Different Effects of Opiate Withdrawal on Dopamine Turnover,Uptake, and Release in the Striatum and Nucleus Accumbens

The purpose of these experiments was to further characterize changes in dopaminergic function that follow withdrawal from chronic opiate treatment. Withdrawal after treatment to a maximum dose of 120 mg/kg of morphine did not alter dopamine concentrations in the substantia nigra, ventral tegmental area, striatum, or nucleus accumbens; but did decrease concentrations of DOPAC and the ratio of DOPAC to dopamine in the lateral striatum and nucleus accumbens. Uptake of tritiated dopamine was diminished for withdrawn slices obtained from the striatum with no effect observed for tissue from the nucleus accumbens. Deficits of in vitro release of tritiated dopamine also occurred following withdrawal, with the nucleus accumbens being sensitive to dependence produced by a lower dose of morphine. In conclusion, opiate withdrawal produces a complex pattern of effects on dopaminergic function that is specific for the striatum and nucleus accumbens.     

Heterogeneous Distribution of Kir3 Potassium Channel Proteins Within Dopaminergic Neurons in the Mesencephalon of the Rat Brain

1. Dopaminergic neurons in the substantia nigra pars compacta (SNc) and the ventral tegmental area (VTA) of the ventral mesencephalon play an important role in the regulation of the parallel basal ganglia loops. 2. We have raised affinity-purified polyclonal rabbit antibodies specific for all four members of the Kir3 family of inwardly rectifying potassium channels (Kir3.1–Kir3.4) to investigate the distribution of the channel proteins in the dopaminergic neurons of the rat mesencephalon at light and electron microscopic level. In addition, immunocytochemical double labeling with tyrosine hydroxylase (TH), a marker of dopaminergic neurons, were performed. 3. All Kir3 channels were present in this region. However, the individual proteins showed differential cellular and subcellular distributions. 4. Kir3.1 immunoreactivity was found in SNc fibers and some neurons of the substantia nigra pars reticulata (SNr). Few Kir3.3-positive neurons were found in the SNc. However, a strong Kir3.3 signal was identified in the SNr neuropil. Weak Kir3.4 staining was detected in neuronal somata as well as in dendritic fibers of both parts of the SN. 5. In the VTA, Kir3.1, Kir3.3, and Kir3.4 showed only weak staining of neuropil structures. The distribution of the Kir3.2 channel protein was especially striking with strong labeling in the SNc and in the lateral but not central VTA. 6. Our results suggest that the heterogeneously distributed Kir3.2 channel proteins could help to discriminate the dopaminergic neurons of VTA and SNc.     

Ventral Tegmental Dopamine Neurons Control the Impulse Vector during Motivated Behavior

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Differential expression of tyrosine hydroxylase mRNA in the developing rat mesencephalon

1. With respect to the mesostriatal projection, the mesencephalon is composed of two dopaminergic (DA) cell populations, called dorsal tier and ventral tier. Strong evidence suggests differences in both the spatial and the temporal sequence of the innervation of the striatum between the two groups, with the ventral tier neurons innervating striatal patches prenatally and dorsal tier cells innervating striatal matrix postnatally. 2. Using in situ hybridization, we have examined the expression of the gene coding for tyrosine hydroxylase (TH) in mesencephalic DA neurons with respect to their postnatal development. Two ontogenic patterns of expression were observed: (a) dorsal tier neurons of the medial mesencephalon exhibited a sharp increase in expression beginning after birth, peaking on day 14, then decreasing and, finally, stabilizing; and (b) ventral tier neurons and dorsal tier cells from the lateral and the medial-dorsal mesencephalon showed only a slight increase in TH mRNA, reaching a plateau at P10. 3. The time course of the observed increase in TH gene expression in the first group, generally parallels the innervation of their target cells in the striatal matrix, suggesting that TH gene expression in these cells may be influenced by their postsynaptic cells or by the innervation process.     

A Neural Circuit Mechanism for Encoding Aversive Stimuli in the Mesolimbic Dopamine System

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Acute administration of cocaine regulates the phosphorylation of serine-19, -31 and -40 in tyrosine hydroxylase

Acute cocaine can inhibit catecholamine biosynthesis by regulating the enzymatic activity of tyrosine hydroxylase via alterations in the phosphorylation state of the enzyme. The mechanisms underlying acute cocaine-dependent regulation of tyrosine hydroxylase phosphorylation have not been determined. In this study, 0, 15 or 30 mg/kg cocaine was administered intraperitoneally to rats and the phosphorylation state of tyrosine hydroxylase in the brain was examined using antibodies specific for the phosphorylated forms of serine-19, -31 and -40 in tyrosine hydroxylase. In the caudate and nucleus accumbens, cocaine dose-dependently decreased the levels of phosphorylated serine-19, -31 and -40. In the ventral tegmental area, the levels of phosphorylated serine-19, but not serine-31 and -40, were decreased by 15 and 30 mg/kg cocaine. In the amygdala, the levels of phosphorylated serine-19, but not serine-31 or -40, were decreased. The functional effects of these alterations in phosphorylation state were assessed by measuring tyrosine hydroxylase activity in vivo (accumulation of DOPA after administration of the decarboxylase inhibitor NSD-1015). Acute administration of 30 mg/kg cocaine significantly decreased l-DOPA production in caudate and accumbens but not in amygdala. These data suggest that the phosphorylation of serine-31 or -40, but not serine-19, is involved in the regulation of tyrosine hydroxylase activity by acute cocaine.     

Neurochemical characterization of the release and uptake of dopamine in ventral tegmental area and serotonin in substantia nigra of the mouse

In the present report, fast-scan cyclic voltammetry was used to identify the monoamines that were released by electrical stimulation in mouse brain slices containing ventral tegmental area (VTA), substantia nigra (SN) -pars compacta (SNc) and -pars reticulata (SNr). We showed that voltammograms obtained in mouse VTA were consistent with detection of a catecholamine, while those in both subregions of the SN were consistent with detection of an indolamine, based on the reduction peak potentials. We used pharmacological blockade and genetic deletion of monoamine transporters to further confirm the identity of released monoamines in mouse midbrain and to assess the control of monoamines by their transporters in each brain region. Inhibition of dopamine and norepinephrine transporters by nomifensine (1 and 10 microm) decreased uptake rates in the VTA, but did not change uptake rates in either subregion of the SN. Serotonin transporter inhibition by fluoxetine (10 microm) decreased uptake rates in the SNc and SNr, but was without effect in the VTA. Selective inhibition of the norepinephrine transporter by desipramine (10 microm) had no effect in any brain region. Using dopamine transporter- and serotonin transporter-knockout mice, we found decreased uptake rates in VTA and SN subregions, respectively. Peak signals recorded in each midbrain region were pulse number dependent and exhibited limited frequency dependence. Thus, dopamine is predominately detected by voltammetry in mouse VTA, while serotonin is predominately detected in mouse SNc and SNr. Furthermore, active uptake occurs in these areas and can be altered only by specific uptake inhibitors, suggesting a lack of heterologous uptake. In addition, somatodendritic dopamine release in VTA was not mediated by monoamine transporters. This work offers an initial characterization of voltammetric signals in the midbrain of the mouse and provides insight into the regulation of monoamine neurotransmission in these areas.     

Repeated exposure to cocaine alters the modulation of mesocorticolimbic glutamate transmission by medial prefrontal cortex Group II metabotropic glutamate receptors

Repeated cocaine exposure enhances glutamatergic output from the medial prefrontal cortex to subcortical brain regions. Loss of inhibitory control of cortical pyramidal neurons may partly account for this augmented cortical glutamate output. Recent research indicated that repeated cocaine exposure reduced the ability of cortical Group II metabotropic glutamate receptors to modulate behavioral and neurochemical responses to cocaine. Thus, experiments described below examined whether repeated cocaine exposure alters metabotropic glutamate receptor regulation of mesocorticolimbic glutamatergic transmission using in vivo microdialysis. Infusion of the Group II metabotropic glutamate receptor antagonist LY341495 into the medial prefrontal cortex enhanced glutamate release in this region, the nucleus accumbens and the ventral tegmental area in sensitized animals, compared to controls, following short-term withdrawal but not after long-term withdrawal. Additional studies demonstrated that vesicular (K(+)-evoked) and non-vesicular (cystine-evoked) glutamate release in the medial prefrontal cortex was enhanced in sensitized animals, compared to controls, that resulted in part from a reduction in Group II metabotropic glutamate receptor modulation of these pools of glutamate. In summary, these findings indicate that the expression of sensitization to cocaine is correlated with an altered modulation of mesocorticolimbic glutamatergic transmission via reduction of Group II metabotropic glutamate receptor function.     

Dopaminergic toxicity associated with oral exposure to the herbicide atrazine in juvenile male C57BL/6 mice

The herbicide atrazine (ATR) is a very commonly used pesticide in the United States. and a major ground water contaminant. It has also been recently implicated as a potential basal ganglia toxicant. In the present study, our objective was to determine the effects of ATR exposure on striatal neurochemistry, on the number of dopaminergic neurons in the substantia nigra pars compacta (SNpc), and, as a reference, in the ventral tegmental area (VTA) of male juvenile C57BL/6 mice. Oral exposure to ATR for 14 days dose-dependently decreased the levels of dopamine (DA) and its metabolites in the striatum for up to a week post-treatment. ATR exposure also time- and dose-dependently decreased the number of tyrosine hydroxylase-positive (TH+) dopaminergic neurons in both SNpc and VTA (with effects being slightly more prominent in SNpc), such that the decreases were most evident at 7 weeks post-cessation of exposure to ATR. Together, these data indicate that, in the juvenile male C57BL/6 mouse, the neurotoxic effects of ATR appear to cause transient neurochemical alterations, whereas the loss of TH+ neurons appears to be persistent, possibly confined to basal ganglia dopaminergic neurons, but not exclusive to the SNpc.     

The effects of antipsychotics on beta-catenin, glycogen synthase kinase-3 and dishevelled in the ventral midbrain of rats

Protein kinase B and glycogen synthase kinase-3 have been identified as susceptibility genes for schizophrenia and altered protein and mRNA levels have been detected in the brains of schizophrenics post-mortem. Recently, we reported that haloperidol, clozapine and risperidone alter glycogen synthase kinase-3 and beta-catenin protein expression and glycogen synthase kinase-3 phosphorylation levels in the rat prefrontal cortex and striatum. In the current study, beta-catenin, adenomatous polyposis coli, Wnt1, dishevelled and glycogen synthase kinase-3 were examined in the ventral midbrain and hippocampus using western blotting. In addition, beta-catenin and GSK-3 were examined in the substantia nigra and ventral tegmental area using confocal and fluorescence microscopy. The results indicate that repeated antipsychotic administration results in significant elevations in glycogen synthase kinase-3, beta-catenin and dishevelled-3 protein levels in the ventral midbrain and hippocampus. Raclopride causes similar changes in beta-catenin and GSK-3 in the ventral midbrain, suggesting that D2 dopamine receptor antagonism mediated the changes observed following antipsychotic administration. In contrast, amphetamine, a drug capable of inducing psychotic episodes, had the opposite effect on beta-catenin and GSK-3 in the ventral midbrain. Collectively, the results suggest that antipsychotics may exert their beneficial effects through modifications to proteins that are associated with the canonical Wnt pathway.     

Circulating Triglycerides Gate Dopamine-Associated Behaviors through DRD2-Expressing Neurons

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D1-D2 dopamine receptor interaction within the nucleus accumbens mediates long-loop negative feedback to the ventral tegmental area (VTA)

The objective of the present study was to examine the effects of perfusion of dopamine (DA) D1- and D2-like receptor agonists in the nucleus accumbens (ACB) on the long-loop negative feedback regulation of mesolimbic somatodendritic DA release in the ventral tegmental area (VTA) of Wistar rats employing ipsilateral dual probe in vivo microdialysis. Perfusion of the ACB for 60 min with the D1-like receptor agonist SKF 38393 (SKF, 1-100 microM) dose-dependently reduced the extracellular levels of DA in the ACB, whereas the extracellular levels of DA in the VTA were not changed. Similarly, application of the D2-like receptor agonist quinpirole (Quin, 1-100 microM) through the microdialysis probe in the ACB reduced the extracellular levels of DA in the ACB in a concentration-dependent manner, whereas extracellular levels of DA in the VTA were not altered. Co-application of SKF (100 microM) and Quin (100 microM) produced concomitant reductions in the extracellular levels of DA in the ACB and VTA. The reduction in extracellular levels of DA in the ACB and VTA produced by co-infusion of SKF and Quin was reversed in the presence of either 100 microM SCH 23390 (D1-like antagonist) or 100 microM sulpiride (D2-like antagonist). Overall, the results suggest that (a) activation of dopamine D1- or D2-like receptors can independently regulate local terminal DA release in the ACB, whereas stimulation of both subtypes is required for activation of the negative feedback pathway to the VTA.     

Eating-Induced Dopamine Release from Mesolimbic Neurons Is Mediated by NMDA Receptors in the Ventral Tegmental Area: A Dual-Probe Microdialysis Study

Abstract: This study was aimed at identifying the neuronal pathways that mediate the eating-induced increase in the release of dopamine in the nucleus accumbens of the rat brain. For that purpose, a microdialysis probe was implanted in the ventral tegmental area and a second probe was placed in the ipsilateral nucleus accumbens. Receptor-specific compounds acting on GABA_A (40 µ M muscimol; 50 µ M bicuculline), GABA_B (50 µ M baclofen), acetylcholine (50 µ M carbachol), NMDA [30 µ M (±)-3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP)], and non-NMDA [300 µ M 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX)] receptors were infused into the ventral tegmental area by retrograde dialysis, whereas extracellular dopamine was recorded in the ipsilateral nucleus accumbens. Intrategmental infusion of muscimol or baclofen decreased extracellular dopamine in the ipsilateral nucleus accumbens; CPP and CNQX were without effect, and bicuculline and carbachol increased dopamine release. During infusion of the various compounds, food-deprived rats were allowed to eat for 10 min. The infusions of muscimol, bicuculline, baclofen, carbachol, and CNQX did not prevent the eating-induced increase in extracellular dopamine in the nucleus accumbens. However, during intrategmental infusion of CPP, the eating-induced increase in extracellular dopamine in the nucleus accumbens was suppressed. These results indicate that a glutamatergic projection to the ventral tegmental area mediates, via an NMDA receptor, the eating-induced increase in dopamine release from mesolimbic dopamine neurons.     

GABAA Receptor Blockade in the Anterior Ventral Tegmental Area Increases Extracellular Levels of Dopamine in the Nucleus Accumbens of Rats

Abstract: Previously, it was shown that microinfusion of the GABA_A antagonist picrotoxin into the anterior ventral tegmental area (VTA) is reinforcing. It was hypothesized that this reinforcing effect of picrotoxin in the anterior VTA is mediated, at least in part, by the activation of the mesoaccumbens dopamine (DA) system. The objective of the present study was to determine if blockade of GABA_A receptors in the anterior VTA can increase extracellular levels of DA in the nucleus accumbens (ACB), using an in vivo microdialysis technique in freely moving rats. Concentrations of picrotoxin (40, 80, and 160 µ M ) that had previously been shown to produce a reinforcing effect increased the extracellular levels of DA and its major metabolites in the ACB. The increased extracellular DA levels induced by intra-VTA injection of picrotoxin was markedly attenuated by coadministration with the GABA_A agonist muscimol, whereas intra-VTA injection of muscimol alone did not have an apparent effect on extracellular DA levels in the ACB. Microinjection of another GABA_A antagonist, bicuculline, into the anterior VTA also increased the extracellular release of DA in the ACB. These results suggest that DA neurons projecting from the anterior VTA to the ACB are tonically inhibited by GABA through its actions at the GABA_A receptors.     

Involvement of GABA and cholinergic receptors in the nucleus accumbens on feedback control of somatodendritic dopamine release in the ventral tegmental area

The objectives of the present study were to examine the involvement of GABA and cholinergic receptors within the nucleus accumbens (ACB) on feedback regulation of somatodendritic dopamine (DA) release in the ventral tegmental area (VTA). Adult male Wistar rats were implanted with ipsilateral dual guide cannulae for in vivo microdialysis studies. Activation of the feedback system was accomplished by perfusion of the ACB with the DA uptake inhibitor GBR 12909 (GBR; 100 microm). To assess the involvement of GABA and cholinergic receptors in regulating this feedback system, antagonists (100 microm) for GABAA (bicuculline, BIC), GABAB (phaclofen, PHAC), muscarinic (scopolamine, SCOP), and nicotinic (mecamylamine, MEC) receptors were perfused through the probe in the ACB while measuring extracellular DA levels in the ACB and VTA. Local perfusion of the ACB with GBR significantly increased (500% of baseline) the extracellular levels of DA in the ACB and produced a concomitant decrease (50% of baseline) in the extracellular DA levels in the VTA. Perfusion of the ACB with BIC or PHAC alone produced a 200-400% increase in the extracellular levels of DA in the ACB but neither antagonist altered the levels of DA in the VTA. Co-perfusion of either GABA receptor antagonist with GBR further increased the extracellular levels of DA in the ACB to 700-800% of baseline. However, coperfusion with BIC completely prevented the reduction in the extracellular levels of DA in the VTA produced by GBR alone, whereas PHAC partially prevented the reduction. Local perfusion of the ACB with either MEC or SCOP alone had little effect on the extracellular levels of DA in the ACB or VTA. Co-perfusion of either cholinergic receptor antagonist with GBR markedly reduced the extracellular levels of DA in the ACB and prevented the effects of GBR on reducing DA levels in the VTA. Overall, the results of this study suggest that terminal DA release in the ACB is under tonic GABA inhibition mediated by GABAA (and possibly GABAB) receptors, and tonic cholinergic excitation mediated by both muscarinic and nicotinic receptors. Activation of GABAA (and possibly GABAB) receptors within the ACB may be involved in the feedback inhibition of VTA DA neurons. Cholinergic interneurons may influence the negative feedback system by regulating terminal DA release within the ACB.     

Specific involvement of neurotensin type 1 receptor in the neurotensin-mediated in vivo dopamine efflux using knock-out mice

Abstract Neurotensin is a tridecapeptide neurotransmitter known to be involved in psychiatric disorders, various physiological processes and several different neurobiological mechanisms, including modulation of accumbal dopamine release. Two neurotensin extracellular binding sites, namely NT1- and NT2-receptor (NT1R and NT2R), have been cloned from the rat brain. These receptors are distinguishable by their different in vitro pharmacological properties but the available pharmacological tools have weak in vivo potency and specificity. The use of genetically engineered knock-out mice has provided a powerful alternative to the classical pharmacological approach to investigate their respective roles. In this study, using in vivo differential pulse amperometry, we show that, in wild-type mice, neurotensin application into the ventral tegmental area dose-dependently evokes dopamine efflux in the nucleus accumbens. This neurotensin-mediated efflux is dramatically decreased in mice lacking NT1R while it is unaffected in NT2R-deleted mice. This finding indicates that a large part of the dopamine efflux evoked by neurotensin in the nucleus accumbens of wild-type mice is mediated via NT1R present in the ventral tegmental area.