Involvement of DA D3 Receptors in Structural Neuroplasticity of Selected Limbic Brain Circuits: Possible Role in Treatment-Resistant Depression.

Structural neuroplasticity in the adult brain is a process involving quantitative changes of the number and size of neurons and of their dendritic arborization, axon branching, spines, and synapses. These changes can occur in specific neural circuits as adaptive response to environmental challenges, exposure to stressors, tissue damage or degeneration. Converging studies point to evidence of structural plasticity in circuits operated by glutamate, GABA, dopamine, and serotonin neurotransmitters, in concert with neurotrophic factors such as Brain Derived Neurotrophic Factor (BDNF) or Insulin Growth Factor 1 (IGF1) and a series of modulators that include circulating hormones. Intriguingly, most of these endogenous agents trigger the activation of the PI3K/Akt/mTOR and ERK1/2 intracellular pathways that, in turn, lead to the production of growth-related structural changes, enhancing protein synthesis, metabolic enzyme functions, mitogenesis for energy, and new lipid-bilayer membrane apposition. The dopamine (DA) D3 receptor has been shown to play a specific role by inducing structural plasticity of the DAergic neurons of the nigrostriatal and mesocorticolimbic circuit, where they are expressed in rodents and humans, via activation of the mTORC1 and ERK1/2 pathways. These effects are BDNF-dependent and require the recruitment of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors to allow the structural changes. Since in mood disorders, depression and anhedonia have been proposed to be associated with impaired neuroplasticity and reduced DAergic tone in brain circuits connecting prefrontal cortex, ventral striatum, amygdala, and ventral mesencephalon, activation of D3 receptors could provide a therapeutic benefit. Sustained improvements of mood and anhedonia were observed in subjects with an unsatisfactory response to serotonin uptake inhibitors (SSRI) when treated with D3-preferential D2/D3 agonists such as pramipexole and ropinirole. The recent evidence that downstream mTOR pathway activation in human mesencephalic DA neurons is also produced by ketamine, probably the most effective antidepressant currently used in subjects with treatment-resistant depression, further supports the rationale for a D3 receptor activation in mood disorders.

Synergic action of L-acetylcarnitine and L-methylfolate in Mouse Models of Stress-Related Disorders and Human iPSC-Derived Dopaminergic Neurons.

The epigenetic agents, L-acetylcarnitine (LAC) and L-methylfolate (MF) are putative candidates as add-on drugs in depression. We evaluated the effect of a combined treatment with LAC and MF in two different paradigms of chronic stress in mice and in human inducible pluripotent stem cells (hiPSCs) differentiated into dopaminergic neurons. Two groups of mice were exposed to chronic unpredictable stress (CUS) for 28 days or chronic restraint stress (CRS) for 21 day, and LAC (30 or 100 mg/kg) and/or MF (0.75 or 3 mg/kg) were administered i.p. once a day for 14 days, starting from the last week of stress. In both stress paradigms, LAC and MF acted synergistically in reducing the immobility time in the forced swim test and enhancing BDNF protein levels in the frontal cortex and hippocampus. In addition, LAC and MF acted synergistically in enhancing type-2 metabotropic glutamate receptor (mGlu2) protein levels in the hippocampus of mice exposed to CRS. Interestingly, CRS mice treated with MF showed an up-regulation of NFκB p65, which is a substrate for LAC-induced acetylation. We could also demonstrate a synergism between LAC and MF in cultured hiPSCs differentiated into dopamine neurons, by measuring dendrite length and number, and area of the cell soma after 3 days of drug exposure. These findings support the combined use of LAC and MF in the treatment of MDD and other stress-related disorders.

Corrigendum: Negative Symptoms of Schizophrenia and Dopaminergic Transmission: Translational Models and Perspectives Opened by iPSC Techniques.

[This corrects the article DOI: 10.3389/fnins.2020.00632.].

Dopaminergic neuromodulation of prefrontal cortex activity requires the NMDA receptor coagonist d-serine.

Prefrontal control of cognitive functions critically depends upon glutamatergic transmission and N-methyl D-aspartate (NMDA) receptors, the activity of which is regulated by dopamine. Yet whether the NMDA receptor coagonist d-serine is implicated in the dopamine-glutamate dialogue in the prefrontal cortex (PFC) and other brain areas remains unexplored. Here, using electrophysiological recordings, we show that d-serine is required for the fine-tuning of glutamatergic neurotransmission, neuronal excitability, and synaptic plasticity in the PFC through the actions of dopamine at D1 and D3 receptors. Using in vivo microdialysis, we show that D1 and D3 receptors exert a respective facilitatory and inhibitory influence on extracellular levels and activity of d-serine in the PFC, with actions expressed primarily via the cAMP/protein kinase A (PKA) signaling cascade. Further, using functional magnetic resonance imaging (fMRI) and behavioral assessment, we show that d-serine is required for the potentiation of cognition by D3R blockade as revealed in a test of novel object recognition memory. Collectively, these results unveil a key role for d-serine in the dopaminergic neuromodulation of glutamatergic transmission and PFC activity, findings with clear relevance to the pathogenesis and treatment of diverse brain disorders involving alterations in dopamine-glutamate cross-talk.

Pharmacology profile of F17464, a dopamine D3 receptor preferential antagonist.

F17464 (N-(3-{4-[4-(8-Oxo-8H-[1,3]-dioxolo-[4,5-g]-chromen-7-yl)-butyl]-piperazin-1-yl}-phenyl)-methanesulfonamide, hydrochloride) is a new potential antipsychotic with a unique profile. The compound exhibits high affinity for the human dopamine receptor subtype 3 (hD3) (Ki = 0.17 nM) and the serotonin receptor subtype 1a (5-HT1a) (Ki = 0.16 nM) and a >50 fold lower affinity for the human dopamine receptor subtype 2 short and long form (hD2s/l) (Ki = 8.9 and 12.1 nM, respectively). [14C]F17464 dynamic studies show a slower dissociation rate from hD3 receptor (t1/2 = 110 min) than from hD2s receptor (t1/2 = 1.4 min) and functional studies demonstrate that F17464 is a D3 receptor antagonist, 5-HT1a receptor partial agonist. In human dopaminergic neurons F17464 blocks ketamine induced morphological changes, an effect D3 receptor mediated. In vivo F17464 target engagement of both D2 and 5-HT1a receptors is demonstrated in displacement studies in the mouse brain. F17464 increases dopamine release in the rat prefrontal cortex and mouse lateral forebrain - dorsal striatum and seems to reduce the effect of MK801 on % c-fos mRNA medium expressing neurons in cortical and subcortical regions. F17464 also rescues valproate induced impairment in a rat social interaction model of autism. All the neurochemistry and behavioural effects of F17464 are observed in the dose range 0.32-2.5 mg/kg i.p. in both rats and mice. The in vitro - in vivo pharmacology profile of F17464 in preclinical models is discussed in support of a therapeutic use of the compound in schizophrenia and autism.

Negative Symptoms of Schizophrenia and Dopaminergic Transmission: Translational Models and Perspectives Opened by iPSC Techniques.

Negative symptoms (NS) represent a heterogeneous dimension of schizophrenia (SCZ), associated with a poor functional outcome. A dysregulated dopamine (DA) system, including a reduced D1 receptor activation in the prefrontal cortex, DA hypoactivity in the caudate and alterations in D3 receptor activity, seems to contribute to the pathogenesis of NS. However, failure to take into account the NS heterogeneity has slowed down progress in research on their neurobiological correlates and discoveries of new effective treatments. A better neurobiological characterization of NS is needed, and this requires objective quantification of their features that can be applied in translational models, such as animal models and human inducible pluripotent stem cells (iPSC). In this review we summarize the evidence for dopaminergic alterations relevant to NS in translational animal models focusing on dysfunctional motivation, a core aspect of NS. Among others, experiments on mutant rodents with an overexpression of DA D2 or D3 receptors and the dopamine deficient mice are discussed. In the second part we summarize the findings from recent studies using iPSC to model the pathogenesis of SCZ. By retaining the genetic background of risk genetic variants, iPSC offer the possibility to study the effect of de novo mutations or inherited polymorphisms from subgroups of patients and their response to drugs, adding an important tool for personalized psychiatry. Given the key role of DA in NS, we focus on findings of iPSC-derived DA neurons. Since implementation of iPSC-derived neurons to study the neurobiology of SCZ is a relatively recent acquisition, the available data are limited. We highlight some methodological aspects of relevance in the interpretation of in vitro testing results, including limitations and strengths, offering a critical viewpoint for the implementation of future pharmacological studies aimed to the discovery and characterization of novel treatments for NS.

The novel hybrid agonist HyNDA-1 targets the D3R-nAChR heteromeric complex in dopaminergic neurons.

In this paper, we designed, synthesized and tested a small set of three new derivatives potentially targeting the D3R-nAChR heteromer, a receptor complex recently identified and characterized as the molecular entity that, in dopaminergic neurons, mediates the neurotrophic effects of nicotine. By means of a partially rigidified spacer of variable length, we incorporated in the new compounds (1a-c) the pharmacophoric substructure of a known ß2-subunit-containing nAChR agonist (A-84543) and that of the D2/D3R agonist drug ropinirole. All the compounds retained the ability to bind with high affinity both ß2-subunit-containing nAChR and D3R. Compound 1a, renamed HyNDA-1, which is characterized by the shortest linker moiety, was the most interesting ligand. We found, in fact, that HyNDA-1 significantly modulated structural plasticity on both mice and human dopaminergic neurons, an effect strongly prevented by co-incubating this ligand with either nAChR or D3R antagonists. Moreover, the neurotrophic effects of HyNDA-1 were specifically lost by disrupting the complex with specific interfering peptides. Interestingly, by using the Bioluminescence Resonance Energy Transfer 2 (BRET2) assay in HEK-293 transfected cells, we also found that HyNDA-1 has the ability to increase the affinity of interaction between nAChR and D3R. Overall, our results indicate that the neurotrophic effects of HyNDA-1 are mediated by activation of the D3R-nAChR heteromeric complex specifically expressed on dopaminergic neurons.

Structural Plasticity Induced by Ketamine in Human Dopaminergic Neurons as Mechanism Relevant for Treatment-Resistant Depression.

The mechanisms underlying the antidepressant effects of ketamine in treatment-resistant depression are only partially understood. Reactivation of neural plasticity in prefrontal cortex has been considered critical in mediating the effects of standard antidepressants, but in treatment-resistant depression patients with severe anhedonia, other components of the affected brain circuits, for example, the dopamine system, could be involved. In a recent article in Molecular Psychiatry, we showed that ketamine induces neural plasticity in human and mouse dopaminergic neurons. Human dopaminergic neurons were differentiated from inducible pluripotent stem cells for over 60 days. Mimicking the pharmacokinetic exposures occurring in treatment-resistant depression subjects, cultures were incubated with either ketamine at 0.1 and 1 µM for 1 h or with its active metabolite (2R,6R)-hydroxynorketamine at 0.1 and 0.5 µM for up to 6 h. Three days after dosing, we observed a concentration-dependent increase in dendritic arborization and soma size. These effects were mediated by the activation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor that triggered the pathways of mammalian target of rapamycin and extracellular signal-regulated kinase via the engagement of brain-derived neurotrophic factor signaling, as previously described in rodent prefrontal cortex. Interestingly, we found that neural plasticity induced by ketamine requires functionally intact dopamine D3 receptors. These data are in keeping with our recent observation that plasticity can be induced in human dopaminergic neurons by the D3 receptor-preferential agonist pramipexole, whose effect as augmentation treatment in treatment-resistant depression has been reported. Overall, the evidence of pharmacologic response in human inducible pluripotent stem cell-derived neurons could provide complementary information to those provided by circuit-based imaging when assessing the potential response to a given augmentation treatment.

Ketamine increases the expression of GluR1 and GluR2 α-amino-3-hydroxy-5-methy-4-isoxazole propionate receptor subunits in human dopaminergic neurons differentiated from induced pluripotent stem cells.

The mechanisms underlying the prolonged antidepressant effects after a single exposure to ketamine are only partially understood. Converging findings indicate a critical role of structural neuroplasticity, recently also proposed for dopaminergic (DA) neurons known to be involved in a depression core symptom, anhedonia. We recently showed that ketamine induces dendritic outgrowth in human DA neurons differentiated in-vitro from induced pluripotent stem cells of healthy donors, a phenomenon blocked by the α-amino-3-hydroxy-5-methy-4-isoxazole propionate receptor antagonist NBQX. As changes in the expression of AMPA receptor subunits GluR1 and GluR2 were observed in neuroplasticity of rodent DA neurons, we aimed to explore this phenomenon in human DA neurons. Using specific antibodies against GluR1 and GluR2 α-amino-3-hydroxy-5-methy-4-isoxazole propionate receptor subunits, we showed that GluR1 levels were significantly higher in soma than in dendrites, whereas for GluR2, levels were significantly higher in dendrites than in soma. One hour exposure to 1 µM ketamine increased the signal of both subunits in dendrites, but only of GluR2 in soma, at 24, 48, and 72 h. Nonlinear polynomial fitting of dendritic expression indicated that the two curves were significantly different, with stronger and more sustained effects on GluR2 expression. Overall, these data support a role for GluR1 and GluR2 dendritic upregulation in driving structural plasticity in human DA neurons depending on ketamine transient exposure, indicating translationally relevant downstream mechanism possibly involved in antidepressant effects.

(2R,6R)-Hydroxynorketamine promotes dendrite outgrowth in human inducible pluripotent stem cell-derived neurons through AMPA receptor with timing and exposure compatible with ketamine infusion pharmacokinetics in humans.

The mechanisms underlying the prolonged antidepressant effects after a single infusion of ketamine are only partially understood. Ketamine's half-life of ∼2 h cannot explain antidepressant effects that last for 1 week, suggesting the triggering of long-lasting neuroplasticity. Recent human pharmacokinetics (PK) data indicate that a ketamine metabolite, (2R,6R)-hydroxynorketamine (HNK), persists in the high submicromolar range for additional 6-12 h. As in rodents HNK can induce dendrite outgrowth through AMPA receptor-mediated mechanisms, in this work, we aimed to show that HNK produces similar effects in human neurons at concentrations and exposure time compatible with human PK after ketamine infusion. Human dopaminergic neurons were differentiated in-vitro from inducible pluripotent stem cells obtained from healthy donors. Exposure to submicromolar HNK for 6 h produced dendrite outgrowth when measured 3 days after exposure. This neuroplasticity was similar to that obtained with exposure to low micromolar concentrations of ketamine for 1 or 6 h. HNK and ketamine effects were blocked by pretreatment with the AMPA receptor antagonists NBQX and GYKI 52466, and by the mammalian target of rapamycin pathway blocker rapamycin. It is reasonable to conclude that the mechanistic similarity between ketamine and HNK and their diachronic brain exposure owing to the different plasma PK observed after single therapeutic ketamine infusion should contribute to the final sustained antidepressant action.

Genetic variation in CHRNA7 and CHRFAM7A is associated with nicotine dependence and response to varenicline treatment.

The role of nicotinic acetylcholine receptors (nAChR) in nicotine dependence (ND) is well established; CHRNA7, encoding the α7 subunit, has a still uncertain role in ND, although it is implicated in a wide range of neuropsychiatric conditions. CHRFAM7A, a hybrid gene containing a partial duplication of CHRNA7, is possibly involved in modulating α7 nAChR function. The aim of this study was to investigate the role of CHRNA7 and CHRFAM7A genetic variants in ND and to test the hypothesis that α7 nAChR variation may modulate the efficacy of varenicline treatment in smoking cessation. We assessed CHRNA7 and CHRFAM7A copy number, CHRFAM7A exon 6 ∆2 bp polymorphism, and sequence variants in the CHRNA7 proximal promoter in an Italian sample of 408 treatment-seeking smokers. We conducted case-control and quantitative association analyses using two smoking measures (cigarettes per day, CPD, and Fagerström Test for Nicotine Dependence, FTND). Next, driven by the hypothesis that varenicline may exert some of its therapeutic effects through activation of α7 nAChRs, we restricted the analysis to a subgroup of 142 smokers who received varenicline treatment. The CHRNA7 promoter variant rs28531779 showed association with both smoking quantitative measures (FNTD p = 0.026, ß = 0.89, 95% CI 0.11-1.67; CPD p = 0.006, ß = 4.82 95% CI 1.42-8.22). Moreover, in the varenicline-treated subgroup we observed association of CHRFAM7A copy number with 6 months smoking abstinence (p = 0.035, OR = 3.18, 95% CI = 1.09-9.30). Thus, our study points to a possible role of genetic variation in CHRNA7 and CHRFAM7A in tobacco addiction mechanisms and response to varenicline treatment.

Alpha6-Containing Nicotinic Acetylcholine Receptors Mediate Nicotine-Induced Structural Plasticity in Mouse and Human iPSC-Derived Dopaminergic Neurons.

Midbrain dopamine (DA) neurons are considered a critical substrate for the reinforcing and sensitizing effects of nicotine and tobacco dependence. While the role of the α4 and ß2 subunit containing nicotinic acetylcholine receptors (α4ß2∗nAChRs) in mediating nicotine effects on DA release and DA neuron activity has been widely explored, less information is available on their role in the morphological adaptation of the DA system to nicotine, eventually leading to dysfunctional behaviors observed in nicotine dependence. In particular, no information is available on the role of α6∗nAChRs in nicotine-induced structural plasticity in rodents and no direct evidence exists regarding the occurrence of structural plasticity in human DA neurons exposed to nicotine. To approach this problem, we used two parallel in vitro systems, mouse primary DA neuron cultures from E12.5 embryos and human DA neurons differentiated from induced pluripotent stem cells (iPSCs) of healthy donors, identified using TH+ immunoreactivity. In both systems, nicotine 1-10 µM produced a dose-dependent increase of maximal dendrite length, number of primary dendrites, and soma size when measured after 3 days in culture. These effects were blocked by pretreatments with the α6∗nAChR antagonists α-conotoxin MII and α-conotoxin PIA, as well as by the α4ß2nAChR antagonist dihydro-ß-erythroidine (DHßE) in both mouse and human DA neurons. Nicotine was also ineffective when the primary DA neurons were obtained from null mutant mice for either the α6 subunit or both the α4 and α6 subunits of nAChR. When pregnant mice were exposed to nicotine from gestational day 15, structural plasticity was also observed in the midbrain DA neurons of postnatal day 1 offspring only in wild-type mice and not in both null mutant mice. This study confirmed the critical role of α4α6∗nAChRs in mediating nicotine-induced structural plasticity in both mouse and human DA neurons, supporting the translational relevance of neurons differentiated from human iPSCs for pharmacological studies.

Ropinirole and Pramipexole Promote Structural Plasticity in Human iPSC-Derived Dopaminergic Neurons via BDNF and mTOR Signaling.

The antiparkinsonian ropinirole and pramipexole are D3 receptor- (D3R-) preferring dopaminergic (DA) agonists used as adjunctive therapeutics for the treatment resistant depression (TRD). While the exact antidepressant mechanism of action remains uncertain, a role for D3R in the restoration of impaired neuroplasticity occurring in TRD has been proposed. Since D3R agonists are highly expressed on DA neurons in humans, we studied the effect of ropinirole and pramipexole on structural plasticity using a translational model of human-inducible pluripotent stem cells (hiPSCs). Two hiPSC clones from healthy donors were differentiated into midbrain DA neurons. Ropinirole and pramipexole produced dose-dependent increases of dendritic arborization and soma size after 3 days of culture, effects antagonized by the selective D3R antagonists SB277011-A and S33084 and by the mTOR pathway kinase inhibitors LY294002 and rapamycin. All treatments were also effective in attenuating the D3R-dependent increase of p70S6-kinase phosphorylation. Immunoneutralisation of BDNF, inhibition of TrkB receptors, and blockade of MEK-ERK signaling likewise prevented ropinirole-induced structural plasticity, suggesting a critical interaction between BDNF and D3R signaling pathways. The highly similar profiles of data acquired with DA neurons derived from two hiPSC clones underpin their reliability for characterization of pharmacological agents acting via dopaminergic mechanisms.

Ketamine effects on mammalian target of rapamycin signaling in the mouse limbic system depend on functional dopamine D3 receptors.

Ketamine is a noncompetitive glutamate N-methyl-D-aspartic acid receptor antagonist. When acutely administered to rodents, it produces a rapid antidepressant effect. There is evidence that N-methyl-D-aspartic acid receptor blockade enhances glutamatergic transmission preferentially engaging α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors leading to mTOR (mammalian target of rapamycin) pathways activation, thus resulting into downstream neuroadaptive changes in limbic structures. Recent in-vitro data on primary neuronal cultures showed that ketamine activates mTOR also in dopaminergic neurons, and this activation depends on the presence of functional dopamine D3 receptors. The aim of this work was to study the in-vivo relevance of viable D3 receptors in mediating the effects of acute ketamine administration on the mTOR downstream substrate p70 ribosomal S6 kinase (p70S6K), an obligatory substrate for mTOR. We compared the effects of single ketamine 5 mg/kg, 10 mg/kg, or vehicle injection in wild-type and D3 receptor knockout mice. Animals were killed after 60 min, and their brains were processed for p-p70S6K immunohistochemistry. Ketamine increased p70S6K phosphorylation in prefrontal cortex, nucleus accumbens core and shell, ventral tegmental area, substantia nigra, hippocampal CA1, CA2, and CA3, and basolateral amygdala of wild-type mice but not in mutant mice. Our study demonstrates that ketamine-induced p70S6K phosphorylation is dependent on viable D3R expressed in most of limbic structures.

Expansion of human midbrain floor plate progenitors from induced pluripotent stem cells increases dopaminergic neuron differentiation potential.

Human induced pluripotent stem cells (hiPSCs) are invaluable to study developmental processes and disease mechanisms particularly in the brain. hiPSCs can be differentiated into mature and functional dopaminergic (DA) neurons. Having robust protocols for the generation of differentiated DA neurons from pluripotent cells is a prerequisite for the use of hiPSCs to study disease mechanisms, for drug discovery, and eventually for cell replacement therapy. Here, we describe a protocol for generating and expanding large numbers of homogeneous midbrain floor plate progenitors (mFPPs) that retain efficient DA neurogenic potential over multiple passages and can be cryobanked. We demonstrate that expanded mFPPs have increased DA neuron potential and differentiate more efficiently and rapidly than progenitors generated by standard protocols. In addition, this novel method results in increased numbers of DA neurons that in vitro show characteristic electrophysiological properties of nigrostriatal DA neurons, produce high levels of dopamine, and integrate into host mice when grafted in vivo. Thus, we describe a robust method for producing human mesencephalic DA neurons from hiPSCs.

Pharmacological targeting of dopamine D3 receptors: Possible clinical applications of selective drugs.

Dopamine D3 receptors have been pharmacologically engaged in humans since the development of the first antipsychotics and ergot-derivative dopamine (DA) agonists, even without knowing it. These agents were generally non-selective, developed primarily to target D2 receptors. In the last 10 years the understanding of the clinical implication of D3 receptors has been progressing also due to the identification of D3 gene polymorphisms, the use of more selective PET ligands such as [(11)C]-(+)-PHNO and the learning regarding the clinical use of the D3-preferential D2/D3 agonists ropinirole and pramipexole. A new specific neuroplasticity role of D3 receptor regarding dendrite arborisation outgrowth in dopaminergic neurons was also proposed to support, at least in part, the slowing of disease observed in subjects with Parkinson׳s Disease treated with DA agonists. Similar mechanisms could be at the basis of the antidepressant-like effects observed with DA agonists when co-administered with standard of care. Severe adverse event occurring with the use of anti-parkinsonian DA agonists in predisposed subjects, i.e., impulse control disorders, are now suggested to be putatively related to overactive D3 receptors. Not surprisingly, blockade of D3 receptors was proposed as treatment for addictive disorders, a goal that could be potentially achieved by repositioning buspirone, an anxiolytic drug with D3-preferential antagonistic features, or with novel selective D3 antagonists or partial agonists currently in development for schizophrenia. At the moment ABT-925 is the only selective D3 antagonist tested in schizophrenic patients in Phase II, showing an intriguing cognitive enhancing effects supported by preclinical data. Finally, exploratory pharmacogenetic analysis suggested that ABT-925 could be effective in a subpopulation of patients with a polymorphism on the D3 receptor, opening to a possible personalised medicine approach.

Structural plasticity in mesencephalic dopaminergic neurons produced by drugs of abuse: critical role of BDNF and dopamine.

Mesencephalic dopaminergic neurons were suggested to be a critical physiopathology substrate for addiction disorders. Among neuroadaptive processes to addictive drugs, structural plasticity has attracted attention. While structural plasticity occurs at both pre- and post-synaptic levels in the mesolimbic dopaminergic system, the present review focuses only on dopaminergic neurons. Exposures to addictive drugs determine two opposite structural responses, hypothrophic plasticity produced by opioids and cannabinoids (in particular during the early withdrawal phase) and hypertrophic plasticity, mostly driven by psychostimulants and nicotine. In vitro and in vivo studies identified BDNF and extracellular dopamine as two critical factors in determining structural plasticity, the two molecules sharing similar intracellular pathways involved in cell soma and dendrite growth, the MEK-ERK1/2 and the PI3K-Akt-mTOR, via preferential activation of TrkB and dopamine D3 receptors, respectively. At present information regarding specific structural changes associated to the various stages of the addiction cycle is incomplete. Encouraging neuroimaging data in humans indirectly support the preclinical evidence of hypotrophic and hypertrophic effects, suggesting a possible differential engagement of dopamine neurons in parallel and partially converging circuits controlling motivation, stress, and emotions.

Dopamine D3 receptor ligands for drug addiction treatment: update on recent findings.

The dopamine D3 receptor is located in the limbic area and apparently mediates selective effects on motivation to take drugs and drug-seeking behaviors, so that there has been considerable interest on the possible use of D3 receptor ligands to treat drug addiction. However, only recently selective tools allowing studying this receptor have been developed. This chapter presents an overview of findings that were presented at a symposium on the conference Dopamine 2013 in Sardinia in May 2013. Novel neurobiological findings indicate that drugs of abuse can lead to significant structural plasticity in rodent brain and that this is dependent on the availability of functional dopamine D3 autoreceptor, whose activation increased phosphorylation in the ERK pathway and in the Akt/mTORC1 pathway indicating the parallel engagement of a series of intracellular signaling pathways all involved in cell growth and survival. Preclinical findings using animal models of drug-seeking behaviors confirm that D3 antagonists have a promising profile to treat drug addiction across drugs of abuse type. Imaging the D3 is now feasible in human subjects. Notably, the development of (+)-4-propyl-9-hydroxynaphthoxazine ligand used in positron emission tomography (PET) studies in humans allows to measure D3 and D2 receptors based on the area of the brain under study. This PET ligand has been used to confirm up-regulation of D3 sites in psychostimulant users and to reveal that tobacco smoking produces elevation of dopamine at the level of D3 sites. There are now novel antagonists being developed, but also old drugs such as buspirone, that are available to test the D3 hypothesis in humans. The first results of clinical investigations are now being provided. Overall, those recent findings support further exploration of D3 ligands to treat drug addiction.