence that both groups find cocaine reinforcing. Differences in individuals that drive variations in use patterns and particularly, drug intake levels, are critical as they influence the likelihood of developing dependence. Our data suggest that Arc/Arg3.1 may contribute to addiction as a regulator of drug-taking vulnerability under different drug availability conditions.
... [67][68][69] Interestingly, dopamine release in the NAc can induce Arc expression in dopamine D1-receptor-expressing neurons. 70 Taken together with our findings, it may be that dopamine-mediated induction of Arc is necessary for normal novelty behavior. When testing the role of Arc in the adult NAc for social novelty preference, we recapitulated our global Arc KO results; however, additional novel object recognition testing revealed an extension of the deficit to objects, indicating a broad role for NAc Arc in regulating novelty discrimination/preference. ...
The activity‐regulated cytoskeleton‐associated protein (Arc, also known as Arg3.1) regulates glutamatergic synapse plasticity and has been linked to neuropsychiatric illness; however, its role in behaviors associated with mood and anxiety disorders remains unclear. We find that stress upregulates Arc expression in the adult mouse nucleus accumbens (NAc) – a brain region implicated in mood and anxiety behaviors. Global Arc knockout mice have altered AMPAR‐subunit surface levels in the adult NAc, and the Arc‐deficient mice show reductions in anxiety‐like behavior, deficits in social novelty preference, and anti‐depressive‐like behavior. Viral‐mediated expression of Arc in the adult NAc of male, global Arc KO mice restores normal levels of anxiety‐like behavior in the elevated plus maze. Consistent with this finding, viral‐mediated reduction of Arc in the adult NAc reduces anxiety‐like behavior in male, but not female, mice in the elevated plus maze. NAc‐specific reduction of Arc also produced significant deficits in both object and social novelty preference tasks. Together our findings indicate that Arc is essential for regulating normal mood‐ and anxiety‐related behaviors and novelty discrimination, and that Arc’s function within the adult NAc contributes to these behavioral effects.
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... Arc is an effector immediate early gene associated with learning and memory, 13,14 triggered by exposure to drugs like cocaine, [15][16][17][18] and specifically involved in drug seeking after abstinence. [19][20][21] BDNF is a nerve growth factor well known for its critical role and dynamic expression in neural and synaptic development, 22,23 as well as its influence on various neuropsychiatric disorders, including substance use. ...
Recreational drug use peaks during adolescence. Our research with adolescent vs adult male rats, however, shows that rats taking cocaine as adolescents have lower levels of cue‐induced reinstatement of drug‐seeking than adults, despite similar levels of intravenous (i.v.) cocaine self‐administration. Lower rates of reinstatement in younger rats could be explained by higher levels of brain plasticity. Two neuroplasticity‐related genes, activity‐regulated cytoskeletal‐associated gene (Arc) and brain‐derived neurotrophic factor (Bdnf), influence cocaine self‐administration and cue‐induced reinstatement. We tested whether reinstatement of cocaine seeking correlates with expression of these genes in reinforcement‐related brain regions. Adolescent and adult male rats (postnatal day 35 or 83‐95 at start) were allowed to acquire lever‐pressing maintained by i.v. infusions of cocaine in daily 2‐h sessions over 13 days. At one of three experimental time points, rats were sacrificed and tissue collected to analyze Arc and Bdnf mRNA by in situ hybridization in the entire medial prefrontal cortex and entire nucleus accumbens, as well as relevant subregions: prelimbic cortex, infralimbic cortex, and nucleus accumbens core and shell. Despite taking similar amounts of cocaine, adolescents reinstated less than adults. Gene expression was most notable in the prelimbic cortex, was generally higher in adolescent‐onset groups, and was higher with longer abstinence. These data partially support the hypothesis that higher levels of Arc and/or Bdnf gene expression in reinforcement‐related brain regions of younger animals contribute to lower rates of extinction responding and/or reinstatement. Future studies should include mechanistic analysis of Arc, Bdnf, and their signaling pathways in age‐dependent effects of cocaine.
... In fact, at variance from the well-known interactions between mesolimbic dopaminergic neurons and c-Fos, DFosB or BDNF-trkB (cited in the dedicated sections), very little is known on the possible interactions among Arc, mesolimbic dopamine, motivation and reward. This in spite of the fact that Arc gene deletion produces phenotypes strictly related to schizophrenia (Managoè t al., 2016), and that D1 receptor agonists, D2 receptor antagonists and psychostimulants (i.e., cocaine and methamphetamine) increase Arc expression in the striatum and mPFC (Fosnaugh et al., 1995;Yamagata et al., 2000;Moro et al, 2007;Waters et al., 2014). Further studies are needed to shed light on the biological role and significance of Arc activation and expression in the brain areas relevant to rewarding behaviors and acquisition of sexual experience as well as in the differences between RLA and RHA rats in Arc expression induced by sexual activity. ...
Male Roman High- (RHA) and Low-Avoidance (RLA) rats display significant differences in sexual behavior (RHA rats exhibit higher sexual motivation and better copulatory performance than RLA rats). These differences are very evident in sexually naïve rats (which copulate with a receptive female rat for the first time), and are still present, although reduced, after five copulatory tests, when sexual experience has been acquired. Since sexual activity is a natural reward that induces neural activation and synaptic plastic changes in limbic brain areas, we studied whether the differences in sexual activity between these rat lines are accompanied by changes in the expression of markers of neural activation and plasticity, i.e., c-Fos, ΔFosB (a truncated form of FosB), Brain-Derived Neurotrophic Factor (BDNF) and its tyrosine kinase receptor B (trkB) and Activity regulated cytoskeleton-associated (Arc) protein in the ventral tegmental area (VTA), nucleus accumbens (Acb) (core and shell) and medial prefrontal cortex (mPFC) of sexually naïve and experienced RHA and RLA rats by Western Blot and/or immunohistochemistry. This study shows that these markers changed differentially in the VTA, Acb and mPFC of RHA and RLA rats, after sexual activity. In both rat lines, the changes were very evident in naïve rats, tended to disappear in experienced rats and were higher in RHA than RLA rats. These findings confirm that sexual activity induces neural activation in limbic brain areas involved in motivation and reward, leading to changes in synaptic plasticity with sexual experience acquisition, and show that these depend on the animals' genotypic/phenotypic characteristics.
... These studies lead us to suggest that activation of muscarinic cholinergic-receptors located in GAD-positive neurons of the DS can also be responsible for arc expression during retrieval. Other neurotransmitters that are released into the striatum that also modulate arc expression are glutamate (Link et al., 1995;Lyford et al., 1995;Wang et al., 2009) and dopamine (Fosnaugh et al., 1995), but the activation of their receptors in the striatum during retrieval remains to be studied. Thus, the observed increase in arc expression in the DS in the present study could be the result of the integrated activity of glutamate, dopamine and acetylcholine. ...
Similar to the hippocampus and amygdala, the dorsal striatum is involved in memory retrieval of inhibitory avoidance, a task commonly used to study memory processes. It has been reported that memory retrieval of fear conditioning regulates gene expression of arc and zif268 in the amygdala and the hippocampus, and it is surprising that only limited effort has been made to study the molecular events caused by retrieval in the striatum. To further explore the involvement of immediate early genes in retrieval, we used real-time PCR to analyze arc and zif268 transcription in dorsal striatum, dorsal hippocampus, and amygdala at different time intervals after retrieval of step-through inhibitory avoidance memory. We found that arc expression in the striatum increased 30 min after retrieval while no changes were observed in zif268 in this region. Expression of arc and zif268 also increased in the dorsal hippocampus but the changes were attributed to context re-exposure. Control procedures indicated that in the amygdala, arc and zif268 expression was not dependent on retrieval. Our data indicate that memory retrieval of inhibitory avoidance induces arc gene expression in the dorsal striatum, caused, very likely, by the instrumental component of the task. Striatal arc expression after retrieval may induce structural and functional changes in the neurons involved in this process.
... To extend our studies of c-Fos activation to other IEGs known to be responsive to acute cocaine administration (Bhat et al., 1992;Fosnaugh et al., 1995;Nestler, 2012), we This article is protected by copyright. All rights reserved. ...
Background and purpose:
The psychostimulant cocaine induces complex molecular, cellular and behavioral responses as a consequence of inhibiting presynaptic dopamine, noradrenaline and serotonin (5-HT) transporters. To elucidate 5-HT transporter (SERT)-specific contributions to cocaine action, we evaluated cocaine effects in the SERT Met172 knock-in mouse, which expresses a SERT coding substitution that eliminates high-affinity cocaine recognition.
Experimental approach:
We validated the impact of SERT Met172 on cocaine antagonism of 5-HT re-uptake using ex vivo synaptosome preparations and in vivo microdialysis. We assessed SERT-dependence of cocaine actions behaviorally through acute and chronic locomotor activation, sensitization, conditioned place preference (CPP), and oral cocaine consumption. We implemented c-Fos, quantitative RT-PCR, and RNA sequencing approaches for insights into cellular and molecular networks supporting SERT-dependent cocaine actions.
Key results:
SERT Met172 mice demonstrated functional insensitivity for cocaine at SERT. Though they displayed wildtype levels of acute cocaine-induced hyperactivity or chronic sensitization, the pattern of acute motor activation was distinct, with a bias toward thigmotaxis. CPP was increased, and a time-dependent elevation in oral cocaine consumption was observed. SERT Met172 mice displayed relatively higher levels of neuronal activation in the hippocampus, piriform cortex and prelimbic cortex (PrL), accompanied by region-dependent changes in immediate early gene (IEG) expression. Distinct SERT-dependent gene expression networks triggered by acute and chronic cocaine administration were identified, including PrL Akt and nucleus accumbens ERK1/2 signaling.
Conclusion and implications:
Our studies reveal distinct SERT contributions to cocaine action, reinforcing the possibility of targeting specific aspects of cocaine addiction by modulation of 5-HT signaling.
... The molecular mechanism underlying Arc selective endocytosis of AMPA receptors remains unknown, although it is clear that high levels of Arc accelerate the internalization of AMPARs, i.e., endocytosis of AMPARs is facilitated, whereas deletion of Arc slows AMPAR endocytosis (Chowdhury et al. 2006;Rial Verde et al. 2006). Arc gene expression is broadly responsive to neuronal activation by physiological stimuli (Lyford et al. 1995;Guzowski et al. 2000) and by dopamine-dependent mechanisms in the striatum (Fosnaugh et al. 1995;Berke et al. 1998), as well as it plays an active role in modifying long-term synaptic responses (Guzowski et al. 2000). ...
The loss of nigral dopaminergic neurons typical in Parkinson’s disease (PD) is responsible for hyperexcitability of medium spiny neurons resulting in abnormal corticostriatal glutamatergic synaptic drive. Considering the neuroprotective effect of exercise, the changes promoted by exercise on AMPA-type glutamate receptors (AMPARs), and the role of activity-regulated cytoskeleton-associated protein (Arc) in the AMPARs trafficking, we studied the impact of short and long-term treadmill exercise during evolution of the unilateral 6-hydroxy-dopamine (6-OHDA) animal model of PD. Wistar rats were divided into sedentary and exercised groups, with and without lesion by 6-OHDA and followed up to 4 months. The exercised groups were subjected to a moderate treadmill exercise 3×/week. We measured the proteins tyrosine hydroxylase (TH), Arc, GluA1, and GluA2/3 in the striatum, substantia nigra, and motor cortex. Our results showed a higher reduction of TH expression in all sedentary groups when compared to all exercised groups in striatum and substantia nigra. In general, larger changes occurred in the striatum in the first and third months after training. After 1 month of exercise, there was significant increase of GluA2/3 with concomitant reduction of GluA1 and Arc. As a balanced system, these changes were reversed in the third month, showing an increase of Arc and GluA1 and decrease of GluA2/3. Similar results for GluAs and Arc were observed in the motor cortex of the exercised animals. These modifications may be relevant for corticostriatal circuits in PD, since the exercise-dependent plasticity can modulate GluAs expression and maybe neuronal excitability.
... Increased Arc/Arg3.1 levels were preferentially found in the D1R-expressing SPNs located in striosome compartments. These results extend earlier studies showing that acute psychostimulant administration upregulates Arc/Arg3.1 transcripts and protein levels in the striatum (Fosnaugh et al., 1995;Tan et al., 2000;Klebaur et al., 2002;Fumagalli et al., 2006;Salery et al., 2016). While Arc/Arg3.1 expression is often used as a marker of neuronal activity, convergent evidence suggests that the induction of this plasticityassociated gene would sustain homeostatic responses. ...
Repeated psychostimulant exposure induces persistent gene expression modifications that contribute to enduring changes in striatal GABAergic spiny projecting neurons (SPNs). However, it remains unclear whether changes in the control of mRNA translation are required for the establishment of these durable modifications. Here we report that repeated exposure to D-amphetamine decreases global striatal mRNA translation. This effect is paralleled by an enhanced phosphorylation of the translation factors, eIF2α and eEF2, and by the concomitant increased translation of a subset of mRNAs, among which the mRNA encoding for the activity regulated cytoskeleton-associated protein, also known as activity regulated gene 3.1 (Arc/Arg3.1). The enrichment of Arc/Arg3.1 mRNA in the polysomal fraction is accompanied by a robust increase of Arc/Arg3.1 protein levels within the striatum. Immunofluorescence analysis revealed that this increase occurred preferentially in D1R-expressing SPNs localized in striosome compartments. Our results suggest that the decreased global protein synthesis following repeated exposure to D-amphetamine favors the translation of a specific subset of mRNAs in the striatum.
... Arc is enriched in neuronal dendrites, where it localizes in a distribution resembling that of F-actin (Lyford et al., 1995), interacts with cytoskeletal proteins (Fujimoto et al., 2004), AMPA receptors (Chowdhury et al., 2006) and promotes CaM kinase II-dependent neurite extension (Donai et al., 2003). It was previously demonstrated that a 6-OHDA lesion could inhibit acute cocaine-induced Arc mRNA expression in the striatum (Fosnaugh et al., 1995, Tan et al., 2000. Therefore, the reduced striatal Arc expression observed here may be involved in the inhibitory effects on motor functions and depressive-like behavior induced by α-synuclein overexpression. ...
... Moreover, transcription factors such as CREB and ΔfosB have also been associated with cocaine (19)(20)(21). Transcriptional targets have been invoked, especially for BDNF, and immediate-early genes, such as c-fos and Arc, may also play important roles (21)(22)(23). ...
Psychotropic actions of cocaine are generally thought to involve its blockade of monoamine transporters leading to increased synaptic levels of monoamines, especially dopamine. Subsequent intracellular events have been less well characterized. We describe a signaling system wherein lower behavioral stimulant doses of cocaine, as well as higher neurotoxic doses, activate a cascade wherein nitric oxide nitrosylates glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to generate a complex with the ubiquitin-E3-ligase Siah1 which translocates to the nucleus. With lower cocaine doses, nuclear GAPDH augments CREB signaling, while at higher doses p53 signaling is enhanced. The drug CGP3466B very potently blocks GAPDH nitrosylation, hindering both signaling cascades and inhibits both behavioral activating and neurotoxic effects of cocaine. This system affords potentially novel approaches to the therapy of cocaine abuse.
... As previously stated, in our experimental protocol, c-Fos expression could mainly reflect cellular reactivity to STN HFS, while Arc expression could reflect both cellular reactivity to cocaine and plasticity-related processes involved together with those induced by STN HFS. As expected (e.g., Besnard et al., 2011;Fosnaugh et al., 1995;Fumagalli et al., 2006;Samaha et al., 2004), cocaine induced a strong Arc activation in the DST, as well as in the NAC shell and core in the OFF group. In addition, no cocaine response was found in any other motor or limbic areas examined. ...
... Arc/Arg3.1 and its encoded protein are thought to play a role in activity-dependent plasticity of dendrites [7]. Arc/Arg3.1 mRNA is greatly increased by long-term potentiation (LTP)-inducing electrical stimuli [8,9]; administration of psycho-stimulant drugs such as cocaine [10], amphetamines/methamphetamines [11][12][13], and phencyclidine [14]; insulin [15], middle cerebral artery occlusion [16], electroconvulsive shock [17,18], olfactory inputs [19], mating [20], stress [21], and other stimuli that prompt neuronal activity [22,23]. The mRNA is then rapidly delivered to the dendrites [9]. ...
Activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) was originally identified in patients with seizures. It is densely distributed in the hip-pocampus and amygdala in particular. Because the expression of Arc/Arg3.1 is regulated by nerve inputs , it is thought to be an immediate early gene. As shown both in vitro and in vivo, Arc/Arg3.1 is involved in synaptic consolidation and regulates some forms of learning and memory in rats and mice [1,2]. Furthermore, a recent study suggests that Arc/Arg3.1 may play a significant role in signal transmission via AMPA-type glutamate receptors [3-5]. Therefore, we conducted a detailed analysis of fear memory in Arc/Arg3.1-deficient mice. As previously reported, the knockout animals exhibited impaired fear memory in both contextual and cued test situations. Although Arc/Arg3.1-deficient mice showed almost the same performance as wild-type littermates 4 hr after a conditioning trial, their performance was impaired in the retention test after 24 hr or longer, either with or without reconsolidation. Immunohistochemical analyses showed an abnormal density of GluR1 in the hippocampus of Arc/Arg3.1-deficient mice; however, an application of AMPA potentiator did not improve memory performance in the mutant mice. Memory impairment in Arc/Arg3.1-deficient mice is so robust that the mice provide a useful tool for developing treatments for memory impairment.
... Arc plays a very important role in learning processes, longterm potentiation and brain plasticity that demand synthesis of new mRNA and proteins. Studies of psychomotor drugs have revealed that Arc transcription is induced in striatal neurons by cocaine ( Fosnaugh et al., 1995) and amphetamine that both alter release of neurotransmitters ( Tan et al., 2000). In adult rats, Arc mRNA was upregulated after exposure to a multifarious environment in the cortex, especially in layers III and V, the hippocampus and striatum (Figure 1) and to a lower extent in the dentate gyrus ( Pinaud et al., 2001). ...
... Expression changes of genes targeted by dopaminergic and glutamatergic transmissions or genes involved in mediating transmission were also initiated during adaptation to drug exposure. Some of these genes have been relatively well analyzed such as CART (Douglass, McKinzie, & Couceyro, 1995), the Fos family (Hope et al., 1992;Nestler, 2008), CREB (Carlezon et al., 2005), Arc (Fosnaugh, Bhat, Yamagata, Worley, & Baraban, 1995), EGR1 (O'Donovan, Tourtellotte, Millbrandt, & Baraban, 1999), Homer-1 (Swanson et al., 2001), MKP-1 (Ujike, Takaki, Kodama, & Kuroda, 2002), Narp (Hyman & Malenka, 2001), NFκB (Ang et al., 2001), and CdK5 (Bibb et al., 2001). During the adaptive process, changes in the striatum take place in synergy with changes in other brain regions, particularly with changes in dopaminergic neurons in the midbrain ventral tegmental area where cocaine-induced glutamate release activates calcium-calmodulin-dependent protein kinases such as CaMKII which are involved in the process of behavioral sensitization (Fernandez-Espejo, Ramiro-Fuentes, Portavella, & Moreno-Paublete, 2008). ...
Addiction is due to changes in the structure and function of the brain, including neuronal networks and the cells that comprise them. Within cells, gene expression changes can track and help explain their altered function. Transcriptional changes induced by addictive agents are dynamic and divergent and range from signal pathway-specific perturbations to widespread molecular and cellular dysregulation that can be measured by "omic" methods and that can be used to identify new pathways. The molecular effects of addiction depend on timing of exposure or withdrawal, the stage of adaptation, the brain region, and the behavioral model, there being many models of addiction. However, the molecular neural adaptations across different drug exposures, conditions, and regions are to some extent shared and can reflect common actions on pathways relevant to addiction. Epigenetic studies of DNA methylation and histone modifications and studies of regulatory RNA networks have been informative for elucidating the mechanisms of transcriptional change in the addicted brain.
... Single or repeated cocaine exposure increases Arc/ Arg3.1 expression (Fosnaugh et al. 1995;Freeman et al. 2002), an effect that is strictly related to the activation of dopamine D1 receptors (Fumagalli et al. 2006). A role for Arc/Arg3.1 in the rat medial prefrontal cortex (mPFC) has been suggested in the associative processing of drugassociated contextual stimuli, in the extinction of cocaineseeking as well as in cue-elicited reinstatement of cocaineseeking (Fumagalli et al. 2009;Hearing et al., 2008Hearing et al., , 2011Ziolkowska et al. 2011), pointing to Arc/Arg3.1 as a critical mediator of cocaine's action. ...
Cocaine is a psychostimulant whose abuse causes a social and economic burden for our society. Most of the published literature deals with acute effects of cocaine or short-term abstinence in adult animals but much less information exists on neuroplastic changes following long-term abstinence. We have recently shown that the long-term abstinence following developmental exposure to cocaine results in increased Activity-Regulated Cytoskeletal-associated protein (Arc/Arg3.1) expression in the crude synaptosomal fraction (Giannotti et al. Int J Neuropsychopharmacology 7(4):625-634, 2014). Given that Arc/Arg3.1 localizes not only at active synapse but also in the nucleus (Okuno et al. Cell 149:886-898, 2012; Korb et al. Nat Neurosci 16:874-883 2013; Bloomer et al. Brain Res 1153:20-33 2007), we investigated Arc/Arg3.1 protein levels in the whole homogenate and the nuclear fraction of animals exposed to cocaine during adolescence. We observed the increased expression of Arc/Arg3.1 in both the fractions, suggesting that up-regulation of Arc/Arg3.1 protein may be partly due to the increased nuclear expression of Arc/Arg3.1 in the medial prefrontal cortex (mPFC) of rats sacrificed at postnatal day 90, following 48 days of abstinence. This effect seems to cause reduced Gria1 transcription. We also found reduced expression of fragile X mental retardation gene (FMR1) which normally inhibits Arc/Arg3.1 translation together with reduced expression of Ubiquitin-protein ligase E3A (Ube3a) that normally causes Arc/Arg3.1 protein degradation via ubiquitination. Further, we found increased expression of metabotropic glutamate receptor 5 (GRM5) which is also involved in the regulation of Arc/Arg3.1 expression. Taken together, our findings show that abstinence from developmental exposure to cocaine is associated with alterations in the finely tuned mechanisms that regulate Arc/Arg3.1 expression.
... The close relationship between neural activity and Arc expression has been exploited to dissect the role of different brain regions in processing behavioral experience [74][75][76][77][78][79] and in spatial memory recall 80 , as well as to monitor the recovery of polysynaptic circuits after trauma 81 . Arc is also used as a histological marker of neuronal activation in studies on subjects as diverse as Alzheimer's disease [82][83][84][85] , stroke 86 , odor processing in the olfactory bulb [87][88][89] , neural processing in avian species [90][91][92] , circadian rhythms 93 , spermiogenesis 94 , the efficacy of antidepressant therapies [95][96][97][98] , and the effects of drugs of abuse [99][100][101][102][103] and other neurotoxic insults 104,105 . Understanding how synaptic activity couples to Arc expression is critical to interpret the use of Arc as a marker. ...
... Similar to Dusp1, Arc was up-regulated in the mPFC amphetamine treated animals, but not present in the OFC where Dusp6 was down-regulated. As Arc has been used as a marker of plastic changes in the brain and is believed to be involved in producing the structural alterations in neurons that are triggered by drugs of abuse [33,34], the opposing downregulation of Dusp6 in OFC and up-regulation of Dusp1 and Arc in the mPFC may contribute to the increased dendritic complexity of the mPFC and the decrease in neuronal branching and spine density present in the OFC. ...
... The induction of multiple transcription factors by this mechanism has already been described. Other psychostimulant induced IEG products that have been described in the striatum include homer-1a, narp, arc, and many others (62,74,75). Some of the genes induced by dopamine and psychostimulants in the striatum have been hypothesized to play a role in hippocampal LTP, making it tempting to speculate that they may ultimately have a role in synaptic remodeling in the striatum (76)(77)(78)(79). ...
Addiction to alcohol, tobacco, and illegal drugs represents a substantial burden to societies worldwide. In terms of health-related outcomes, addiction results in enormous di- rect medical costs, premature mortality (tobacco alone may be responsible for 450,000 deaths yearly in the United States), and disability. In terms of broader social costs, ad- diction results in crime, negative impacts on families, de- railed lives, and personal suffering. The major categories of drugs most likely to produce addiction are psychostimulants (including cocaine and amphetamines), opiates, ethanol, nicotine, marijuana, and phencyclidine-like drugs. Under- standing the molecular and cellular actions of addictive drugs is obligatory if we are to better understand pathophys- iology and develop potent pharmacotherapies to treat addic- tion. Of course, the molecular and cellular information pre- sented in this chapter cannot be applied directly to the behavioral expression of addiction without putting it into the context of systems level neuroscience described in other chapters. Acutely, addictive drugs are both rewarding (i.e., inter- preted by the brain as intrinsically positive)and reinforcing (i.e., behaviors associated with drug use tend to be repeated). With repeated use, however, addictive drugs produce mo- lecular changes that, within a vulnerable brain, promote continued drug-taking behavior in a manner that becomes increasingly difficult to control. The central feature of ad- diction is compulsive drug use—the loss of control over the apparently voluntary acts of drug seeking and drug tak- ing. Once it has taken hold, addiction tends to follow a chronic course with periods of abstinence (that may or may not follow treatment), followed by relapse to active drug use. Even after extended periods of drug abstinence, the risk of relapse remains high. From the point of view of develop- ing treatments, a central problem in addiction research in-
... Immediate early genes: Arc is induced after cocaine-exposure 215 . Arc shows an immediate response to morphine-exposure (Fig. 3D), in addition to an up-regulation after repeated exposure (day 14). ...
Use of addictive substances like cocaine produces enduring associations between the drug experience and cues in the drug‐taking environment. In individuals with a substance use disorder (SUD) and attempting to remain abstinent, these powerful drug‐cue associations can trigger a return to active drug use, but the molecular mechanisms regulating drug‐cue associations remain poorly understood. The activity‐regulated cytoskeleton‐associated protein (Arc) is induced by cocaine in the nucleus accumbens (NAc), an important brain reward region, but Arc's NAc function in SUD‐related behaviour remains unclear. We show here that cocaine self‐administration (SA) in rats produced a significant upregulation of Arc protein in both the core and shell subregions of the NAc. Subregion‐specific Arc reduction (shRNA) in the medial NAc Shell enhanced both context‐associated and cue‐reinstated cocaine seeking, but without altering the motivation to work for cocaine, the sensitivity to the reinforcing effects of cocaine or the ability of cocaine priming to reinstate drug seeking. In contrast, we observed no effects of Arc knockdown in the NAc core on any aspect of cocaine SA, extinction or reinstated cocaine seeking, suggesting that Arc functions within the medial NAc shell, but not NAc core, to limit the strength of drug‐context and drug‐cue associations that promote cocaine‐seeking behaviour.
DNA double-strand breaks (DSBs), classified as the most harmful type of DNA damage based on the complexity of repair, lead to apoptosis or tumorigenesis. In aging, DNA damage increases and DNA repair decreases. This is exacerbated in disease, as post-mortem tissue from patients diagnosed with mild cognitive impairment (MCI) or Alzheimer’s disease (AD) show increased DSBs. A novel role for DSBs in immediate early gene (IEG) expression, learning, and memory has been suggested. Inducing neuronal activity leads to increases in DSBs and upregulation of IEGs, while increasing DSBs and inhibiting DSB repair impairs long-term memory and alters IEG expression. Consistent with this pattern, mice carrying dominant AD mutations have increased baseline DSBs, and impaired DSB repair is observed. These data suggest an adaptive role for DSBs in the central nervous system and dysregulation of DSBs and/or repair might drive age-related cognitive decline (ACD), MCI, and AD. In this review, we discuss the adaptive role of DSBs in hippocampus-dependent learning, memory, and IEG expression. We summarize IEGs, the history of DSBs, and DSBs in synaptic plasticity, aging, and AD. DSBs likely have adaptive functions in the brain, and even subtle alterations in their formation and repair could alter IEGs, learning, and memory.
Biological psychiatry is the part of psychiatry which investigates and considers mental disorders in a neurobiological perspective by reducing them to neurochemical phenomena. Psychopharmacology complements biological psychiatry by providing pharmaceutical agents for the treatment of mental disorders. The chapter describes the basic biological models of mental disorders as well as the basic principles of psychopharmacology and the basic classes of medications.
Epigenetic mechanisms have gained increasing attention as regulators of synaptic plasticity and responsiveness to drugs of abuse. In particular, it has been shown that the activity of the DNA methyltransferase 3a (Dnmt3a) mediates certain long-lasting effects of cocaine. Here we examined the role of the Dnmt isoforms, Dnmt3a1 and Dnmt3a2, within the nucleus accumbens (NAc) on transcriptional activity of immediate early genes (IEGs) and acute and long-lasting responsiveness to cocaine and cocaine conditioned cues. Using primary striatal cultures, we show that transcription of Dnmt3a2, but not that of Dnmt3a1, is activated by dopamine D1 receptor signaling and that knockdown of Dnmt3a2 using viral vector-mediated expression of Dnmt3a2-specific shRNAs impairs induction of the IEGs, Arc, FosB, and Egr2. Acute cocaine administration increases expression of Dnmt3a2 but not that of Dnmt3a1 in the NAc shell. In contrast, in the NAc core, expression of Dnmt3a1 and Dnmt3a2 was unaffected by cocaine administration. shRNA-mediated knockdown of Dnmt3a2 in vivo impairs the induction of IEGs, including Egr2 and FosB indicating that Dnmt3a2 regulates cocaine-dependent expression of plasticity genes in the rat NAc shell. Cocaine self-administration experiments in rats revealed that Dnmt3a2 regulates drug cue memories that drive reinstatement of cocaine seeking as well as incubation of this phenomenon within the NAc shell. Dnmt3a2 does not influence the primary reinforcing effects of cocaine. Thus, Dnmt3a2 mediates long-lasting cocaine cue memories within the NAc shell. Targeting Dnmt3a2 expression or function may interfere with cocaine craving and relapse.
Many of the decisions we make in our everyday lives are sequential and entail sparse rewards. While sequential decision-making has been extensively investigated in theory (e.g., by reinforcement learning models) there is no systematic experimental paradigm to test it. Here, we developed such a paradigm and investigated key components of reinforcement learning models: the eligibility trace (i.e., the memory trace of previous decision steps), the external reward, and the ability to exploit the statistics of the environment's structure (model-free vs. model-based mechanisms). We show that the eligibility trace decays not with sheer time, but rather with the number of discrete decision steps made by the participants. We further show that, unexpectedly, neither monetary rewards nor the environment's spatial regularity significantly modulate behavioral performance. Finally, we found that model-free learning algorithms describe human performance better than model-based algorithms.
Immediate–early genes (IEGs) are transiently and rapidly activated in response to various cellular stimuli. IEGs mediate diverse functions during pathophysiologic events by regulating cellular signal transduction. We investigated the temporal expression of several IEGs, including c-fos, early growth response protein-1 (Egr-1), and activity-regulated cytoskeleton-associated protein (Arc), in trimethyltin (TMT)-induced hippocampal neurodegeneration. Mice (7 weeks old, C57BL/6) administered TMT (2.6 mg/kg intraperitoneally) presented severe neurodegenerative lesions in the dentate gyrus (DG) and showed behavioral seizure activity on days 1–4 post-treatment, after which the lesions and behavior recovered spontaneously over time. c-fos, Egr-1, and Arc mRNA and protein levels significantly increased in the mouse hippocampus after TMT treatment. Immunohistochemical analysis showed that nuclear c-fos expression increased mainly in the DG, whereas nuclear Egr-1 expression was increased extensively in cornu ammonis (CA) 1, CA3, and the DG after TMT treatment. Increased Arc levels were detected in the cellular somata/dendrites of the hippocampal subregions after TMT treatment. Therefore, we suggest that increased IEGs are associated with TMT-induced pathological events in mouse hippocampus.
Widespread abuse of amphetamine and methamphetamine remains a significant global health problem. Intervention efforts to reduce its use have focused on developing treatments that reverse or mitigate the neurological effects of these drugs that support addictive behaviors. Exposing rodents to amphetamine or methamphetamine and analyzing their brain tissue for genetic responses has proven to be a valuable tool for identifying the most relevant brain circuits for targeted treatment of drug abuse. Immediate early genes (IEGs), including transcription factors that coordinate stimulus-driven effects across a range of genes and signaling pathways, have been widely studied in efforts to map the neural responses to psychoactive drugs. This chapter provides an overview of studies that have examined the effects of amphetamine and methamphetamine treatment on expression patterns of the IEGs c-fos, arc, zif/268, and bdnf to characterize the neural effects of acute and chronic drug exposure.
Immediate early gene (IEG) expression is dynamically regulated in brain neurons, in response to natural activity, and linked to neural plasticity. Because of the demonstrated importance of synaptic activity in neuronal function and development, it is anticipated that IEGs contribute to adult and developmental cortical plasticity. The IEG response includes proteins that function as transcription factors, as well as enzymes and growth factor proteins that can directly modify neuronal and synaptic function. An understanding of the mechanistic contribution of the IEG response to neuronal plasticity will provide information not only about basic developmental processes, and their disturbances in mental retardation, but may also contribute to new diagnostic and therapeutic approaches for these developmental disorders. (C) 1999 Wiley-Liss. Inc.
(±)3,4-Methylenedioxymethamphetamine (MDMA) is a relatively selective dopaminergic neurotoxin in mice. This study was designed to evaluate whether MDMA exposure affects their recognition memory and hippocampal expression of plasticity markers. Mice were administered with increasing doses of MDMA once per week for 8 weeks (three times in 1 day, every 3 h) and killed 2 weeks (2w) or 3 months (3m) later. The treatment did not modify hippocampal tryptophan hydroxylase 2, a serotonergic indicator, but induced an initial reduction in dopaminergic markers in substantia nigra, which remained stable for at least 3 months. In parallel, MDMA produced a decrease in dopamine (DA) levels in the striatum at 2w, which were restored 3 months later, suggesting dopaminergic terminal regeneration (sprouting phenomenon). Moreover, recognition memory was assessed using the object recognition test. Young (2w) and mature (3m) adult mice exhibited impaired memory after 24-h but not after just 1-h retention interval. Two weeks after the treatment, animals showed constant levels of CREB but an increase in its phosphorylated form and in c-Fos expression. Brain-derived neurotrophic factor (BDNF) and especially Arc overexpression was sustained and long-lasting. We cannot rule out the absence of MDMA injury in the hippocampus being due to the generation of BDNF. The levels of NMDAR2B, PSD-95, and synaptophysin were unaffected. In conclusion, the young mice exposed to MDMA showed increased expression of early key markers of plasticity, which sometimes remained for 3 months, and suggests hippocampal maladaptive plasticity that could explain memory deficits evidenced here.
Abstract Adenosine and the adenosine receptor antagonist, caffeine, modulate locomotor activity and striatal neuropeptide expression through interactions with the dopaminergic system by mechanisms which remain partially undetermined. We adressed this question by using quantitative immunocytochemistry and in situ hybridization, combined with retrograde tracing of striatal neurons, to characterize the mechanism(s) leading to the striatal increase in the immediate early genes (IEG), c-fos, zif-268 and arc, following a single injection of caffeine or the A1 antagonist, 1,3-dipropyl-8-cyclopentylxanthine (DPCPX). Caffeine and DPCPX induced c-fos, zif-268 and arc expression, both at mRNA and protein levels, in large proportions of striatonigral and striatopallidal neurons. The involvement of dopamine systems was evaluated by manipulations of the dopaminergic transmission. Quinpirole, a D2 agonist, almost completely blocked the caffeine-induced IEG increase in both striatopallidal and striatonigral neurons. Conversely, the lesion of the nigrostriatal pathway and the D1 antagonist SCH23390 abolished the caffeine effects in striatonigral neurons but had no or slight effect, respectively, on its action in striatopallidal neurons. These observations demonstrate that caffeine- and DPCPX-mediated IEG inductions involved different mechanisms in striatonigral and striatopallidal neurons through blockade of A1 receptors. Immediate early gene inductions result from a stimulation of dopamine release in striatonigral neurons and from activation of glutamate release and probably also acetylcholine release in striatopallidal neurons. These results also support the idea that, besides A2A receptors, adenosine acting at the A1 receptor plays pivotal functions in the basal ganglia physiology and that blockade of these receptors by specific or nonspecific antagonists, DPCPX and caffeine, may influence a broad range of neuronal functions in the striatum.
As immediate-early genes (IEGs) are thought to play a critical role in mediating stimulus-induced neural plasticity, IEG response induced by methamphetamine (METH) has been characterized to define the changes in gene expression that may underlie its long-lasting behavioral effects. Although activation of several transcription factor IEGs has been described, little is known about effector IEGs. Here, we have examined whether METH administration affects expression of an effector IEG arc (activity-regulated, cytoskeleton-associated) that encodes a protein with homology to spectrin. Using in situ hybridization, we observed that METH caused a rapid and transient dose-dependent increase in arc mRNA level in the striatum and cortex that was abolished by pretreatment with the specific dopamine D1 receptor antagonist SCH-23390 but not by an atypical neuroleptic clozapine. METH induced arc mRNA in layers IV and VI of the cortex which dopamine receptor are localized to. These results suggest that D1 receptors are coupled to activation of arc gene, which may be involved in functional or structural alterations underlying neural plasticity triggered by METH.
1.1. Ethanol and cocaine are drugs of abuse that can produce long-lived changes in behavior, including dependence.2.2. A common set of neural pathways appears to mediate the addictive actions of ethanol and cocaine.3.3. Many prominent aspects of drug dependence may be the result of alterations in intracellular signals as well as specific patterns of gene expression.4.4. For instance, changes in G proteins and cAMP, phosphorylation of proteins and induction of c-fos and zif/268 in specific drug-sensitive brain regions may represent adaptive changes in response to a drug-dependent state.5.5. The concurrent use of ethanol and cocaine is the most prevalent pattern of drug abuse in humans. However, the number of studies investigating the behavioral and molecular effects of this combination are few.6.6. Emerging evidence indicates a possible antagonistic effect of ethanol and cocaine action on transcription factor function. In addition, cocaethylene (a psychoactive metabolite derived from combined ethanol and cocaine exposure) has significant effects on gene expression as well.
Antipsychotic drugs are monoamine receptor antagonists. However, the mechanisms by which these direct actions are translated into therapeutic effects are unknown. Candidate mechanisms include receptor-mediated regulation of gene expression in target neurons. Inducible transcription factors, including certain immediate early genes (IEGs), may mediate between receptor-activated second messenger systems and expression of genes involved in the differentiated functions of neurons. We examined the specificity of induction of the IEGs c-fos and zif268 after acute administration of several antipsychotic drugs and, for comparison, the stimulant amphetamine, which has pharmacologic effects relatively opposite to those of antipsychotics. Antipsychotic drugs with potent dopamine D2 receptor antagonist properties, such as haloperidol, induced both c-fos and zif268 mRNA in the caudate-putamen; however, the atypical antipsychotic drug clozapine induced zif268 but not c-fos mRNA in that region. Similarly, haloperidol, but not clozapine, induced c-Fos-like immunoreactivity in the caudate-putamen. In contrast, both drugs induced c-Fos-like immunoreactivity in the nucleus accumbens. Like haloperidol, amphetamine induced both c-fos and zif268 mRNA in the caudate-putamen, but the anatomic patterns of induction of c-Fos-like immunoreactivity by the two drugs were dramatically different. Haloperidol and amphetamine induced AP-1 binding activity in cell extracts from the caudate-putamen, indicating that drug-induced IEG expression results in protein products that may function in the regulation of target gene expression. Thus these data demonstrate that inductions of IEG expression by haloperidol, clozapine, and amphetamine are specific, may be biologically relevant, and suggest avenues for further investigation.
The ventral tegmental area (VTA) and its dopaminergic projections appear to mediate some of the rewarding properties of opiates, cocaine, and other drugs of abuse. In a previous study, we demonstrated that chronic morphine and cocaine exert common actions on tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, in this dopaminergic brain reward region (Beitner-Johnson and Nestler, 1991). In the present study, we investigated the effects of chronic morphine and cocaine on other phosphoproteins in the VTA by back phosphorylation and two-dimensional electrophoretic analysis. It was found that a number of phosphoproteins, in addition to tyrosine hydroxylase, were regulated similarly by the two drug treatments in this brain region. Several of these morphine- and cocaine-regulated phosphoproteins were identified as neurofilament (NF) proteins. Chronic, but not acute, administration of either morphine or cocaine was found to decrease levels of the three NF proteins, NF-200 (NF-H), NF-160 (NF-M), and NF-68 (NF-L), by between 15% and 50% in the VTA by back phosphorylation, immunolabeling, and Coomassie blue staining. Such regulation of NF proteins was selective, in that no detectable changes were observed in the levels of eight other major cytoskeletal or cytoskeletal-associated proteins analyzed. Furthermore, NF levels were not altered by chronic treatment with either imipramine or haloperidol, two psychotropic drugs without reinforcing properties, or by chronic stress. Morphine and cocaine regulation of NFs showed regional specificity, as NF levels were not altered in the substantia nigra, or other parts of the brain or spinal cord, by these drug treatments. NFs are thought to function as determinants of neuronal morphology and to be associated with axonal transport. Thus, decreased NF levels in the VTA in response to chronic morphine and chronic cocaine could lead to drug-induced alterations in the structural and functional properties of this brain region, which may represent, in turn, part of a common biochemical basis of morphine and cocaine addiction and craving.
The expression of immediate-early genes of the fos/jun leucine zipper family can be regulated in striatal neurons by stimuli affecting the dopaminergic nigrostriatal system. The regulatory effects are gene specific, region specific, and striatal compartment specific. In the experiments reported here, we have explored the possibility that dopaminergic stimulation might also affect striatal expression of NGFI-A, a member of the zinc finger family of immediate-early genes. We treated healthy adult rats with amphetamine or cocaine and monitored the acute response of striatal neurons by in situ hybridization with oligonucleotide probes for NGFI-A mRNA. Both drugs evoked rapid, anatomically patterned increases in NGFI-A mRNA expression in the dorsal striatum (caudoputamen) and in the ventral striatum (nucleus accumbens, olfactory tubercle). The main response to each drug was in medium-sized neurons, known to be the projection neurons of the striatum. At every dose of amphetamine eliciting a response, the increased NGFI-A mRNA expression was preferentially concentrated in striosomes of the rostral caudoputamen, whereas cocaine at each dose given induced expression of NGFI-A mRNA in both striosomes and matrix at these striatal levels. The two indirect agonists evoked NGFI-A expression in both striatal compartments farther caudally, especially in the central and medial caudoputamen. Activation by both drugs was blocked by pretreatment with the D1-selective dopamine receptor antagonist SCH23390. These patterns of NGFI-A activation are remarkably similar to those found for Fos-like immunoreactivity following acute amphetamine and cocaine treatments, suggesting that coordinate activation of members of at least two immediate-early gene families occurs in the striatum following catecholaminergic stimulation. Such patterns of induction strongly support the view that the genomic responsiveness of the striosome and of the matrix compartments of the rostral striatum are distinct at the level of early-response gene expression. These findings raise the interesting possibility that some of the well-known effects of dopaminergic stimulation on neuropeptide and neurotransmitter expression in the striatum may reflect particular combinatorial patterns of immediate-early gene activation.
Chronic treatment of rats with cocaine leads to long-term biochemical changes in the nucleus accumbens (NAc), a brain region implicated in mediating the reinforcing effects of cocaine and other drugs of abuse. Immediate early genes (IEGs) and their protein products appear to play an important role in transducing extracellular stimuli into altered patterns of cellular gene expression and, therefore, into long-term changes in cellular functioning. We therefore examined changes in the mRNA levels for the IEGs c-fos, c-jun, fosB, junB, and zif268 in the NAc of rats treated acutely and chronically with cocaine. A single cocaine injection increased the mRNA levels of all of the IEGs examined. Following chronic cocaine treatment, however, IEG expression had returned to control levels and was not significantly increased following a further acute challenge with cocaine, suggesting desensitization in the ability of cocaine to induce these IEGs. Similarly, levels of Fos-like immunoreactivity, which are increased in the NAc by acute cocaine, were reduced to control levels in chronic cocaine-treated rats. Fos, Jun, and a number of related proteins activate or repress transcription of genes by binding to DNA response elements called AP-1 sites. As would be expected from the RNA data and immunohistochemistry, acute cocaine administration increased AP-1 binding activity in the NAc, an effect that reverted completely to control levels within 8-12 hr. In contrast, AP-1 binding activity in the NAc of animals treated chronically with cocaine remained elevated at acute levels 18 hr after the last chronic injection, a time at which c-fos and c-jun mRNA levels and Fos-like immunoreactivity had returned to control values. An additional acute cocaine challenge did not further increase AP-1 binding. The data suggest that chronic cocaine treatment leads to a persistent increase in AP-1 binding activity, which may be involved in some of the physiological and behavioral aspects of cocaine addiction.
The protooncogene c-fos produces a phosphoprotein, Fos, which regulates gene transcription processes. In neuronal systems, Fos has been proposed to couple synaptic transmission to changes in gene expression by acting in the cell nucleus in concert with other proteins to form complexes in the promoter regions of target genes. We report here that the acute administration of a single dose of the indirect-acting dopaminergic agonist cocaine increases multiple Fos proteins in rat caudate nucleus. The increase is dose-dependent and is apparent immunocytochemically at 1 hr, maximal at 2 hr, and absent 48 hr after treatment. The increase seen immunocytochemically is composed of several molecular weight species as assessed by Western blotting of proteins from isolated striatal cell nuclei. Administration of the specific dopaminergic receptor antagonists sulpiride and SCH-23390 prior to cocaine support a significant role for D1 but not for D2 receptors in mediating this effect. These data indicate that D1 dopamine receptors are linked to a cellular immediate-early gene system(s) and suggest an action of cocaine at one or more levels of gene expression via modulation of transcriptional processes in activated cells.
Amphetamine and cocaine are stimulant drugs that act on central monoaminergic neurons to produce both acute psychomotor activation and long-lasting behavioral effects including addiction and psychosis. Here we report that single doses of these drugs induce rapid expression of the nuclear proto-oncogene c-fos in the forebrain and particularly in the striatum, an extrapyramidal structure implicated in addiction and in long-term drug-induced changes in motor function. The two drugs induce strikingly different patterns of c-fos expression in the striosome-matrix compartments and limbic subdivisions of the striatum, and their effects are pharmacologically distinct, although both are sensitive to dopamine receptor blockade. We propose that differential activation of immediate-early genes by psychostimulants may be an early step in drug-specific molecular cascades contributing to acute and long-lasting psychostimulant-induced changes in behavior.
A single learning event initiates several memory processes with different time courses of retention. While short term memory involves covalent modification of pre-existing proteins, the finding that long-term memory requires the expression, during learning, of additional genes, makes it possible to analyse in molecular terms the induction and retention of long-term memory.
Neuronal activity is an essential stimulus for induction of plasticity and normal development of the CNS. We have used differential cloning techniques to identify a novel immediate-early gene (IEG) cDNA that is rapidly induced in neurons by activity in models of adult and developmental plasticity. Both the mRNA and the encoded protein are enriched in neuronal dendrites. Analysis of the deduced amino acid sequence indicates a region of homology with alpha-spectrin, and the full-length protein, prepared by in vitro transcription/translation, coprecipitates with F-actin. Confocal microscopy of the native protein in hippocampal neurons demonstrates that the IEG-encoded protein is enriched in the subplasmalemmal cortex of the cell body and dendrites and thus colocalizes with the actin cytoskeletal matrix. Accordingly, we have termed the gene and encoded protein Arc (activity-regulated cytoskeleton-associated protein). Our observations suggest that Arc may play a role in activity-dependent plasticity of dendrites.
Neuronal activity results in long term cellular changes that underlie normal brain development and synaptic plasticity. To examine the molecular basis of activity-dependent plasticity, we have used differential cloning techniques to identify genes that are rapidly induced in brain neurons by synaptic activity. Here we describe an inducible novel member of the Ras family of small GTP-binding proteins we have termed Rheb. rheb mRNA is rapidly and transiently induced in hippocampal granule cells by seizures and by NMDA-dependent synaptic activity in the long term potentiation paradigm. The predicted amino acid sequence of Rheb is most closely homologous to yeast Ras1 and human Rap2. The putative GTP binding regions are highly conserved. A bacterial fusion protein of Rheb binds GTP and exhibits intrinsic GTPase activity. Like Ha-Ras, the carboxylterminal sequence encodes a CAAX box that is predicted to signal post-translational farnesylation and to target Rheb to specific membranes. rheb mRNA is expressed at comparatively high levels in normal adult cortex as well as a number of peripheral tissues, including lung and intestine. In the developing brain, rheb mRNA is expressed at relatively high levels in embryonic day 19 cortical plate, and expression remains at stable levels throughout the remainder of prenatal and postnatal development. Its close homology with ras and its rapid inducibility by receptor-dependent synaptic activity suggest that rheb may play an important role in long term activity-dependent neuronal responses.
In cells in culture, specific stimuli induce selective patterns of immediate-early gene induction. In the present study, we tested for such selectivity of stimulated gene expression by monitoring the expression of fos/jun gene mRNAs in the striatum in rats treated in vivo with the indirect dopamine agonist cocaine. We found by Northern blot and in situ hybridization analysis that cocaine induces the coordinate expression of c-fos and jun B mRNAs in neurons of the rat's striatum. By contrast, another immediate-early gene of the leucine-zipper family, c-jun, was not induced in striatal neurons by cocaine at any time tested from 1 to 24 hr after treatment. With the same probe, we could detect the induction of c-jun mRNA (as well as that of c-fos and jun B mRNAs) in the hippocampus following administration of pentylenetetrazol. The induction of expression of c-fos and jun B was rapid and transient, with peak expression occurring at approximately 1 hr after cocaine administration, and the induction of the two genes was in similar striatal sites. These results establish that differential patterns of expression of fos/jun genes occur in striatal neurons following exposure to cocaine, a potent psychomotor stimulant. We suggest that these tissue-specific patterns of gene expression may contribute to the response specificity of striatal neurons to stimulation by monoamines including dopamine.
Recent studies have shown that dopamine receptor agonists induce expression of Fos-like immunoreactivity in rat striatal neurons. The protooncogene c-fos belongs to a family of immediate early genes that are rapidly induced in fibroblasts by growth factors. In light of previous findings that several immediate early gene mRNAs that encode proven or putative transcription factors are differentially regulated by neuronal stimulation in vivo, we have examined the effect of dopaminergic agents on mRNA levels of several such genes using in situ hybridization and northern blot analysis. d-Amphetamine (2.5-10 mg/kg i.p.) causes a rapid but transient dose-dependent increase in zif268 and jun-B mRNA levels in striatum that was abolished by striatal 6-hydroxydopamine lesions or by pretreatment with the specific D1 receptor antagonist SCH-23390 but not by specific D2 receptor antagonists. Apomorphine, a dopamine agonist that acts at both D1 and D2 receptors, and SKF-38393, a specific D1 receptor agonist, produce similar mRNA changes in rats pretreated with either 6-hydroxydopamine or reserpine, whereas LY-171,555, a specific D2 receptor agonist, has no effect. Direct dopamine agonist effects on these immediate early gene mRNA levels are also blocked by D1 but not by D2 antagonists. We observed similar, although less robust, changes in c-fos and fos-B mRNA levels. These results demonstrate that striatal D1 dopamine receptors are coupled to activation of multiple transcription factor genes, including zif268 and jun-B as well as members of the fos family.
The mechanisms by which the atypical neuroleptic clozapine produces its therapeutic effects in the treatment of schizophrenia without causing the extrapyramidal side effects that are characteristic of most antipsychotic drugs remain unclear. Recently, a single injection of the typical antipsychotic haloperidol has been shown to increase c-fos expression in the striatum [Dragunow et al. (1990) Neuroscience 37, 287–294]. C-fos is a proto-oncogene that encodes a 55,000 mol. wt phosphoprotein. Fos, which is thought to assist in the regulation of “target genes” containing an AP-1 binding site. Because a wide variety of physiological and pharmacological stimuli increase c-fos expression, it has been proposed that Fos immunohistochemistry might be useful in mapping functional pathways in the central nervous system. The present experiments examined some potential neuroanatomical differences in the actions of clozapine and haloperidol by comparing their effects on c-fos expression in the medial prefrontal cortex, nucleus accumbens, striatum and lateral septum. The effects of the selective dopamine receptor antagonists SCH 23390 (D1) and raclopride (D2) were also examined.
Transcription regulatory factors are rapidly induced in brain by a wide variety of stimuli and may be important in coordinating changes in gene expression underlying neuronal plasticity. In addition to the transient activation profile typically displayed by many of these genes, zif268 exhibits prominent basal expression in the brain that is dependent on synaptic activity. Accordingly, zif268 may play a key role in regulating neuronal gene expression induced by naturally occurring stimuli. Acute cocaine administration (15 mg/kg i.p.) produces a robust and transient induction of several transcription factors in the brain, including zif268. In the present study we report that, in contrast to the acute effects of a single dose, chronic cocaine treatment (15 mg/kg i.p., twice daily for a total of 10 injections), produces a widespread suppression of basal zif268 mRNA levels in rat forebrain between 8 and 24 hr after the last cocaine injection. This reduction is not evident after a single injection of cocaine or comparable chronic treatment with a structural analog, procaine, that does not share cocaine's uptake inhibitor properties. The suppression of basal zif268 expression may provide a mechanism for producing widespread effects of chronic cocaine administration on neuronal gene expression.
The effect of selective dopamine D2 receptor-acting drugs on striatal c-fos mRNA expression in the rat has been investigated by Northern hybridization and autoradiography to determine a possible role for c-fos in the initiation of adaptive changes in D2 receptor number by neuroleptic drugs. The neuroleptic drug haloperidol, a D2 receptor antagonist, was found to produce a rapid and transient induction of c-fos mRNA expression as compared with the expression in animals treated with saline. This induction by haloperidol was found to be dose dependent and D2 receptor mediated, inasmuch as a D2 agonist completely reversed the induction and the inactive isomer of the neuroleptic butaclamol, which does not produce an increase in D2 receptors, had no effect on c-fos mRNA expression. From these data, it can be concluded that c-fos expression in striatum is under dopamine D2 receptor-mediated inhibitory control. It is suggested that c-fos may play a role in the initiation of the increase in D2 receptor number produced by chronic neuroleptic drug treatment.
Rats injected with haloperidol, which binds to both D2 dopamine and sigma receptors or the specific D2 dopamine receptor antagonist YM 09151-2, but not the specific D1 dopamine receptor antagonist SCH 23390, showed induction of c-fos protein and c-fos-related antigens in striatal neurons. This effect of haloperidol and YM 09151-2 was inhibited by the N-methyl-D-aspartate antagonist MK801 but was not affected by 1,3-di-O-tolylguanidine, a selective sigma receptor ligand. Two different antisera were used to detect c-fos protein: one was specific for c-fos protein itself while the other recognized c-fos protein as well as c-fos protein-related antigens. In time-course immunocytochemical studies, the c-fos protein was induced maximally by 1 h and had returned to baseline by 24 h. However, c-fos protein-related antigens were induced maximally after 2 h and remained elevated for at least three days after haloperidol injection. Furthermore, the c-fos protein-specific antiserum detected two to three times fewer immunopositive striatal cells than the antiserum which detected both c-fos protein-related antigens and c-fos protein in haloperidol-treated rats. This result suggests that some striatal neurons express c-fos protein-related antigens but not c-fos protein after haloperidol injection. In some striatal sections from haloperidol-injected rats immunostained with the antiserum which recognizes both c-fos protein and c-fos protein-related antigens, there were large areas of immunopositive neurons interspersed with "areas" of striatum devoid of immunostaining. The implications of these results for theories concerning the biochemical mechanism of action of haloperidol are discussed.
Excitation of neurons results in a series of finely orchestrated responses that occur over a time frame ranging from fractions of a second to hours or days. In the short term, stimulation evokes an array of biochemical and biophysical events that represent the execution of the neurophysiological phenotype of a particular cell. These processes, which contribute to the overall behavior of a neural circuit, do not require de novo protein synthesis. In contrast, stimulation is also linked to long-term phenotypic changes that require alterations in gene expression. Thus, one or more mechanisms must exist that couple cell-surface stimuli to the transcriptional regulatory apparatus of the neuron. In this article James Morgan and Tom Curran detail a stimulus-transcription coupling cascade, involving the products of the proto-oncogenes, c-fos and c-jun, that operates in many cell types including neurons.
Actin filaments are highly concentrated in postsynaptic densities at central excitatory synapses, but their influence on postsynaptic glutamate receptors is unknown. We tested whether actin depolymerization influences NMDA channel activity in whole-cell recording on cultured hippocampal neurons. The ATP- and calcium-dependent rundown of NMDA channels was prevented when actin depolymerization was blocked by phalloidin. Rundown of AMPA/kainate receptors was unaffected by phalloidin. Cytochalasins, which enhance actin-ATP hydrolysis, induced NMDA channel rundown, whereas taxol or colchicine, which stabilize or disrupt microtubule assembly, had no effect. Protease inhibitors also had no effect. Our results suggest that calcium and ATP can influence NMDA channel activity by altering the state of actin polymerization and are consistent with a proposed model in which actin filaments compartmentalize a channel regulatory protein.
Many different types of voltage-dependent Ca2+ channels inactivate when intracellular ATP declines or intracellular Ca2+ rises. An inside-out, patch-clamp technique was applied to the Ca2+ channels of Lymnaea neurons to determine the mechanism(s) underlying these two phenomena. Although no evidence was found for a phosphorylation mechanism, agents that act on the cytoskeleton were found to alter Ca2+ channel activity. The cytoskeletal disrupters colchicine and cytochalasin B were found to speed Ca2+ channel decline in ATP, whereas the cytoskeletal stabilizers taxol and phalloidin were found to prolong Ca2+ channel activity without ATP. In addition, cytoskeletal stabilizers reduced Ca(2+)-dependent channel inactivation, suggesting that both channel metabolic dependence and Ca(2+)-dependent inactivation result from a cytoskeletal interaction.
Acute administration of cocaine increases expression of the transcription factor genes c-fos and zif268 in the striatum. This response is thought to be mediated via D1 dopamine (DA) receptors, as it is blocked by the selective D1 receptor antagonist SCH 23390. However, the directly acting D1 receptor agonists, apomorphine and SKF 38393, do not mimic cocaine's activation of these genes raising the possibility that D1 receptor activation is necessary, but not sufficient, to trigger transcription factor expression. Because cocaine blocks uptake of norepinephrine (NE) and serotonin (5-HT), as well as DA, we examined whether cocaine's ability to inhibit NE and 5-HT uptake may contribute to its induction of c-fos and zif268 expression in striatum. In examining the effects of selective monoamine uptake inhibitors, we observed that fluoxetine or citalopram, selective inhibitors of 5-HT uptake, potentiated the ability of mazindol, a DA and NE uptake inhibitor, to induce zif268 and c-fos expression, even though these 5-HT uptake inhibitors had no effect when administered alone. In contrast, the selective NE uptake inhibitor, desipramine, administered alone, or in combination with fluoxetine, did not increase expression of zif268 or c-fos. Furthermore, selective denervation of 5-HT projections by p-chloroamphetamine treatment attenuated the increase in zif268 and c-fos expression induced by cocaine in the striatum. In contrast, selective lesions of NE projections with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride failed to block cocaine's activation of these genes in the striatum. Taken together, these findings indicate that cocaine's ability to induce striatal expression of c-fos and zif268 is mediated by its effects on both the 5-HT and DA systems.
Prostaglandins play important and diverse roles in the CNS. The first step in prostaglandin synthesis involves enzymatic oxidation of arachidonic acid, which is catalyzed by prostaglandin H(PGH) synthase, also referred to as cyclooxygenase. We have cloned an inducible form of this enzyme from rat brain that is nearly identical to a murine, mitogen-inducible cyclooxygenase identified from fibroblasts. Our studies indicate that this gene, here termed COX-2, is expressed throughout the forebrain in discrete populations of neurons and is enriched in the cortex and hippocampus. Neuronal expression is rapidly and transiently induced by seizures or NMDA-dependent synaptic activity. No expression is detected in glia or vascular endothelial cells. Basal expression of COX-2 appears to be regulated by natural synaptic activity in the developing and adult brain. Both basal and induced expression of COX-2 are inhibited by glucocorticoids, consistent with COX-2 regulation in peripheral tissues. Our studies indicate that COX-2 expression may be important in regulating prostaglandin signaling in brain. The marked inducibility in neurons by synaptic stimuli suggests a role in activity-dependent plasticity.
The requirement of protein and messenger RNA synthesis for long-term memory suggests that neural activity induced by learning initiates a cascade of gene expression. Here we use differential screening to identify five immediate-early genes induced by neuronal activity. One of these is tissue-plasminogen activator (tPA), an extracellular serine protease, which is induced with different spatial patterns in the brain by three activity-dependent events: (1) convulsive seizure increases expression of tPA in the whole brain; (2) stimulation of the perforant path produces an epileptiform after-discharge that ultimately leads to kindling increases the levels of tPA throughout the hippocampus bilaterally; and (3) brief high-frequency stimulation of the perforant path that produces long-term potentiation (LTP) causes an NMDA (N-methyl-D-aspartate) receptor-mediated increase in the levels of tPA mRNA which is restricted to the granule cells of the ipsilateral dentate gyrus. As release of tPA is correlated with morphological differentiation, the increased expression of tPA may play a role in the structural changes that accompany activity-dependent plasticity.
Resubmitted manuscript received February 13 Address correspondence and reprint requests to Dr. J. M. Baraban at Department of Neuroscience
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- Research Biochemicals
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-OHDA) were from Research Biochemicals, Inc. (Natick, Resubmitted manuscript received February 13, 1995 ; accepted February 13, 1995. Address correspondence and reprint requests to Dr. J. M. Baraban at Department of Neuroscience, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205 U.S.A. The present address of Dr. R. V. Bhat is Department of Pharmacol-ogy, Cephalon, Inc., 145 Brandywine Parkway, West Chester, PA 19380, U.S.A. Abbreviations used : arc, activity-regulated, cytoskeleton-associ-ated ;
Synaptic regulation of immediate early genes in brain. Cold Spring Harbor Symposium Quant
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- A J Cole
- T H Murphy
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- J M Basban
Worley P. F., Cole A. J., Murphy T. H., Christy B. A., Nakabeppu Y., and Basban J. M. (1990) Synaptic regulation of immediate early genes in brain. Cold Spring Harbor Symposium Quant. Biol. 54, 213-223.