Mouse Ventral Tegmental Area Research Articles

Histone Deacetylase Inhibitor Suberanilohydroxamic Acid Treatment Reverses Hyposensitivity to γ-Aminobutyric Acid in the Ventral Tegmental Area During Ethanol Withdrawal.

BackgroundThe ventral tegmental area (VTA) is important for alcohol‐related reward and reinforcement. Mouse VTA neurons are hyposensitive to γ‐aminobutyric acid (GABA) during ethanol (EtOH) withdrawal, and GABA responsiveness is normalized by in vitro treatment with histone deacetylase inhibitors (HDACi). The present study examined the effect of a systemically administered HDACi, suberanilohydroxamic acid (SAHA) on GABA sensitivity, and related molecular changes in VTA neurons during withdrawal after chronic EtOH intake in rats.MethodsSprague Dawley male adult rats were fed with Lieber‐DeCarli diet (9% EtOH or control diet) for 16 days. Experimental groups included control diet‐fed and EtOH diet‐fed (0‐ or 24‐hour withdrawal) rats treated with either SAHA or vehicle injection. Single‐unit recordings were used to measure the response of VTA neurons to GABA. Immunohistochemistry was performed to examine levels of HDAC2, acetylated histone H3 lysine 9 (acH3K9), and GABAA receptor α1 and α5 subunits in the VTA; quantitative polymerase chain reaction was performed to examine the mRNA levels of HDAC2 and GABAA receptor subunits.ResultsVTA neurons from the withdrawal group exhibited GABA hyposensitivity. In vivo SAHA treatment 2 hours before sacrifice normalized the sensitivity of VTA neurons to GABA. EtOH withdrawal was associated with increased HDAC2 and decreased acH3K9 protein levels; SAHA treatment normalized acH3K9 levels. Interestingly, no significant change was observed in the mRNA levels of HDAC2. The mRNA levels, but not protein levels, of GABAA receptor α1 and α5 subunits were increased during withdrawal.ConclusionsWithdrawal from chronic EtOH exposure results in a decrease in GABA‐mediated inhibition, and this GABA hyposensitivity is normalized by in vivo SAHA treatment. Disruption of signaling in the VTA produced by alteration of GABA neurotransmission could be 1 neuroadaptive physiological process leading to craving and relapse. These results suggest that HDACi pharmacotherapy with agents like SAHA might be an effective treatment for alcoholism.

Downregulation of thioredoxin-1 in the ventral tegmental area delays extinction of methamphetamine-induced conditioned place preference.

Drug addiction is characterized by compulsive drug use and relapse. Thioredoxin-1 is emerging as an important modulator involved in the cellular protective response against a variety of toxic stressors. Previous study has reported that thioredoxin-1 overexpression prevents the acquisition of methamphetamine-conditioned place preference. Here, we aimed to investigate the effect of thioredoxin-1 on methamphetamine-conditioned place preference extinction and the possible mechanism. (a) An extinction procedure in mice was employed to investigate the effect of thioredoxin-1 on the extinction of methamphetamine-conditioned place preference. After the acquisition of methamphetamine-conditioned place preference, mice underwent the following procedures: the injection of thioredoxin-1 small interfering RNA in the ventral tegmental area followed by the post-conditioned place preference test, four days of extinction training followed by four days of recovery after surgery. (b) The levels of thioredoxin-1, dopamine D1 receptor, tyrosine hydroxylase, phosphorylated extracellular regulated kinase, and phosphorylated cyclic adenosine monophosphate response element binding protein were examined by using Western blot analysis. Thioredoxin-1 downregulation in the ventral tegmental area delayed methamphetamine-conditioned place preference extinction. The expression of thioredoxin-1 was decreased in the ventral tegmental area of mice in control and negative groups after methamphetamine-conditioned place preference extinction, but not in the thioredoxin-1 siRNA group. The levels of dopamine D1 receptor, tyrosine hydroxylase, phosphorylated extracellular regulated kinase, and phosphorylated cyclic adenosine monophosphate response element binding protein were decreased in the ventral tegmental area, nucleus accumbens, and prefrontal cortex of mice in the control and negative groups after methamphetamine-conditioned place preference extinction, but were inversely increased in thioredoxin-1 siRNA group. The results suggest that downregulation of thioredoxin-1 in the ventral tegmental area may delay methamphetamine-conditioned place preference extinction by regulating the mesocorticolimbic dopaminergic signaling pathway.

Opposing actions of CRF-R1 and CB1 receptors on VTA-GABAergic plasticity following chronic exposure to ethanol.

Dopamine neurons in the ventral tegmental area (VTA) influence learned behaviors and neuropsychiatric diseases including addiction. The stress peptide corticotrophin-releasing factor (CRF) contributes to relapse to drug and alcohol seeking following withdrawal, although the cellular actions are poorly understood. In this study, we show that presynaptic CRF type 1 receptors (CRF-R1) potentiate GABA release onto mouse VTA dopamine neurons via a PKC-Ca2+ signaling mechanism. In naive animals, activation of CRF-R1 by bath application of CRF or ethanol enhanced GABAA inhibitory postsynaptic currents (IPSCs). Following 3 days of withdrawal from four weekly cycles of chronic intermittent ethanol (CIE) vapor exposure, spontaneous IPSC frequency was enhanced while CRF and ethanol potentiation of IPSCs was intact. However, withdrawal for 3 weeks or more was associated with reduced spontaneous IPSC frequency and diminished CRF and ethanol responses. Long-term withdrawal was also accompanied by decreased sensitivity to the CB1 receptor agonist WIN55212 as well as greatly enhanced sensitivity to the CB1 antagonist AM251. Inclusion of BAPTA in the internal recording solution restored the responsiveness to CRF or ethanol and reduced the potentiating actions of AM251. Together, these data suggest that GABAA inhibition of VTA dopamine neurons is regulated by presynaptic actions of CRF and endocannabinoids and that long-term withdrawal from CIE treatment enhances endocannabinoid-mediated inhibition, thereby suppressing CRF facilitation of GABA release. Such findings have implications for understanding the impact of chronic alcohol on stress-related, dopamine-mediated alcohol-seeking behaviors.

Nicotinamide phosphoribosyltransferase regulates cocaine reward through Sirtuin 1

Nicotinamide phosphoribosyltransferase (NAMPT), a rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis in mammals, converts nicotinamide into nicotinamide mononucleotide (NMN). NMN is subsequently converted to NAD, a component that is critical for cell energy metabolism and survival. Sirtuin 1 (SIRT1), an NAD-dependent histone deacetylase, plays an important role in mediating memory and synaptic plasticity. Here, we found that NAMPT was significantly upregulated in the ventral tegmental area (VTA) of cocaine-conditioned mice. Intraperitoneal or intra-VTA injection of FK866, a specific inhibitor of NAMPT, significantly attenuated cocaine reward. However, such effects were clearly repressed by intra-VTA expression of NAMPT or supplementation with NMN. Using 1H-nuclear magnetic resonance metabolomic analysis, we found that the content of NAD and NMN were increased in the VTA of cocaine-conditioned mice; moreover, the expression of SIRT1 was also upregulated. Interestingly, the inhibitory effect of FK866 on cocaine reward was significantly weakened in Sirt1 midbrain conditional knockout mice. Our results suggest that NAMPT-mediated NAD biosynthesis may modify cocaine behavioral effects through SIRT1. Moreover, our findings reveal that the interplay between NAD biosynthesis and SIRT1 regulation may comprise a novel regulatory pathway that responds to chronic cocaine stimuli.

Oxytocin functions as a spatiotemporal filter for excitatory synaptic inputs to VTA dopamine neurons.

The experience of rewarding or aversive stimuli is encoded by distinct afferents to dopamine (DA) neurons of the ventral tegmental area (VTA). Several neuromodulatory systems including oxytocin regulate DA neuron excitability and synaptic transmission that process socially meaningful stimuli. We and others have recently characterized oxytocinergic modulation of activity in mouse VTA DA neurons, but the mechanisms underlying oxytocinergic modulation of synaptic transmission in DA neurons remain poorly understood. Here, we find that oxytocin application or optogenetic release decrease excitatory synaptic transmission, via long lasting, presynaptic, endocannabinoid-dependent mechanisms. Oxytocin modulation of excitatory transmission alters the magnitude of short and long-term depression. We find that only some glutamatergic projections to DA neurons express CB1 receptors. Optogenetic stimulation of three major VTA inputs demonstrates that oxytocin modulation is limited to projections that show evidence of CB1R transcripts. Thus, oxytocin gates information flow into reward circuits in a temporally selective and pathway-specific manner.

CaV3.1 isoform of T-type calcium channels supports excitability of rat and mouse ventral tegmental area neurons

Recent data have implicated voltage-gated calcium channels in the regulation of the excitability of neurons within the mesolimbic reward system. While the attention of most research has centered on high voltage L-type calcium channel activity, the presence and role of the low voltage-gated T-type calcium channel (T-channels) has not been well explored. Hence, we investigated T-channel properties in the neurons of the ventral tegmental area (VTA) utilizing wild-type (WT) rats and mice, CaV3.1 knock-out (KO) mice, and TH-eGFP knock-in (KI) rats in acute horizontal brain slices of adolescent animals. In voltage-clamp experiments, we first assessed T-channel activity in WT rats with characteristic properties of voltage-dependent activation and inactivation, as well as characteristic crisscrossing patterns of macroscopic current kinetics. T-current kinetics were similar in WT mice and WT rats but T-currents were abolished in CaV3.1 KO mice. In ensuing current-clamp experiments, we observed the presence of hyperpolarization-induced rebound burst firing in a subset of neurons in WT rats, as well as dopaminergic and non-dopaminergic neurons in TH-eGFP KI rats. Following the application of a pan-selective T-channel blocker TTA-P2, rebound bursting was significantly inhibited in all tested cells. In a behavioral assessment, the acute locomotor increase induced by a MK-801 (Dizocilpine) injection in WT mice was abolished in CaV3.1 KO mice, suggesting a tangible role for 3.1 T-type channels in drug response. We conclude that pharmacological targeting of CaV3.1 isoform of T-channels may be a novel approach for the treatment of disorders of mesolimbic reward system.

Determination of neurotensin projections to the ventral tegmental area in mice

Pharmacologic treatment with the neuropeptide neurotensin (Nts) modifies motivated behaviors such as feeding, locomotor activity, and reproduction. Dopamine (DA) neurons of the ventral tegmental area (VTA) control these behaviors, and Nts directly modulates the activity of DA neurons via Nts receptor-1. While Nts sources to the VTA have been described in starlings and rats, the endogenous sources of Nts to the VTA of mice remain incompletely understood, impeding determination of which Nts circuits orchestrate specific behaviors in this model. To overcome this obstacle we injected the retrograde tracer Fluoro-Gold into the VTA of mice that express GFP in Nts neurons. Identification of GFP-Nts cells that accumulate Fluoro-Gold revealed the Nts afferents to the VTA in mice. Similar to rats, most Nts afferents to the VTA of mice arise from the medial and lateral preoptic areas (POA) and the lateral hypothalamic area (LHA), brain regions that are critical for coordination of feeding and reproduction. Additionally, the VTA receives dense input from Nts neurons in the nucleus accumbens shell (NAsh) of mice, and minor Nts projections from the amygdala and periaqueductal gray area. Collectively, our data reveal multiple populations of Nts neurons that provide direct afferents to the VTA and which may regulate specific aspects of motivated behavior. This work lays the foundation for understanding endogenous Nts actions in the VTA, and how circuit-specific Nts modulation may be useful to correct motivational and affective deficits in neuropsychiatric disease.

Effects of Mitochondrial Antioxidant SkQ1 on Biochemical and Behavioral Parameters in a Parkinsonism Model in Mice.

According to one hypothesis, Parkinson's disease pathogenesis is largely caused by dopamine catabolism that is catalyzed on mitochondrial membranes by monoamine oxidase. Reactive oxygen species are formed as a byproduct of these reactions, which can lead to mitochondrial damage followed by cell degeneration and death. In this study, we investigated the effects of administration of the mitochondrial antioxidant SkQ1 on biochemical, immunohistochemical, and behavioral parameters in a Parkinson-like condition caused by protoxin MPTP injections in C57BL/6 mice. SkQ1 administration increased dopamine quantity and decreased signs of sensory-motor deficiency as well as destruction of dopaminergic neurons in the substantia nigra and ventral tegmental area in mice with the Parkinson-like condition.

Antimanic Efficacy of a Novel Kv3 Potassium Channel Modulator.

Kv3.1 and Kv3.2 voltage-gated potassium channels are expressed on parvalbumin-positive GABAergic interneurons in corticolimbic brain regions and contribute to high-frequency neural firing. The channels are also expressed on GABAergic neurons of the basal ganglia, substantia nigra, and ventral tegmental area (VTA) where they regulate firing patterns critical for movement control, reward, and motivation. Modulation of Kv3.1 and Kv3.2 channels may therefore have potential in the treatment of disorders in which these systems have been implicated, such as bipolar disorder. Following the recent development of a potassium channel modulator, AUT1-an imidazolidinedione compound that specifically increases currents mediated by Kv3.1 and Kv3.2 channels in recombinant systems-we report that the compound is able to reverse 'manic-like' behavior in two mouse models: amphetamine-induced hyperactivity and ClockΔ19 mutants. AUT1 completely prevented amphetamine-induced hyperactivity in a dose-dependent manner, similar to the atypical antipsychotic, clozapine. Similar efficacy was observed in Kv3.2 knockout mice. In contrast, AUT1 was unable to prevent amphetamine-induced hyperactivity in mice lacking Kv3.1 channels. Notably, Kv3.1-null mice displayed baseline hyperlocomotion, reduced anxiety-like behavior, and antidepressant-like behavior. In ClockΔ19 mice, AUT1 reversed hyperactivity. Furthermore, AUT1 application modulated firing frequency and action potential properties of ClockΔ19 VTA dopamine neurons potentially through network effects. Kv3.1 protein levels in the VTA of ClockΔ19 and WT mice were unaltered by acute AUT1 treatment. Taken together, these results suggest that the modulation of Kv3.1 channels may provide a novel approach to the treatment of bipolar mania.

Morphine activation of mu opioid receptors causes disinhibition of neurons in the ventral tegmental area mediated by \u03b2-arrestin2 and c-Src

The tyrosine kinase, c-Src, participates in mu opioid receptor (MOP) mediated inhibition in sensory neurons in which β-arrestin2 (β-arr2) is implicated in its recruitment. Mice lacking β-arr2 exhibit increased sensitivity to morphine reinforcement; however, whether β-arr2 and/or c-Src participate in the actions of opioids in neurons within the reward pathway is unknown. It is also unclear whether morphine acts exclusively through MOPs, or involves delta opioid receptors (DOPs). We examined the involvement of MOPs, DOPs, β-arr2 and c-Src in the inhibition by morphine of GABAergic inhibitory postsynaptic currents (IPSCs) recorded from neurons in the mouse ventral tegmental area. Morphine inhibited spontaneous IPSC frequency, mainly through MOPs, with only a negligible effect remaining in MOP−/− neurons. However, a reduction in the inhibition by morphine for DOP−/− c.f. WT neurons and a DPDPE-induced decrease of IPSC frequency revealed a role for DOPs. The application of the c-Src inhibitor, PP2, to WT neurons also reduced inhibition by morphine, while the inactive PP3, and the MEK inhibitor, SL327, had no effect. Inhibition of IPSC frequency by morphine was also reduced in β-arr2−/− neurons in which PP2 caused no further reduction. These data suggest that inhibition of IPSCs by morphine involves a β-arr2/c-Src mediated mechanism.

Immediate-early c-fos and tyrosine hydroxylase protein expression in different brain regions in high-fat induced obesity in mice

To examine brain dopamine expression in chronic high-fat diet(HFD)-induced obese mice. Ten male mice were fed by a high-fat diet (HF:45% of calories from fat) for 12 weeks and then classified as HFD group. Ten male mice were fed a low-fat diet (LF:10% of calories from fat) and used as control group (NCD). In the 10th week, the blood of the caudal vein was collected to determine the basal blood glucose level after both groups mice were fast for 12 h. Intraperitoneal (IP) glucose tolerance test (GTT) and insulin tolerance (ITT) were performed in HFD and NCD mice in the 12th week. Animals were sacrificed after fasting for 4 hours at the 12th week. Brain tissues were processed for Fos-ir and TH-ir by immunohistochemistry. After 12 weeks of feeding, body weight was significant higher in HFD mice than that in NCD ones. During GTT and ITT, HFD mice had significantly decreased glucose tolerance and insulin tolerance at 15 min and 30 min respectively than NCD ones (P<0.05). There were higher plasma insulin concentration and leptin concentration in HFD mice than those in NCD ones (P<0.05). High fat-induced increased body weight was associated with increased cellular activation, indicated by Fos immunoreactive (ir) staining, in nucleus accumbens(NAcc), paraventricular nucleus (PVN), ventral tegmental area (VTA) and substantia nigra (SN) than those of NCD ones (P<0.05); and also significantly associated with enhanced in the number of cells labeled for tyrosine hydroxylase (TH-ir), and the number of cells co-labeled for TH-ir/Fos-ir in the VTA and SN than those of NCD ones (P<0.01). Moreover, there was significantly relationship TH-ir positive cell numbers with final body weight in VTA and SN in HFD mice (P<0.05). The results showed that chronic consumption of high-fat food was associated with plasticity-related changes in reward circuitry in mice.

Roundabout receptor 2 maintains inhibitory control of the adult midbrain.

The maintenance of excitatory and inhibitory balance in the brain is essential for its function. Here we find that the developmental axon guidance receptor Roundabout 2 (Robo2) is critical for the maintenance of inhibitory synapses in the adult ventral tegmental area (VTA), a brain region important for the production of the neurotransmitter dopamine. Following selective genetic inactivation of Robo2 in the adult VTA of mice, reduced inhibitory control results in altered neural activity patterns, enhanced phasic dopamine release, behavioral hyperactivity, associative learning deficits, and a paradoxical inversion of psychostimulant responses. These behavioral phenotypes could be phenocopied by selective inactivation of synaptic transmission from local GABAergic neurons of the VTA, demonstrating an important function for Robo2 in regulating the excitatory and inhibitory balance of the adult brain.

Brain cannabinoid receptor 2: expression, function and modulation.

Cannabis sativa (marijuana) is a fibrous flowering plant that produces an abundant variety of molecules, some with psychoactive effects. At least 4% of the world's adult population uses cannabis annually, making it one of the most frequently used illicit drugs in the world. The psychoactive effects of cannabis are mediated primarily through cannabinoid receptor (CBR) subtypes. The prevailing view is that CB1Rs are mainly expressed in the central neurons, whereas CB2Rs are predominantly expressed in peripheral immune cells. However, this traditional view has been challenged by emerging strong evidence that shows CB2Rs are moderately expressed and function in specific brain areas. New evidence has demonstrated that brain CB2Rs modulate animal drug-seeking behaviors, suggesting that these receptors may exist in brain regions that regulate drug addiction. Recently, we further confirmed that functional CB2Rs are expressed in mouse ventral tegmental area (VTA) dopamine (DA) neurons and that the activation of VTA CB2Rs reduces neuronal excitability and cocaine-seeking behavior. In addition, CB2R-mediated modulation of hippocampal CA3 neuronal excitability and network synchronization has been reported. Here, we briefly summarize recent lines of evidence showing how CB2Rs modulate function and pathophysiology in the CNS.

Midbrain dopamine neurons signal aversion in a reward-context-dependent manner.

Dopamine is thought to regulate learning from appetitive and aversive events. Here we examined how optogenetically-identified dopamine neurons in the lateral ventral tegmental area of mice respond to aversive events in different conditions. In low reward contexts, most dopamine neurons were exclusively inhibited by aversive events, and expectation reduced dopamine neurons' responses to reward and punishment. When a single odor predicted both reward and punishment, dopamine neurons' responses to that odor reflected the integrated value of both outcomes. Thus, in low reward contexts, dopamine neurons signal value prediction errors (VPEs) integrating information about both reward and aversion in a common currency. In contrast, in high reward contexts, dopamine neurons acquired a short-latency excitation to aversive events that masked their VPE signaling. Our results demonstrate the importance of considering the contexts to examine the representation in dopamine neurons and uncover different modes of dopamine signaling, each of which may be adaptive for different environments.

Oxytocin receptors are expressed on dopamine and glutamate neurons in the mouse ventral tegmental area that project to nucleus accumbens and other mesolimbic targets.

The mesolimbic dopamine (DA) circuitry determines which behaviors are positively reinforcing and therefore should be encoded in the memory to become a part of the behavioral repertoire. Natural reinforcers, like food and sex, activate this pathway, thereby increasing the likelihood of further consummatory, social, and sexual behaviors. Oxytocin (OT) has been implicated in mediating natural reward and OT-synthesizing neurons project to the ventral tegmental area (VTA) and nucleus accumbens (NAc); however, direct neuroanatomical evidence of OT regulation of DA neurons within the VTA is sparse. To phenotype OT-receptor (OTR) expressing neurons originating within the VTA, we delivered Cre-inducible adeno-associated virus that drives the expression of fluorescent marker into the VTA of male mice that had Cre-recombinase driven by OTR gene expression. OTR-expressing VTA neurons project to NAc, prefrontal cortex, the extended amygdala, and other forebrain regions but less than 10% of these OTR-expressing neurons were identified as DA neurons (defined by tyrosine hydroxylase colocalization). Instead, almost 50% of OTR-expressing cells in the VTA were glutamate (GLU) neurons, as indicated by expression of mRNA for the vesicular GLU transporter (vGluT). About one-third of OTR-expressing VTA neurons did not colocalize with either DA or GLU phenotypic markers. Thus, OTR expression by VTA neurons implicates that OT regulation of reward circuitry is more complex than a direct action on DA neurotransmission. J. Comp. Neurol. 525:1094-1108, 2017. © 2016 Wiley Periodicals, Inc.

Activation of the reward system boosts innate and adaptive immunity.

Positive expectations contribute to the clinical benefits of the placebo effect. Such positive expectations are mediated by the brain's reward system; however, it remains unknown whether and how reward system activation affects the body's physiology and, specifically, immunity. Here we show that activation of the ventral tegmental area (VTA), a key component of the reward system, strengthens immunological host defense. We used 'designer receptors exclusively activated by designer drugs' (DREADDs) to directly activate dopaminergic neurons in the mouse VTA and characterized the subsequent immune response after exposure to bacteria (Escherichia coli), using time-of-flight mass cytometry (CyTOF) and functional assays. We found an increase in innate and adaptive immune responses that were manifested by enhanced antibacterial activity of monocytes and macrophages, reduced in vivo bacterial load and a heightened T cell response in the mouse model of delayed-type hypersensitivity. By chemically ablating the sympathetic nervous system (SNS), we showed that the reward system's effects on immunity are, at least partly, mediated by the SNS. Thus, our findings establish a causal relationship between the activity of the VTA and the immune response to bacterial infection.

VTA Projection Neurons Releasing GABA and Glutamate in the Dentate Gyrus.

Both dopamine and nondopamine neurons from the ventral tegmental area (VTA) project to a variety of brain regions. Here we examine nondopaminergic neurons in the mouse VTA that send long-range projections to the hippocampus. Using a combination of retrograde tracers, optogenetic tools, and electrophysiological recordings, we show that VTA GABAergic axons make synaptic contacts in the granule cell layer of the dentate gyrus, where we can elicit small postsynaptic currents. Surprisingly, the currents displayed a partial sensitivity to both bicuculline and NBQX, suggesting that these mesohippocampal neurons corelease both GABA and glutamate. Finally, we show that this projection is functional in vivo and its stimulation reduces granule cell-firing rates under anesthesia. Altogether, the present results describe a novel connection between GABA and glutamate coreleasing of cells of the VTA and the dentate gyrus. This connection could be relevant for a variety of functions, including reward-related memory and neurogenesis.

SHANK3 controls maturation of social reward circuits in the VTA.

SummaryHaploinsufficiency of SHANK3, encoding the synapse scaffolding protein SHANK3, leads to a highly penetrant form of Autism Spectrum Disorder (ASD). How SHANK3 insufficiency affects specific neural circuits and this is related to specific ASD symptoms remains elusive. Here we used shRNA to model Shank3 insufficiency in the Ventral Tegmental Area (VTA) of mice. We identified dopamine (DA) and GABA cell-type specific changes in excitatory synapse transmission that converge to reduce DA neuron activity and generate behavioral deficits, including impaired social preference. Administration of a positive allosteric modulator of the type 1 metabotropic glutamate receptors (mGluR1) during the first postnatal week restored DA neuron excitatory synapse transmission and rescued the social preference defects, while optogenetic DA neuron stimulation was sufficient to enhance social preference. Collectively, these data reveal the contribution of impaired VTA function to social behaviors and identify mGluR1 modulation during postnatal development as a potential treatment strategy.

Neuronal calcium/calmodulin-dependent protein kinase II mediates nicotine reward in the conditioned place preference test in mice.

Several recent studies have indicated the involvement of calcium-dependent mechanisms, in particular the abundant calcium-activated kinase, calcium/calmodulin-dependent kinase II (CaMKII), in behaviors associated with nicotine dependence in mice. Behavioral and biochemical studies have shown that CaMKII is involved in acute and chronic nicotine behaviors and nicotine withdrawal; however, evidence of a role for CaMKII in nicotine reward is lacking. Thus, the goal of the current study was to examine the role of CaMKII in nicotine reward. Using pharmacological and genetic tools, we tested nicotine conditioned place preference (CPP) in C57Bl/6 mice after administration of CaMKII antagonists and in α-CaMKII wild-type (+/+) and heterozygote (±) mice. CaMKII antagonists blocked expression of nicotine CPP, and the preference score was significantly reduced in α-CaMKII ± mice compared with their +/+ counterparts. Further, we assessed CaMKII activity in the ventral tegmental area (VTA), nucleus accumbens (NAc), prefrontal cortex, and hippocampus after nicotine CPP and found significant increases in CaMKII activity in the mouse VTA and NAc that were blocked by CaMKII antagonists. The findings from this study show that CaMKII mediates nicotine reward and suggest that increases in CaMKII activity in the VTA and NAc are relevant to nicotine reward behaviors.

Long-range projection neurons of the mouse ventral tegmental area: a single-cell axon tracing analysis.

Pathways arising from the ventral tegmental area (VTA) release dopamine and other neurotransmitters during the expectation and achievement of reward, and are regarded as central links of the brain networks that create drive, pleasure, and addiction. While the global pattern of VTA projections is well-known, the actual axonal wiring of individual VTA neurons had never been investigated. Here, we labeled and analyzed the axons of 30 VTA single neurons by means of single-cell transfection with the Sindbis-pal-eGFP vector in mice. These observations were complemented with those obtained by labeling the axons of small populations of VTA cells with iontophoretic microdeposits of biotinylated dextran amine. In the single-cell labeling experiments, each entire axonal tree was reconstructed from serial sections, the length of terminal axonal arbors was estimated by stereology, and the dopaminergic phenotype was tested by double-labeling for tyrosine hydroxylase immunofluorescence. We observed two main, markedly different VTA cell morphologies: neurons with a single main axon targeting only forebrain structures (FPN cells), and neurons with multibranched axons targeting both the forebrain and the brainstem (F + BSPN cells). Dopaminergic phenotype was observed in FPN cells. Moreover, four “subtypes” could be distinguished among the FPN cells based on their projection targets: (1) “Mesocorticolimbic” FPN projecting to both neocortex and basal forebrain; (2) “Mesocortical” FPN innervating the neocortex almost exclusively; (3) “Mesolimbic” FPN projecting to the basal forebrain, accumbens and caudateputamen; and (4) “Mesostriatal” FPN targeting only the caudateputamen. While the F + BSPN cells were scattered within VTA, the mesolimbic neurons were abundant in the paranigral nucleus. The observed diversity in wiring architectures is consistent with the notion that different VTA cell subpopulations modulate the activity of specific sets of prosencephalic and brainstem structures.