High doses of amphetamine augment, rather than disrupt, exocytotic dopamine release in the dorsal and ventral striatum of the anesthetized rat

High doses of amphetamine (AMPH) are thought to disrupt normal patterns of action potential-dependent dopaminergic neurotransmission by depleting vesicular stores of dopamine (DA) and inducing robust non-exocytotic DA release or efflux via dopamine transporter (DAT) reversal. However, these cardinal AMPH actions have been difficult to establish definitively in vivo. Here, we use fast-scan cyclic voltammetry (FSCV) in the urethane-anesthetized rat to evaluate the effects of 10 and 20 mg/kg AMPH on vesicular DA release and DAT function in dorsal and ventral striata. An equivalent high dose of cocaine (40 mg/kg) was also examined for comparison to psychostimulants acting preferentially by DAT inhibition. Parameters describing exocytotic DA release and neuronal DA uptake were determined from dynamic DA signals evoked by mild electrical stimulation previously established to be reinforcing. High-sensitivity FSCV with nanomolar detection was used to monitor changes in the background voltammetric signal as an index of DA efflux. Both doses of AMPH and cocaine markedly elevated evoked DA levels over the entire 2-h time course in the dorsal and ventral striatum. These increases were mediated by augmented vesicular DA release and diminished DA uptake typically acting concurrently. AMPH, but not cocaine, induced a slow, DA-like rise in some baseline recordings. However, this effect was highly variable in amplitude and duration, modest, and generally not present at all. These data thus describe a mechanistically similar activation of action potential-dependent dopaminergic neurotransmission by AMPH and cocaine in vivo. Moreover, DA efflux appears to be a unique, but secondary, AMPH action.     

7-Nitroindazole enhances amphetamine-evoked dopamine release in rat striatum. an in vivo microdialysis and voltammetric study.

The intracellular second messenger nitric oxide (NO) is implicated in a variety of physiological functions, including release and uptake of dopamine (DA). In the described study, in vivo microdialysis and differential pulse voltammetric techniques were used to determine the involvement of NO in release of DA and its metabolites (dihydroxyphenylalanine, DOPAC; homovanillic acid, HVA) in neostriatum of freely moving rats. While the NO donor molsidomine (30.0 mg/kg; MOLS) and neuronal NO synthase- (nNOS-) inhbitor 7-nitroindazole (10.0 mg/kg; 7-NI) had no effect on the basal in vivo microdialysate level of DA, 7-NI specifically enhanced D,L-amphetamine-(1.0 mg/kg i.p.; AMPH) evoked release of DA. Basal or AMPH effects on DOPAC and HVA levels were not influenced by MOLS or 7-NI. Findings indicate that nitrergic systems have an important role in mediating effects of AMPH on dopaminergic systems.     

Methamphetamine neurotoxicity decreases phasic, but not tonic, dopaminergic signaling in the rat striatum

Neurotoxic doses of methamphetamine (METH) are known to cause depletions in striatal dopamine (DA) tissue content. However, the effects of METH-induced insults on dopaminergic neurotransmission are not fully understood. Here, we employed fast-scan cyclic voltammetry at a carbon-fiber microelectrode in the anesthetized rat striatum to assess the effects of a neurotoxic regimen of METH on phasic and tonic modes of dopaminergic signaling and underlying mechanisms of DA release and uptake. Extracellular DA was electrically evoked by stimulation of the medial forebrain bundle mimicking tonic and phasic firing patterns for dopaminergic cells and was monitored simultaneously in both the dorsomedial and dorsolateral striatum. Kinetic analysis of evoked recordings determined parameters describing DA release and uptake. Striatal DA tissue content was quantified by high performance liquid chromatography with electrochemical detection. METH-pretreatment (four doses of 7.5 or 10.0 mg/kg s.c.) induced DA depletions of ～ 40% on average, which are reported in both striatal subregions. METH pre-treatment significantly decreased the amplitude of signals evoked by phasic, but not tonic, stimulation. Parameters for DA release and uptake were also similarly reduced by ～ 40%, consistent with effects on evoked phasic-like responses and DA tissue content. Taken together, these results suggest that METH-pretreatment selectively diminishes phasic, but not tonic, dopaminergic signaling in the dorsal striatum.     

Amphetamine-Induced Dopamine Release in the Rat Striatum: An In Vivo Microdialysis Study

The effects of a number of biochemical and pharmacological manipulations on amphetamine (AMPH)-induced alterations in dopamine (DA) release and metabolism were examined in the rat striatum using the in vivo brain microdialysis method. Basal striatal dialysate concentrations were: DA, 7 nM; dihydroxyphenylacetic acid (DOPAC), 850 nM; homovanillic acid (HVA), 500 nM; 5-hydroxyindoleacetic acid (5-HIAA), 300 nM; and 3-methoxytyramine (3-MT), 3 nM. Intraperitoneal injection of AMPH (4 mg/kg) induced a substantial increase in DA efflux, which attained its maximum response 20-40 min after drug injection. On the other hand, DOPAC and HVA efflux declined following AMPH. The DA response, but not those of DOPAC and HVA, was dose dependent within the range of AMPH tested (2-16 mg/kg). High doses of AMPH (greater than 8 mg/kg) also decreased 5-HIAA and increased 3-MT efflux. Depletion of vesicular stores of DA using reserpine did not affect significantly AMPH-induced dopamine efflux. In contrast, prior inhibition of catecholamine synthesis, using alpha-methyl-p-tyrosine, proved to be an effective inhibitor of AMPH-evoked DA release (less than 35% of control). Moreover, the DA releasing action of AMPH was facilitated in pargyline-pretreated animals (220% of control). These data suggest that AMPH releases preferentially a newly synthesised pool of DA. Nomifensine, a DA uptake inhibitor, was an effective inhibitor of AMPH-induced DA efflux (18% of control). On the other hand, this action of AMPH was facilitated by veratrine and ouabain (200-210% of control). These results suggest that the membrane DA carrier may be involved in the actions of AMPH on DA efflux.     

Amphetamine augments vesicular dopamine release in the dorsal and ventral striatum through different mechanisms

Amphetamine has well‐established actions on pre‐synaptic dopamine signaling, such as inhibiting uptake and degradation, activating synthesis, depleting vesicular stores, and promoting dopamine‐transporter reversal and non‐exocytotic release. Recent in vivo studies have identified an additional mechanism: augmenting vesicular release. In this study, we investigated how amphetamine elicits this effect. Our hypothesis was that amphetamine enhances vesicular dopamine release in dorsal and ventral striata by differentially targeting dopamine synthesis and degradation. In urethane‐anesthetized rats, we employed voltammetry to monitor dopamine, electrical stimulation to deplete stores or assess vesicular release and uptake, and pharmacology to isolate degradation and synthesis. While amphetamine increased electrically evoked dopamine levels, inhibited uptake, and up‐regulated vesicular release in both striatal sub‐regions in controls, this psychostimulant elicited region‐specific effects on evoked levels and vesicular release but not uptake in drug treatments. Evoked levels better correlated with vesicular release compared with uptake, supporting enhanced vesicular release as an important amphetamine mechanism. Taken together, these results suggested that amphetamine enhances vesicular release in the dorsal striatum by activating dopamine synthesis and inhibiting dopamine degradation, but targeting an alternative mechanism in the ventral striatum. Region‐distinct activation of vesicular dopamine release highlights complex cellular actions of amphetamine and may have implications for its behavioral effects.     

Hypoinsulinemia regulates amphetamine-induced reverse transport of dopamine

The behavioral effects of psychomotor stimulants such as amphetamine (AMPH) arise from their ability to elicit increases in extracellular dopamine (DA). These AMPH-induced increases are achieved by DA transporter (DAT)-mediated transmitter efflux. Recently, we have shown that AMPH self-administration is reduced in rats that have been depleted of insulin with the diabetogenic agent streptozotocin (STZ). In vitro studies suggest that hypoinsulinemia may regulate the actions of AMPH by inhibiting the insulin downstream effectors phosphotidylinositol 3-kinase (PI3K) and protein kinase B (PKB, or Akt), which we have previously shown are able to fine-tune DAT cell-surface expression. Here, we demonstrate that striatal Akt function, as well as DAT cell-surface expression, are significantly reduced by STZ. In addition, our data show that the release of DA, determined by high-speed chronoamperometry (HSCA) in the striatum, in response to AMPH, is severely impaired in these insulin-deficient rats. Importantly, selective inhibition of PI3K with LY294002 within the striatum results in a profound reduction in the subsequent potential for AMPH to evoke DA efflux. Consistent with our biochemical and in vivo electrochemical data, findings from functional magnetic resonance imaging experiments reveal that the ability of AMPH to elicit positive blood oxygen level–dependent signal changes in the striatum is significantly blunted in STZ-treated rats. Finally, local infusion of insulin into the striatum of STZ-treated animals significantly recovers the ability of AMPH to stimulate DA release as measured by high-speed chronoamperometry. The present studies establish that PI3K signaling regulates the neurochemical actions of AMPH-like psychomotor stimulants. These data suggest that insulin signaling pathways may represent a novel mechanism for regulating DA transmission, one which may be targeted for the treatment of AMPH abuse and potentially other dopaminergic disorders.     

Regional Differences in Striatal Dopamine Uptake and Release Associated with Recovery from MPTP-Induced Parkinsonism

Abstract : This study directly assessed striatal dopamine (DA) uptake rates and peak release in response to KCl in normal, symptomatic, and recovered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated cats using in vivo electrochemistry. DA uptake rates measured after direct application of known concentrations of DA to the striatum were slowed significantly in both dorsal and ventral striatum in symptomatic cats compared with rates recorded in normal animals. DA uptake rates remained significantly slowed in recovered cats and were not significantly different from the rates recorded in symptomatic animals. In symptomatic cats, both DA uptake rates and the signal recorded in response to KCl stimulation were significantly decreased from normal in all dorsal and ventral striatal regions sampled. Reduction/oxidation (redox) ratios recorded in response to KCl stimulation suggested DA to be the predominant electroactive species. In spontaneously recovered MPTP-treated cats, recordings in the ventral striatum subsequent to KCl stimulation again suggested DA to be the predominant electroactive species released, and peak levels were significantly higher than those recorded in symptomatic animals. In the dorsal striatum of recovered cats, redox ratios recorded subsequent to KCl stimulation suggested serotonin rather than DA to be the predominant electroactive species released. Peak levels of release in the dorsal striatum were not significantly greater than those recorded in symptomatic animals. These results suggest that in spontaneously recovered MPTP-treated cats, there is partial recovery of ventral striatal DAergic terminals, persistent loss of dorsal striatal DAergic terminals, and a down-regulation of DA transporter number/function throughout the striatum. These processes may contribute to volume transmission of DA in the striatum and promote functional recovery.     

The long-term effects of repeated amphetamine treatment in vivo on amphetamine, KCl and electrical stimulation evoked striatal dopamine release in vitro

Exposure to amphetamine (AMPH) in vivo produces an enduring enhancement ('sensitization') in AMPH-stimulated striatal DA release in vitro. Experiments were conducted to determine whether striatal DA release evoked by neuronal depolarization is altered by AMPH pretreatment in a similar manner. It was found that AMPH pretreatment produced a long-lasting (at least one week) enhancement in striatal DA release evoked by AMPH, KCl or electrical field stimulation. In contrast, the basal rate of DA efflux was not altered by pretreatment condition. A mechanism by which a single change in the intracellular distribution of DA could enhance both AMPH- and depolarization-induced DA release is proposed.     

Selective Deletion of PTEN in Dopamine Neurons Leads to Trophic Effects and Adaptation of Striatal Medium Spiny Projecting Neurons

The widespread distribution of the tumor suppressor PTEN in the nervous system suggests a role in a broad range of brain functions. PTEN negatively regulates the signaling pathways initiated by protein kinase B (Akt) thereby regulating signals for growth, proliferation and cell survival. Pten deletion in the mouse brain has revealed its role in controlling cell size and number. In this study, we used Cre-loxP technology to specifically inactivate Pten in dopamine (DA) neurons (Pten KO mice). The resulting mutant mice showed neuronal hypertrophy, and an increased number of dopaminergic neurons and fibers in the ventral mesencephalon. Interestingly, quantitative microdialysis studies in Pten KO mice revealed no alterations in basal DA extracellular levels or evoked DA release in the dorsal striatum, despite a significant increase in total DA tissue levels. Striatal dopamine receptor D1 (DRD1) and prodynorphin (PDyn) mRNA levels were significantly elevated in KO animals, suggesting an enhancement in neuronal activity associated with the striatonigral projection pathway, while dopamine receptor D2 (DRD2) and preproenkephalin (PPE) mRNA levels remained unchanged. In addition, PTEN inactivation protected DA neurons and significantly enhanced DA-dependent behavioral functions in KO mice after a progressive 6OHDA lesion. These results provide further evidence about the role of PTEN in the brain and suggest that manipulation of the PTEN/Akt signaling pathway during development may alter the basal state of dopaminergic neurotransmission and could provide a therapeutic strategy for the treatment of Parkinson''s disease, and other neurodegenerative disorders.     

Rapid substrate-induced down-regulation in function and surface localization of dopamine transporters: rat dorsal striatum versus nucleus accumbens

The dopamine transporter (DAT) substrates dopamine, d-amphetamine (AMPH), and methamphetamine are known to rapidly and transiently reduce DAT activity and/or surface expression in dorsal striatum and heterologous expression systems. We sought to determine if similar substrate-induced regulation of DATs occurs in rat nucleus accumbens. In dorsal striatum synaptosomes, brief (15-min) in vitro substrate pre-exposure markedly decreased maximal [³H]dopamine uptake velocity whereas identical substrate pre-exposure in nucleus accumbens synaptosomes produced a smaller, non-significant reduction. However, 45 min after systemic AMPH administration, maximal ex vivo [³H]dopamine uptake velocity was significantly reduced in both brain regions. Protein kinase C inhibition blocked AMPH's down-regulation of DAT activity. DAT synaptosomal surface expression was not modified following either the brief in vitro or in vivo AMPH pre-exposure but was reduced after a longer (1-h) in vitro pre-exposure in both brain regions. Together, our findings suggest that relatively brief substrate exposure results in greater down-regulation of DAT activity in dorsal striatum than in nucleus accumbens. Moreover, exposure to AMPH appears to regulate striatal DATs in a biphasic manner, with an initial protein kinase C-dependent decrease in DAT-mediated uptake velocity and then, with longer exposure, a reduction in DAT surface expression.     

Amphetamine-induced in vivo release of dopamine in the striatum is influenced by local pH

The effect of local pH on the in vivo efflux of endogenous dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) following administration of d-amphetamine (AMPH) was examined in the striatum of the anesthetized rat using two bilaterally placed push-pull cannulae. At both pH 7.3 and 6.4, the baseline efflux values for DA and DOPAC were approximately 0.2 and 25 pmoles/15 min, respectively. Subcutaneous injection of 2 mg/kg AMPH induced a 3-fold increase of DA release at pH 7.3 and a 21-fold increase of DA release at pH 6.4. In both cases, the maximum was reached at about 30 min after the drug administration. Following the administration of AMPH, the efflux of DOPAC was reduced to the same degree (20% of control values) under both pH conditions. In vitro data showed that the lower pH did not alter the recovery of DA or DOPAC. In addition, release of DA produced by local perfusion with 5 uM AMPH was also greater at the lower pH (50-fold increase over baseline) than at the physiological pH (10-fold increase over baseline). The stimulated DA release produced by local perfusion with 35 mM K+, however, was the same at both pH values. Preliminary experiments also indicated that there was a pH effect for AMPH-induced serotonin (5-HT) release but that the difference in the amount of 5-HT in the two media was not nearly as large as that obtained for DA. The markedly elevated level of extracellular DA at the lower pH might be due to a higher affinity of the DA uptake system for AMPH, thereby producing greater inhibition of DA uptake as well as enhanced DA release. The data also suggest an enhanced affinity of AMPH for 5-HT uptake sites at the lower pH.     

Tonic autoinhibition contributes to the heterogeneity of evoked dopamine release in the rat striatum

Electrically evoked dopamine release as measured by voltammetry in the rat striatum is heterogeneous in both amplitude and temporal profile. Previous studies have attributed this heterogeneity to variations in the density of dopamine (DA) terminals at the recording site. We reach the alternate conclusion that the heterogeneity of evoked DA release derives from variations in the extent to which DA terminals are autoinhibited. We demonstrate that low-amplitude, slow evoked DA responses occur even though recording electrodes are close to DA terminals. Moreover, the D₂ agonist and antagonist, quinpirole and raclopride, respectively, affect the slow responses in a manner consistent with the known functions of pre-synaptic D₂ autoreceptors. Recording sites that exhibit autoinhibited responses are prevalent in the dorsal striatum. Autoinhibition preceded electrical stimulation, which is consistent with our prior reports that the striatum contains a tonic pool of extracellular DA at basal concentrations that exceed the affinity of D₂ receptors. We conclude that the striatum contains DA terminals operating on multiple time courses, determined at least in part by the local variation in autoinhibition. Thus, we provide direct, real-time observations of the functional consequence of tonic and phasic DAergic signaling in vivo .     

Reserpine pretreatment prevents increases in extracellular striatal dopamine following L-DOPA administration in rats with nigrostriatal denervation

The influence of L-DOPA and reserpine on extracellular dopamine (DA) levels in the striatum of intact and dopaminergic denervated rats was studied using the brain microdialysis technique. In intact rats, reserpine (5 mg/kg s.c.) reduced extracellular DA levels to 4% of basal values. L-DOPA (50 mg/kg i.p.) had no effect on extracellular DA levels in reserpine-pretreated rats. In rats with 6-hydroxydopamine-induced lesion of the nigrostriatal dopaminergic system, basal levels of extracellular DA were low but markedly increased by L-DOPA (50 mg/kg i.p.). In 6-hydroxydopamine-lesioned rats, pretreatment with reserpine (5 mg/kg s.c.) diminished L-DOPA (50 mg/kg i.p.)-induced increases in extracellular DA levels to 16% of those obtained in denervated animals not pretreated with reserpine (p<0.01). These results suggest that in the intact striatum, extracellular DA stems mainly from vesicular storage sites and that in the striatum with dopaminergic denervation, a large part of the L-DOPA-derived extracellular DA is also derived from a vesicular pool that is released by an exocytosis mechanism.     

Evoked dopamine overflow is augmented in the striatum of calcitriol treated rats

Calcitriol, the active metabolite of vitamin D, has been shown to have significant effects on the brain. These actions include reducing the severity of some central nervous system lesions, possibly by upregulating trophic factors such as glial cell line-derived neurotrophic factor (GDNF). GDNF has substantial effects on the nigrostriatal dopamine (DA) system of young adult, aged and lesioned animals. Thus, the administration of calcitriol may lead to significant effects on nigrostriatal DA neuron functioning. The present experiments were designed to examine the ability of calcitriol to alter striatal DA release, and striatal and nigral tissue levels of DA. Male Fischer-344 rats were administered vehicle or calcitriol (0.3, 1.0, or 3.0 μg/kg, s.c.) once daily for eight consecutive days. Three weeks later in vivo microdialysis experiments were conducted to measure basal and stimulus evoked overflow of DA from the striatum. Basal levels of extracellular DA were not significantly affected by the calcitriol treatments. However, the 1.0 and 3.0 μg/kg doses of calcitriol led to increases in both potassium and amphetamine evoked overflow of striatal DA. Although post-mortem tissue levels of striatal DA were not altered by the calcitriol injections, nigral tissue levels of DA and its main metabolites were increased by both the 1.0 and 3.0 μg/kg doses of calcitriol. In a separate group of animals GDNF levels were augmented in the striatum and substantia nigra after eight consecutive daily injections of calcitriol. These results suggest that systemically administered calcitriol can upregulate dopaminergic release processes in the striatum and DA levels in the substantia nigra. Increases in the levels of endogenous GDNF following calcitriol treatment may in part be responsible for these changes. The ability of calcitriol to lead to augmented DA release in the striatum suggests that calcitriol may be beneficial in disease processes involving dopaminergic dysfunction.     

Protein Phosphatases but not Reactive Oxygen Species Play Functional Role in Acute Amphetamine-Mediated Dopamine Release

Drug abuse-induced neurodegeneration can be triggered by elevated production of reactive oxygen species (ROS). Involvement of oxidative stress in acute amphetamine (AMPH)-mediated dopamine (DA) release, however, has not been completely understood yet. In order to elucidate the dopaminergic response of PC12 cells to a single dose of 10 μM AMPH, ROS production was measured as related to the extracellular DA level. Due to the spontaneous oxidation of peroxide-sensitive fluorophore 2′,7′-dichlorofluorescin diacetate (DCFH-DA) to 2′,7′-dichlorofluorescein (DCF), the increase in fluorescence could not be unambiguously attributed to AMPH-triggered ROS production. Based on Amplex Red fluorescence, no ROS production was detected after acute AMPH application. Our data strongly suggest that ROS development was not the main triggering factor for immediate DA release after acute AMPH treatment. On the other hand, AMPH-induced elevation of DA levels in rat brain striatal slices was quenched by the water soluble antioxidant, N-acetylcysteine (NAC) at 10 mM. In this study, we also investigated the contribution of protein phosphatases to the AMPH-induced rat brain striatal dopaminergic response. The experimental protocol, double AMPH challenge was applied for screening the effect of NAC and cantharidin on AMPH-mediated DA release. Here we show that AMPH-mediated DA release increased nearly twofold in striatal rat brain slices pretreated for 30 min with 1000 μM cantharidin, a selective PP1 and PP2A inhibitor. These findings prove the lack of ROS inhibitory action on protein phosphatase activity in acute AMPH-mediated DA efflux.     

New findings on nuclear gangliosides: overview on metabolism and function

Dopamine (DA) is an important transmitter in both motor and limbic pathways. We sought to investigate the role of D(1)-receptor activation in axonal DA release regulation in dorsal striatum using a D(1)-receptor antagonist, SKF-83566. Evoked DA release was monitored in rat striatal slices using fast-scan cyclic voltammetry. SKF-83566 caused a concentration-dependent increase in peak single-pulse evoked extracellular DA concentration, with a maximum increase of ～ 65% in 5 μM SKF-83566. This was accompanied by a concentration-dependent increase in extracellular DA concentration clearance time. Both effects were occluded by nomifensine (1 μM), a dopamine transporter (DAT) inhibitor, suggesting that SKF-83566 acted via the DAT. We tested this by examining [(3)H]DA uptake into LLc-PK cells expressing rat DAT, and confirmed that SKF-83566 is a competitive DAT inhibitor with an IC(50) of 5.7 μM. Binding studies with [(3)H]CFT, a cocaine analog, showed even more potent action of SKF-83566 at the DAT cocaine binding site (IC(50) = 0.51 μM). Thus, data obtained using SKF-83566 as a D(1) DA-receptor antagonist may be confounded by concurrent DAT inhibition. More positively, however, SKF-83566 might be a candidate to attenuate cocaine effects in vivo because of the greater potency of this drug at the cocaine versus DA binding site of the DAT.     

Modulation of dopaminergic neurotransmission in rat striatum upon in vitro and in vivo diclofenac treatment

Diclofenac (DCF) is a widely used non-steroidal anti-inflammatory drug, which also act as a mitochondrial toxin. As it is known that selective mitochondrial complex I inhibition combined with mild oxidative stress causes striatal dopaminergic dysfunction, we tested whether DCF also compromise dopaminergic function in the striatum. [3H]Dopamine ([3H]DA) release was measured from rat striatal slices after in vitro (2 h, 10-25 micromol/L) or in vivo (3 mg/kg i.v. for 28 days) DCF treatment. In vitro treatment significantly decreased [3H]DA uptake and dopamine (DA) content of the slices. H2O2 (0.1 mmol/L)-evoked DA release was enhanced. Intracellular reactive oxygen species production was not significantly changed in the presence of DCF. After in vivo DCF treatment no apparent decrease in striatal DA content was observed and the uptake of [3H]DA into slices was increased. The intensity of tyrosine hydroxylase immunoreactivity in the striatum was highly variable, and both decrease and increase were observed in individual rats. The H2O2-evoked [3H]DA release was significantly decreased and the effluent contained a significant amount of [3H]octopamine, [3H]tyramine, and [3H]beta-phenylethylamine. The ATP content and adenylate energy charge were decreased. In conclusion, whereas in vitro DCF pre-treatment resembles the effect of the mitochondrial toxin rotenone, in vivo it rather counteracts than aggravates dopaminergic dysfunction.     

Macroautophagy can press a brake on presynaptic neurotransmission

The mechanistic target of rapamycin (MTOR) has been implicated in regulating synaptic plasticity and neurodegeneration, but MTOR’s role in modulating presynaptic function through autophagy is unexplored. We studied presynaptic function in ventral dopamine neurons, a system from which neurotransmitter release can be measured directly by cyclic voltammetry. We generated mutant mice that were specifically deficient for macroautophagy in dopaminergic neurons by deleting the Atg7 gene in cells that express the dopamine uptake transporter. Dopamine axonal profiles in the mutant dorsal striatum were ~one third larger in the mutant mice, released ~50% more stimulus-evoked dopamine release, and exhibited more rapid presynaptic recovery than controls. Rapamycin reduced dopamine neuron axon profile size by ~30% in control mice, but had no effect on macroautophagy deficient axons. Acute rapamycin decreased dopaminergic synaptic vesicle density by ~25% and inhibited evoked dopamine release by ~25% in control mice, but not in the Atg7 deficient mutants. Thus, both basal and induced macroautophagy can provide a brake on presynaptic activity in vivo, perhaps by regulating the turnover of synaptic vesicles, and further regulates terminal volume and the kinetics of transmitter release.     

Attenuation of estradiol on the reduction of striatal dopamine by amphetamine in ovariectomized rats

Amphetamine (AMPH) is a highly addictive drug of abuse which exhibits toxicity to dopaminergic neurons in long‐term abusers. Estrogen seems to show neuroprotection in dopamine (DA) deficit caused by AMPH. The present study was to investigate the effects of estradiol on the levels of striatal DA in ovariectomized (Ovx) rats treated with or without AMPH. Female rats were Ovx for 2 weeks before administration of AMPH (5 mg/kg/day, i.p.) with or without 17β‐estradiol benzoate (EB) (25 µg/kg/day, s.c.) for 7 days. The striatal tissues were collected, homogenized with DA mobile phase, and centrifuged. The concentrations of DA in the supernatants were detected by HPLC. The protein expressions of dopamine transporter (DAT), vesicular monoamine transporter 2 (VMAT‐2), and tyrosine hydroxylase (TH) were analyzed by Western blotting. The results indicated that AMPH could attenuate DA level significantly in striatum (P < 0.01). Comparing to control groups, administration of either EB or EB plus AMPH increased DA level (P < 0.01). The protein expression of striatal DAT was significant greater (P < 0.01) in rats treated with AMPH plus EB than AMPH treated animals. These results suggest that the DA levels in striatum can be enhanced by EB via an increase of DAT expression following administration of AMPH. J. Cell. Biochem. 108: 1318–1324, 2009. © 2009 Wiley‐Liss, Inc.     

Conserved dorsal–ventral gradient of dopamine release and uptake rate in mice,rats and rhesus macaques

Although the vast majority of research on the dopamine system has been performed in rodents, and it is assumed that this work will inform us about the human condition, there have been very few direct comparisons of presynaptic dopamine terminal function across multiple species. Because it is difficult to query rapid sub-second dopamine signaling in humans using voltammetric methods, we chose to compare dopamine signals across multiple striatal subregions in slices from C57BL/6J mice, Sprague–Dawley rats and rhesus macaques. We found a dorsal to ventral gradient of dopamine uptake rates with highest levels in the dorsal striatum and lowest levels in the nucleus accumbens shell, which is conserved across species. In addition to uptake rates, there was also a dorsal to ventral, high to low, gradient in the magnitude of stimulated DA release observed in monkeys, mice, and rats. These data demonstrate that there is considerable functional homology across striatal regions in non-human primates and rodents, lending support to the use of rodents as model systems to study dopamine-related circuitry and disorders that are clinically relevant to the human population.