Dopamine release is impaired in a mouse model of DYT1 dystonia

Early onset torsion dystonia, the most common form of hereditary primary dystonia, is caused by a mutation in the TOR1A gene, which codes for the protein torsinA. This form of dystonia is referred to as DYT1. We have used a transgenic mouse model of DYT1 dystonia [human mutant-type (hMT)1 mice] to examine the effect of the mutant human torsinA protein on striatal dopaminergic function. Analysis of striatal tissue dopamine (DA) and metabolites using HPLC revealed no difference between hMT1 mice and their non-transgenic littermates. Pre-synaptic DA transporters were studied using in vitro autoradiography with [(3)H]mazindol, a ligand for the membrane DA transporter, and [(3)H]dihydrotetrabenazine, a ligand for the vesicular monoamine transporter. No difference in the density of striatal DA transporter or vesicular monoamine transporter binding sites was observed. Post-synaptic receptors were studied using [(3)H]SCH-23390, a ligand for D(1) class receptors, [(3)H]YM-09151-2 and a ligand for D(2) class receptors. There were again no differences in the density of striatal binding sites for these ligands. Using in vivo microdialysis in awake animals, we studied basal as well as amphetamine-stimulated striatal extracellular DA levels. Basal extracellular DA levels were similar, but the response to amphetamine was markedly attenuated in the hMT1 mice compared with their non-transgenic littermates (253 +/- 71% vs. 561 +/- 132%, p < 0.05, two-way anova). These observations suggest that the mutation in the torsinA protein responsible for DYT1 dystonia may interfere with transport or release of DA, but does not alter pre-synaptic transporters or post-synaptic DA receptors. The defect in DA release as observed may contribute to the abnormalities in motor learning as previously documented in this transgenic mouse model, and may contribute to the clinical symptoms of the human disorder.     

Paradoxical modulation of short-term facilitation of dopamine release by dopamine autoreceptors

Electrophysiological studies have demonstrated that dopaminergic neurons burst fire during certain aspects of reward-related behavior; however, the correlation between dopamine release and cell firing is unclear. When complex stimulation patterns that mimic intracranial self-stimulation were employed, dopamine release was shown to exhibit facilitated as well as depressive components (Montague et al. 2004). Understanding the biological mechanisms underlying these variations in dopamine release is necessary to unravel the correlation between unit activity and neurotransmitter release. The dopamine autoreceptor provides negative feedback to dopamine release, inhibiting release on the time scale of a few seconds. Therefore, we investigated this D(2) receptor to see whether it is one of the biological mechanisms responsible for the history-dependent modulation of dopamine release. Striatal dopamine release in anesthetized rats was evoked with stimulus trains that were designed to promote the variability of dopamine release. Consistent with the well established D(2)-mediated autoinhibition, the short-term depressive component of dopamine release was blocked by raclopride, a D(2) antagonist, and enhanced by quinpirole, a D(2)-receptor agonist. Surprisingly, these same drugs exerted a similar effect on the short-term facilitated component: a decrease with raclopride and an increase with quinpirole. These data demonstrate that the commanding control exerted by dopamine autoreceptors over short-term neuroadaptation of dopamine release involves both inhibitory and paradoxically, facilitatory components.     

Characterization of the striatal dopaminergic neurotransmission in MEN2B mice with elevated cerebral tissue dopamine

The Ret receptor tyrosine kinase is the common signaling receptor for the glial cell line-derived neurotrophic factor (GDNF) family ligands. The Met918Thr mutation leads to constitutive activation of Ret and is responsible for dominantly inherited cancer syndrome MEN2B. Previously, we found that the mice carrying the mutation (MEN2B mice) have profoundly increased tissue dopamine (DA) concentrations in the striatum as well as increased striatal levels of tyrosine hydroxylase (TH) and dopamine transporter. The aim of this study was to characterize the striatal dopaminergic neurotransmission in MEN2B mice and to clarify the mechanisms by which they compensate their over-production of DA. We found that tyrosine hydroxylase activity and DA synthesis are increased in MEN2B mice. Augmented effects of α-methyl-para-tyrosine (αMT, an inhibitor of TH) and tetrabenazine (VMAT2 blocker) on DA levels suggest that also storage of DA is increased in MEN2B mice. There was no difference in the basal extracellular DA concentrations or potassium-evoked DA release between the genotypes. The effects of cocaine and haloperidol were also similar between the genotypes as assessed by in vivo microdialysis. However, with in vivo voltammetry we found increase in stimulated DA release in MEN2B mice and detailed analysis of DA overflow showed that uptake of DA was also enhanced in MEN2B mice. Thus, our data show that enhanced synthesis of DA leading to increased storage and releasable pools in pre-synaptic terminals in MEN2B mice apparently also leads to increased DA release, which in turn is compensated by higher dopamine transporter activity.     

Selective stimulation of serotonin2c receptors blocks the enhancement of striatal and accumbal dopamine release induced by nicotine administration

The effects of acute and repeated nicotine administration on the extracellular levels of dopamine (DA) in the corpus striatum and the nucleus accumbens were studied in conscious, freely moving rats by in vivo microdialysis. Acute intraperitoneal (i.p.) injection of nicotine (1 mg/kg) increased DA outflow both in the corpus striatum and the nucleus accumbens. Repeated daily injection of nicotine (1 mg/kg, i.p.) for 10 consecutive days caused a significant increase in basal DA outflow both in the corpus striatum and the nucleus accumbens. Acute challenge with nicotine (1 mg/kg, i.p.) in animals treated repeatedly with this drug enhanced DA extracellular levels in both brain areas. However, the effect of nicotine was potentiated in the nucleus accumbens, but not in the corpus striatum. To test the hypothesis that stimulation of 5-HT (5-hydroxytryptamine, serotonin)(2C) receptors could affect nicotine-induced DA release, the selective 5-HT(2C) receptor agonist RO 60-0175 was used. Pretreatment with RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently prevented the enhancement in DA release elicited by acute nicotine in the corpus striatum, but was devoid of any significant effect in the nucleus accumbens. RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently reduced the stimulatory effect on striatal and accumbal DA release induced by an acute challenge with nicotine (1 mg/kg, i.p.) in rats treated repeatedly with this alkaloid. However, only the effect of 3 mg/kg RO 60-0175 reached statistical significance. The inhibitory effect of RO 60-0175 on DA release induced by nicotine in the corpus striatum and the nucleus accumbens was completely prevented by SB 242084 (0.5 mg/kg, i.p.) and SB 243213 (0.5 mg/kg, i.p.), two selective antagonists of 5-HT(2C) receptors. It is concluded that selective activation of 5-HT(2C) receptors can block the stimulatory action of nicotine on central DA function, an effect that might be relevant for the reported antiaddictive properties of RO 60-0175.     

Evidence on extracellular dopamine level in rat striatum: implications for the validity of quantitative microdialysis

Microdialysis zero-net-flux (ZNF) method is commonly used to monitor drug-induced changes in neurotransmitter baseline and release/uptake processes. Recent studies in this field suggest that microdialysis ZNF method seriously underestimates the resting concentration of extracellular dopamine in the rat neostriatum because probe implantation preferentially damages nearby dopamine release sites and that dopamine uptake inhibition increases the relative recovery of dopamine by microdialysis. This study assessed the validity of these claims by examining current data on extracellular dopamine levels at rest and after drug application obtained by voltammetry, a technique thought to induce less tissue disruption than microdialysis. To obtain the extracellular baseline value for dopamine from the evoked overflow data, we modified the existing dopamine kinetic model to suit both the resting and stimulated circumstances. It was found that dopamine uptake inhibition did in fact decrease the microdialysis relative recovery of dopamine, implying that the average basal extracellular dopamine level is within the range of 7-20 nm in rat striatum. This study concludes that the microdialysis ZNF method indeed underestimates the extracellular dopamine concentration, although not by as much as had been thought. Chronic microdialysis damages both neurotransmitter release and uptake, but it does so in a somewhat relative and proportional way for both processes. Thus the validity of the microdialysis ZNF method is not seriously undermined.     

An inverse correlation between the apparent rate of dopamine clearance and tonic autoinhibition in subdomains of the rat striatum: a possible role of transporter-mediated dopamine efflux

The dopaminergic terminal field in the rat striatum is compartmentalized into sub-domains that exhibit distinct dynamics of electrically evoked dopamine release. The fast striatal domains, where dopamine release is predominantly vesicular, exhibit conventional dopaminergic activity. However, vesicular dopamine release is tonically autoinhibited in the slow domains, which suggests that dopamine reaches the autoreceptors via a non-vesicular route. Hence, it appears that the domains use distinct mechanisms to regulate the basal dopamine concentration available to activate, or not, pre-synaptic autoinhibitory receptors. However, direct detection of local variations in tonic extracellular dopamine concentrations is not yet possible. So, the present study employed voltammetry to test the hypothesis that the apparent rate of dopamine clearance from the extracellular space should be domain-dependent. The apparent rate of dopamine clearance is equal to the difference in the rates of dopamine release and uptake that determine extracellular dopamine concentrations. This study confirms that the apparent rate of dopamine clearance is slower in the slow striatal domains where vesicular dopamine release is tonically autoinhibited. These findings support the view that the basal concentration in slow domains is maintained by a non-vesicular release process, possibly transporter-mediated efflux.     

Direct Comparison of the Response of Voltammetry and Microdialysis to Electrically Evoked Release of Striatal Dopamine

Abstract: Carbon fiber microelectrodes either were implanted directly into striatal tissue or were mounted into the outlet of microdialysis probes that were implanted into striatal tissue. This allowed voltammetry and microdialysis to be used under identical in vivo experimental conditions to monitor extracellular dopamine levels during electrical stimulation of the medial forebrain bundle both before and after uptake inhibition with nomifensine. The marked differences between the results obtained with each technique cannot be explained on the basis of their inherent analytical attributes (sensitivity, temporal response, etc.). The results demonstrate that the microdialysis recovery factor for endogenous dopamine increases after uptake inhibition, an observation that stands in contradiction to the existing theory and practice of the microdialysis technique. The observations led to the development of a numerical model that rationalizes the observations reported herein and that allows in vivo voltammetry and in vivo microdialysis results to be interpreted within a single theoretical framework.     

Kinetic diversity of dopamine transmission in the dorsal striatum

Dopamine (DA), a highly significant neurotransmitter in the mammalian central nervous system, operates on multiple time scales to affect a diverse array of physiological functions. The significance of DA in human health is heightened by its role in a variety of pathologies. Voltammetric measurements of electrically evoked DA release have brought to light the existence of a patchwork of DA kinetic domains in the dorsal striatum (DS) of the rat. Thus, it becomes necessary to consider how these domains might be related to specific aspects of DA's functions. Responses evoked in the fast and slow domains are distinct in both amplitude and temporal profile. Herein, we report that responses evoked in fast domains can be further classified into four distinct types, types 1–4. The DS, therefore, exhibits a total of at least five distinct evoked responses (four fast types and one slow type). All five response types conform to kinetic models based entirely on first‐order rate expressions, which indicates that the heterogeneity among the response types arises from kinetic diversity within the DS terminal field. We report also that functionally distinct subregions of the DS express DA kinetic diversity in a selective manner. Thus, this study documents five response types, provides a thorough kinetic explanation for each of them, and confirms their differential association with functionally distinct subregions of this key DA terminal field.

     

Role of vesicle pools in action potential pattern-dependent dopamine overflow in rat striatum in vivo

Action potential (AP) patterns and dopamine (DA) release are known to correlate with rewarding behaviors, but how codes of AP bursts translate into DA release in vivo remains elusive. Here, a given AP pattern was defined by four codes, termed total AP number, frequency, number of AP bursts, and interburst time [N, f, b, i].. The 'burst effect' was calculated by the ratio (γ) of DA overflow by multiple bursts to that of a single burst when total AP number was fixed. By stimulating the medial forebrain bundle using AP codes at either physiological (20 Hz) or supraphysiological (80 Hz) frequencies, we found that DA was released from two kinetically distinct vesicle pools, the fast-releasable pool (FRP) and prolonged-releasable pool (PRP), in striatal dopaminergic terminals in vivo. We examined the effects of vesicle pools on AP-pattern dependent DA overflow and found, with given 'burst codes' [b=8, i=0.5 s], a large total AP number [N = 768, f = 80 Hz] produced a facilitating burst-effect (γ[b8/b1] = 126 ± 3%), while a small total AP number [N=96, 80 Hz] triggered a depressing-burst-effect (γ[b8/b1] = 29 ± 4%). Furthermore, we found that the PRP (but not the FRP) predominantly contributed to the facilitating-burst-effect and the FRP played an important role in the depressing-burst effect. Thus, our results suggest that striatal DA release captures pre-synaptic AP pattern information through different releasable pools.     

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 .     

Differential regulation of somatodendritic and nerve terminal dopamine release by serotonergic innervation of substantia nigra

Nigrostriatal dopaminergic neurons release dopamine from dendrites in substantia nigra and axon terminals in striatum. The cellular mechanisms for somatodendritic and axonal dopamine release are similar, but somatodendritic and nerve terminal dopamine release may not always occur in parallel. The current studies used in vivo microdialysis to simultaneously measure changes in dendritic and nerve terminal dopamine efflux in substantia nigra and ipsilateral striatum respectively, following intranigral application of various drugs by reverse dialysis through the nigral probe. The serotonin releasers (+/-)-fenfluramine (100 micro m) and (+)-fenfluramine (100 micro m) significantly increased dendritic dopamine efflux without affecting extracellular dopamine in striatum. The non-selective serotonin receptor agonist 1-(m-chlorophenyl)-piperazine (100 micro m) elicited a similar pattern of dopamine release in substantia nigra and striatum. NMDA (33 micro m) produced an increase in nigral dopamine of a similar magnitude to mCPP or either fenfluramine drug. However, NMDA also induced a concurrent increase in striatal dopamine. The D2 agonist quinpirole (100 micro m) had a parallel inhibitory effect on dopamine release from dendritic and terminal sites as well. Taken together, these data suggest that serotonergic afferents to substantia nigra may evoke dendritic dopamine release through a mechanism that is uncoupled from the impulse-dependent control of nerve terminal dopamine release.     

Transient changes in mesolimbic dopamine and their association with 'reward'

Mesolimbic dopaminergic neurons modulate complex circuitry in the ventral forebrain involved in reward processing, although the precise function of the dopaminergic input is debated. Electrophysiological measurements have revealed that mesolimbic dopaminergic neurons can fire in either tonic or phasic modes, and that phasic firing accompanies the alerting or anticipatory phases of reward. However, the neurochemical relevance of this rapid neuronal discharge within the reward processing circuitry is not yet clear, in part because of difficulty in interpretation of extracellular dopamine measurements. Herein, the nature of the information provided by different neurochemical techniques is critically discussed. Classical methods of monitoring dopamine reveal changes in extracellular dopamine resulting from tonic neuronal activity, but do not have the temporal resolution to distinguish concentration transients. However, recent advances in dopamine sensors now enable transient dopamine concentrations resulting from phasic firing to be positively identified and followed on a physiologically relevant timescale. This has enabled demonstrations of discrete, phasic dopamine signals accompanying rewarding or alerting stimuli. Thus, enhanced dopamine release at terminals appears to be coincident with phasic electrical activity at cell bodies. These accumulating data promise to help unravel the precise role of phasic dopamine transmission in reward processing.     

Nicotine increases dopamine clearance in medial prefrontal cortex in rats raised in an enriched environment

Environmental enrichment results in differential behavioral and neurochemical responsiveness to nicotine. The present study investigates dopamine clearance (CL_DA) in striatum and medial prefrontal cortex (mPFC) using in vivo voltammetry in rats raised in enriched (EC) or impoverished conditions (IC) and administered nicotine (0.4 mg/kg) or saline. Baseline CL_DA in striatum or mPFC was not different between EC and IC. Across repeated DA application, striatal CL_DA increased in saline-control EC and IC. CL_DA increased in mPFC in saline-control IC; CL_DA did not change in saline-control EC. Thus, enrichment differentially alters dynamic responses of the dopamine transporter (DAT) to repeated DA application in mPFC, but not in striatum. In EC, nicotine increased mPFC CL_DA compared to saline-control, but had no effect on CL_DA in IC; nicotine had no effect in striatum in EC or IC. Compared to respective saline-controls, nicotine increased dihydroxyphenylacetic acid content in striatum and mPFC in EC, but not in IC. Nicotine also had no effect on DA content in striatum or mPFC in EC or IC. Results indicate that enrichment eliminated the dynamic response of mPFC DAT to repeated DA application in saline-control and augmented the nicotine-induced increase in DAT function in mPFC, but not in striatum.     

In vivo monitoring of brain neurotransmitter release for the assessment of neuroendocrine interactions

Summary 1. The neurotransmitter mechanisms regulating neuroendocrine processes have been traditionally inferred from the effects of drugs purportedly acting through specific transmitter systems. The direct appraisal of changes in endogenous neuromediators had to rely initially on analyses of brain samples obtained post-morten.2. Currently, a more physiological assessment is available through the monitoring ot the extracellular levels of neurotransmitters and their metabolites in discrete brain areas of living animals. Two methodologies, namely in vivo voltammetry and microdialysis, are being increasingly used for this purpose. This article summarizes their principles, relative merits, and limitations and presents some relevant applications.3. Thus, microdialysis data show a differential response in the amphetamine-induced dopamine release in the nucleus accumbens in adult male and female rats castrated prepuberally. Given their high time-resolution, in vivo electrochemistry techniques seem especially suited for studying the fast, non-genomic effects of steroid hormones. This is illustrated by the voltammetric detection of a rapid release of dopamine in the corpus striatum induced by progesterone in males.4. These methodologies should be regarded as complementary tools for the assessment of the neurochemical correlates of neuroendocrine interactions.     

Ischemic flow threshold for striatal dopamine release in rats

To determine the level of cerebral blood flow reduction which causes striatal dopamine release, extracellular dopamine and cerebral blood flow was simultaneously determined using in vivo brain dialysis and a hydrogen clearance method, respectively, in the striatum of spontaneously hypertensive rats, before and during experimental cerebral ischemia. The ischemic flow threshold for neurotransmitter dopamine release was found to be 20% of the resting value or 8–10 ml/100g/min of cerebral blood flow, being similar to those for energy and membrane failures.     

Microdialysis and mass spectrometric monitoring of dopamine and enkephalins in the globus pallidus reveal reciprocal interactions that regulate movement

Pallidal dopamine, GABA and the endogenous opioid peptides enkephalins have independently been shown to be important controllers of sensorimotor processes. Using in vivo microdialysis coupled to liquid chromatography-mass spectrometry and a behavioral assay, we explored the interaction between these three neurotransmitters in the rat globus pallidus. Amphetamine (3 mg/kg i.p.) evoked an increase in dopamine, GABA and methionine/leucine enkephalin. Local perfusion of the dopamine D(1) receptor antagonist SCH 23390 (100 μM) fully prevented amphetamine stimulated enkephalin and GABA release in the globus pallidus and greatly suppressed hyperlocomotion. In contrast, the dopamine D(2) receptor antagonist raclopride (100 μM) had only minimal effects suggesting a greater role for pallidal D(1) over D(2) receptors in the regulation of movement. Under basal conditions, opioid receptor blockade by naloxone perfusion (10 μM) in the globus pallidus stimulated GABA and inhibited dopamine release. Amphetamine-stimulated dopamine release and locomotor activation were attenuated by naloxone perfusion with no effect on GABA. These findings demonstrate a functional relationship between pallidal dopamine, GABA and enkephalin systems in the control of locomotor behavior under basal and stimulated conditions. Moreover, these findings demonstrate the usefulness of liquid chromatography-mass spectrometry as an analytical tool when coupled to in vivo microdialysis.     

Catecholamine mapping within nucleus accumbens: differences in basal and amphetamine-stimulated efflux of norepinephrine and dopamine in shell and core

The nucleus accumbens is believed to play a critical role in mediating the behavioral responses to rewarding stimuli. Although most studies of the accumbens focus on dopamine, it receives afferents from many other nuclei, including noradrenergic cell groups in the brainstem. We used in vivo microdialysis to measure extracellular levels of both norepinephrine and dopamine in the accumbens shell and core. Regional analysis of shell and core and border regions demonstrated that norepinephrine was high in shell and decreased from medial shell to lateral core, where baseline levels were low or undetectable. Conversely, extracellular dopamine in core was twice the level seen in shell. Both catecholamines increased following a single injection of amphetamine (2 mg/kg, i.p.). The norepinephrine response was greater and long-lasting in shell compared with core. The maximal dopamine response was higher in core than in shell, but the duration of the effect was comparable in both regions. The distinct neurochemical characteristics of shell and core are likely to contribute to the functional heterogeneity of the two subregions. Furthermore, norepinephrine may be involved in many of the functions generally attributed to the accumbens, either directly or indirectly via modulation of extracellular dopamine.     

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.     

Inhibition of NMDA-induced striatal dopamine release and behavioral activation by the neuroactive steroid 3alpha-hydroxy-5beta-pregnan-20-one hemisuccinate

Our laboratory has previously shown that the synthetic neuroactive steroid 3alpha-hydroxy-5beta-pregnan-20-one hemisuccinate (3alpha5betaHS) is a negative modulator of NMDA receptors in vitro. Similarly, 3alpha5betaHS exhibits rapid sedative, analgesic, anticonvulsive, and neuroprotective effects in vivo. Here we report a study designed to investigate whether a negatively charged neuroactive steroid, 3alpha5betaHS, modulates the action of NMDA receptors in vivo. Our results indicate that peripherally administered 3alpha5betaHS enters the CNS and inhibits NMDA-mediated motor activity and dopamine release in the rat striatum. The increase in motor activity induced by intrastriatal microinjection of NMDA was blocked by the systemic administration of 3alpha5betaHS and the NMDA-induced increase in extracellular dopamine in the striatum was also attenuated by both systemically administered and intrastriatally administered (by in vivo microdialysis) 3alpha5betaHS. These data indicate that 3alpha5betaHS acts through striatal NMDA receptors in vivo. When taken together, these results suggest that neuroactive steroids may prove to be effective in the treatment of neurological and psychiatric disorders involving over-stimulation of NMDA receptors in the mesotelencephalic dopamine system.