Hydroxyl radicals release in rat striatum involves metabotropic glutamate receptors

Disruption of glutamate homeostasis frequently leads to oxidative stress and to the release of hydroxyl radicals (radical OH). Here, we investigated, via a microdialysis approach, the possible involvement of metabotropic glutamate receptors in the glutamate-induced release of hydroxyl radicals in adult rat striatum. Glutamate was applied at low amount, resulting in a moderate release that was not inhibited by dizocilpine (MK-801), a specific NMDA receptor antagonist. (RS)-alpha-methyl-4-carboxyphenylglycine (MCPG), a broad spectrum metabotropic antagonist, that does not exert any effect on the basal release of radical OH suppressed their response to glutamate. (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (t-ACPD), a non-selective metabotropic glutamate receptors agonist, promoted an radical OH release almost similar to that observed after glutamate, which was similarly impaired by co-infusion with MCPG. By contrast, infusion of (RS)-3,5-dihydroxyphenylglycine (DHPG), a more specific group I metabotropic glutamate receptors agonist, did not result in any appreciable radical OH response. Thus, beside NMDA receptors, some metabotropic glutamate receptors may also be involved in the glutamate-induced release of hydroxyl radicals.     

The dopamine agonist pramipexole scavenges hydroxyl free radicals induced by striatal application of 6-hydroxydopamine in rats: an in vivo microdialysis study

Hydroxyl free radical production seems to play an important role in the pathogenesis of Parkinson's disease. In the present study, we investigated the dopamine agonists pramipexole and pergolide as well as the nitrone compound S-PBN (N-tert-butyl-alpha-(2-sulfophenyl)nitrone) to reduce hydroxyl radical formation. Microdialysis experiments were carried out in non-anaesthetized Wistar rats. Salicylate was incorporated into the perfusion fluid to measure indirectly hydroxyl radicals indicated by 2,3-dihydroxybenzoic acid (2,3-DHBA). Local perfusion with 0.2 or 2 nmol/2 microl/min 6-hydroxydopamine (6-OHDA) via the microdialysis probe significantly increased 2,3-DHBA levels 14-fold and 47-fold, respectively. Systemic application of either pergolide (0.05 mg/kg) or pramipexole (1 mg/kg) failed to significantly reduce 6-OHDA-induced hydroxyl radical production. In contrast, a 40 min pretreatment with pramipexole (2 and 10 nmol/2 microl/min via the probe) before onset of 6-OHDA perfusion, significantly attenuated 2, 3-DHBA levels compared with vehicle controls. S-PBN pretreatment (2 nmol/2 microl/min) was not effective to reduce 2,3-DHBA levels. In conclusion, pramipexole was able to reduce hydroxyl radical levels induced by 6-OHDA in vivo after local application. This property of pramipexole may be beneficial under conditions of enhanced hydroxyl radical formation in parkinsonian brains and may add to its well known dopamine D(2)-like receptor agonistic effects.     

Effects of pergolide treatment on in vivo hydroxyl free radical formation during infusion of 6-hydroxydopamine in rat striatum

This study focused on the early neurochemical events involved in 6-hydroxydopamine (6-OHDA) neurotoxicity and the putative neuroprotective effects of pergolide. 6-OHDA in 0.1% ascorbic acid/saline was delivered into rat striatum by means of microdialysis and 2,3-dihydroxybenzoic acid (2,3-DHBA) was measured as an index of hydroxyl free radical formation using salicylate trapping. Infusion of 6-OHDA (2–20 mM) via the dialysis probe for 15 min was associated with an immediate and striking increase in the extracellular levels of 2,3-DHBA and dopamine, and this effect was dose-dependent. An infusion of 10 mM 6-OHDA, equivalent to a direct injection of 4 μg free base, resulted in dopamine overflow with a maximum approx. 200-fold above the baseline. This massive overflow of toxic amounts of dopamine, much greater than expected of reuptake inhibition, seems to be the earliest response of nigrostriatal neurones to 6-OHDA. In rats treated with pergolide mesylate (7 days 0.5 mg/kg/day, i.p.), the average amount of 2,3-DHBA associated with 6-OHDA striatal infusion was significantly smaller than that in controls. This suggests that pergolide treatment leads to an increased ability of striatal tissue to quench hydroxyl radical formation in vivo.     

Rapid reduction of ATP synthesis and lack of free radical formation by MPP+ in rat brain synaptosomes and mitochondria

MPTP is a neurotoxin thought to damage dopaminergic neurons through free radical formation. MPTP is metabolized in the brain to MPP(+), which is taken up into dopaminergic neurons via the dopamine transporter and assumed to impair mitochondrial function. We used striatal synaptosomes and telencephalic mitochondria to further investigate MPP(+) mechanism of action. For comparison, the respiratory toxins FCCP, a cyanide analog that uncouples mitochondrial ATP production, and rotenone, a NADH dehydrogenase inhibitor, were also tested. FCCP, MPP(+) and rotenone caused a rapid but stable decrease in [3H]dopamine (DA) uptake by striatal synaptosomes. Two free radical scavengers, the salen-manganese complex EUK-134, and the spin trap s-PBN, did not prevent MPP(+)-induced decrease in DA uptake. However, addition of ATP during synaptosome preparation resulted in partial recovery of MPP(+)-induced [3H]DA uptake decrease. Generation of oxygen free radicals by treatment of telencephalic mitochondria with MPP(+), FCCP, or rotenone, was evaluated by measuring DCF fluorescence, while light emission by the luciferin-luciferase complex was used to determine ATP levels. MPP(+), unlike rotenone, did not produce oxygen free radicals, but rather blocked ATP production in mitochondria, as did FCCP and rotenone. Taken together, these results suggest that MPP(+) toxicity, at least during its initial stages, is primarily due to a decrease in ATP synthesis by mitochondria and not to free radical formation.     

N-methyl-D-aspartate but not glutamate induces the release of hydroxyl radicals in the neonatal rat: modulation by group I metabotropic glutamate receptors

Although they likely involve activation of N-methyl-D-aspartate (NMDA) receptors, the mechanisms giving rise to perinatal hypoxic-ischemic-induced damages remained unclear. The purpose of the present study was to investigate in vivo the mechanisms regulating the glutamate-induced release of toxic hydroxyl radicals (.OH) in neonatal rat. Anesthetized 7-day-old Wistar rat pups bearing a microdialysis cannula implanted in the striatum were perfused with a solution containing salicylate as an.OH trap. Hydroxyl radicals formation was evaluated, after a 3 hr postoperative delay, by measuring the 2,3-DHBA levels by HPLC/EC before, during and over 3 hr after the administration of glutamatergic agonists or antagonists. Administration of NMDA and of ibotenate dramatically increased the efflux of.OH, 17-fold and sixfold, respectively. Glutamate, used at the same concentration did not produce any significant increase in the.OH release and may even decrease this efflux when given at larger concentrations. The NMDA-induced.OH response was partially but progressively reduced by glutamate coinjection and completely blunted by DHPG [(RS)-3, 5-dihydroxyphenylglycine], a group I metabotropic glutamate receptor agonist. Conversely, AIDA [(RS)-1-aminoindan-1,5-dicarboxylic acid], an antagonist of the same receptors, unmasked an.OH response to glutamate. These results are evidence that the glutamate-induced activation of a group I metabotropic glutamate receptor normally protected the neonatal brain from any glutamate activation of NMDA receptor, which otherwise would produce the release of toxic hydroxyl radicals. Targeting group I metabotropic glutamate receptors and/or.OH might contribute to protecting the neonatal brain against perinatal hypoxic-ischemic induced lesions.     

Neuronal nitric oxide synthase inhibition reduces MPP+-evoked hydroxyl radical formation but not dopamine efflux in rat striatum

Summary. Nigral cell degeneration induced by 1-methyl-4-phenyl-1,2,3,6-tertrahydropyridine (MPTP) or its metabolite 1-methy1-4-phenyl pyridinium (MMP⁺) may involve toxicity induced by nitric oxide. In the present study a microdialysis procedure incorporating salicylate hydroxylation was used to measure striatal hydroxyl radical production through the formation of 2,3-dihydroxybenzoic acid (2,3-DHBA). MPP⁺ (5–20 mM for 20 min) increased 2,3-DHBA formation in the rat striatum in a concentration-dependent manner with a concomitant increase in dopamine release and decrease in 3,4-dihydroxyphenyl acetic acid (DOPAC) formation. Inhibition of NO synthesis following N^G-nitro-L-arginine methyl ester (L-NAME; 1 mM) and 7-nitroindazole monosodium salt (7-NINA; 1 mM), but not N^G-nitro-D-arginine methyl ester (D-NAME; 1 mM) attenuated the MPP⁺-induced increase in hydroxyl radical formation. However, neither L-NAME nor 7-NINA had any effect on the MPP⁺-induced increase in dopamine efflux measured in vivo by microdialysis or in vitro using superfused striatal slices, although nomifensine (10 μM) abolished the MPP⁺-evoked dopamine efflux in vitro. These data suggest that NO formation is necessary for the production of hydroxyl radical following MPP⁺ treatment, but is not involved in the MPP⁺-evoked dopamine release. Received September 11, 1998; accepted November 10, 1998     

The biogenic trace amine tyramine induces a pronounced hydroxyl radical production via a monoamine oxidase dependent mechanism: an in vivo microdialysis study in mouse striatum

Tyramine is a biogenic trace amine that releases monoamines and is a good substrate for monoamine oxidase (MAO)-A/B. Here we investigated whether tyramine affects hydroxyl radical formation in the intact and lesioned dopaminergic system. Male C57bl/6 mice received systemic and local tyramine administrations. Hydroxyl radical formation and dopamine (DA) overflow were determined in the striatum using in vivo microdialysis in combination with the salicylate hydroxylation assay. Systemic injection of tyramine neither enhanced extracellular dopamine nor induced hydroxyl radical formation. In contrast, when tyramine was incorporated into the dialysate fluid, hydroxyl radical formation and extracellular dopamine levels were significantly enhanced. Systemic pretreatment with the MAO-A/B inhibitor tranylcypromine or with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) significantly diminished the tyramine-induced hydroxyl radical formation by 73.1% and 80.6%, respectively. We conclude that the mechanism of tyramine-induced hydroxyl free radical formation involves MAO metabolism and requires an intact dopaminergic system. Pharmacological intervention on the MAO-mediated formation of hydroxyl free radicals seems to be a promising strategy to prevent oxidative damage in the nigrostriatal dopaminergic system.     

Comparison of the novel drug Ensaculin with MK-801 on the reduction of hydroxyl radical production in rat striatum after local application of glutamate

Ensaculin interacts with various neurotransmitter systems (e.g., dopaminergic, serotoninergic, glutamatergic) and was originally designed for the treatment of dementia. In the present study Ensaculin was tested for its possible reduction of glutamate-induced hydroxyl free radical formation in vivo. The microdialysis experiment was carried out in non-anaesthetized Wistar rats, which were implanted with a microdialysis probe into the striatum. Salicylate (10 nmol/2 microl/min) was incorporated into the perfusion fluid to measure indirectly hydroxyl radicals indicated by 2,3-dihydroxybenzoic acid (DHBA) formation. After baseline recording, glutamate (100 or 500 nmol/2 microl/min) was perfused through the microdialysis probe (CMA 12, 4 mm, flow rate 2 microl/min). Ensaculin (0.1, 1 and 10 mg/kg), MK-801 (1 mg/kg) or saline was injected i.p. 20 min after the onset of glutamate perfusion (500 nmol/2 microl/min). Glutamate (100 nmol/2 microl/min) and (500 nmol/2 microl/min) perfusion produced a 2.6- and 17-fold increase of 2,3-DHBA, respectively. Treatment with Ensaculin (1 and 10 mg/kg i.p. ) significantly antagonized the formation of 2,3-DHBA, to values of 60.5% and 56.7% of control levels, respectively. In comparison, MK-801 attenuated 2,3-DHBA levels, to values of 65.8% compared to control values. Ensaculin may be useful in the treatment of neurodegenerative disorders associated with elevated hydroxyl free radicals and excitotoxicity.     

Cerebral oligaemia episode triggers free radical formation and late cognitive deficiencies

Sixty minutes of cerebral oligaemic hypoxia, induced by bilateral clamping of the carotid arteries (BCCA) in pentobarbital-anaesthetized normotensive rats, induces a late progressive cognitive decline when compared with sham-operated controls. Analysis at BCCA of hippocampal metabolism using microdialysis showed increased release of glutamate, aspartate and gamma-aminobutyric acid, followed by a progressive rise in the formation of hydroxyl free radicals measured as 2,3-dihydroxybenzoic acid (2,3-DHBA), their reaction product with salicylate, though only in the re-perfusion phase. In the striatum increased dopamine release occurred during BCCA, whereas glutamate and aspartate showed an increase only during the late re-perfusion phase. gamma-Aminobutyric acid (GABA) concentration increased during BCCA and early re-perfusion. An increase in 2,3-DHBA was seen during BCCA, and persisted over 2 h of re-perfusion. Six and 13 months after surgery, though not as early as 3 months, BCCA-treated rats perform worse than sham-operated controls in a water-maze, where decreased swimming speed reveals striatal dysfunction, while hippocampal dysfunction manifested as diminished spatial bias. These results show that cerebral oligaemia, similarly to cerebral ischaemia, leads to increased extracellular dopamine, aspartate and glutamate, and the production of hydroxyl radicals in structures associated with learning and memory processes. Unlike cerebral ischaemia, in cerebral oligaemia the appearance of spatial memory deficits is delayed.     

Methamphetamine-induced neurotoxicity: Roles for glutamate and dopamine efflux

The neurotoxic effect of methamphetamine (METH) on striatal dopaminergic neurons have been hypothesized to be mediated by excess dopamine (DA) release. In addition, N-methyl-D-aspartate (NMDA) receptor antagonists block METH-induced DA depletions. This suggests that glutamate also mediates the toxic effects of METH. The purpose of this study is to demonstrate that DA and glutamate efflux contribute to METH-inducted neurotoxicity. In vivo microdialysis in rats was used to measure extracellular concentrations of striatal DA and glutamate following 3 injections of METH (10 mg/kg, i.p.), each injection given 2 hours apart. One week following the dialysis experiment, rats were sacrificed and the ventral lateral striata were assayed for DA content. Glutamate concentrations in the dialysate increased by over 4-fold after the third METH injection. In these same animals, striatal DA tissue content was significantly depleted. In separate groups of rats, pretreatment with haloperidol (2 mg/kg at the first METH injection) significantly increased METH-induced DA efflux. The haloperidel pretreatment attenuated the extracellular increase in glutamate produced by METH and blocked subsequent neurotoxicity to DA neurons. In contrast, pretreatment with the DA uptake blocker, GBR-12909 (10 mg/kg, 30 min before each METH injection) significantly attenuated the increased DA release produced by METH but did not change glutamate efflux. However, pretreatment with GBR-12909 did protect against the tissue content depletion of DA in the striatum. Based on these findings, it appears that increased DA and glutamate release in the striatum are important and possibly interact in the development of METH-induced neurotoxicity. © 1994 Wiley-Liss, Inc.     

Nuclear factor-kappaB activation is not involved in a MPTP model of Parkinson's disease

In the present study the involvement of hydroxyl free radicals and nuclear factor-kappaB (NF-kappaB) activation was investigated in the MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) model of Parkinson's disease. MPTP (30 mg/kg, s.c.) produced a significant 2-fold increase in hydroxyl free radicals in the striatum of C57BL/6 mice determined by microdialysis in combination with the salicylate hydroxylation assay. Electrophoretic mobility shift assays did not detect NF-kappaB activation after MPTP treatment. Furthermore, p50-deficient mice showed only minor differences in striatal dopamine and metabolite levels as well as tyrosine hydroxylase immunoreactivity after MPTP administration in comparison to wildtype mice. We postulate that, although hydroxyl radical production was enhanced, NF-kappaB plays only a minor role in the MPTP model because neither neurochemical nor immunocytochemical parameters were altered in p50-deficient mice in comparison to controls.     

Involvement of corticostriatal glutamatergic terminals in striatal dopamine release elicited by stimulation of delta-opioid receptors

We have previously shown that striatal dopamine release induced locally by a delta-opioid receptor agonist was totally inhibited by a glutamate N-methyl-D-aspartate receptor antagonist, indicating the involvement of glutamatergic receptors in this effect. The aim of the present study was to specify this mechanism. Firstly, we investigated the effect of [D-Pen2,D-Pen5]-enkephalin (DPDPE) on glutamate release in rats by intrastriatal microdialysis. The infusion of DPDPE (10 microm) enhanced the glutamate content in dialysate by approximately 34%, an effect which did not appear to result from inhibition of glutamate uptake. We then considered the consequences of a unilateral thermocoagulation of the frontal cortex on either glutamate or dopamine release induced by stimulation of delta-opioid receptors 2 days later. This lesion, which decreased the glutamate content in ipsilateral striatum by approximately 30%, totally prevented the increase in dialysate levels of glutamate induced by DPDPE. Moreover, whereas DPDPE (10 microm) was found to increase the striatal dopamine release in intact animals by approximately 59%, this effect was also completely suppressed by the cortical lesion. Finally, we studied the effect of the lesion on the [3H]-DPDPE binding to striatal membranes prepared from the whole striatum. In the ipsilateral striatum a significant decrease in this [3H]-DPDPE binding (by approximately 18%) was found 2 days after the lesion. Our results indicate that the increase in striatal dopamine release induced by DPDPE probably depends on glutamate release from corticostriatal glutamatergic afferents in response to the stimulation of delta-opioid receptors located on terminals of these neurons.     

Regulation of striatal dopamine release by metabotropic glutamate receptors

In vivo microdialysis in conscious rats was used to assess the effect of metabotropic glutamate receptor stimulation on striatal dopamine release. Local application of the metabotropic glutamate agonist (±)-trans-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD), via a microdialysis probe, produced a concentration-dependent response: infusion of 50 μM ACPD did not produce a significant effect on extracellular dopamine levels, while application of 100 μM or 500 μM ACPD increased dopamine release by approximately 50% or 100%, respectively. To examine the contribution of impulse flow and multisynaptic mechanisms to the ACPD-induced increase in dopamine release, 500 μM ACPD were coapplied with 2 μM tetrodotoxin (TTX). An increase in extracellular dopamine levels was observed after the application of 500 μM ACPD, despite the presence of TTX. To further study the actions of metabotropic glutamate receptor-stimulation on terminal release characteristics of dopamine, the effect of ACPD on 40 mM K⁺-stimulated dopamine release was investigated. It was found that application of ACPD reduces dopamine release in response to K⁺ stimulation. These data suggest that during basal conditions, metabotropic glutamate receptor activation facilitates striatal dopamine release, possibly through presynaptic, impulse-independent mechanisms. However, during conditions of hyperstimulation, activation of metabotropic receptors, in contrast to ionotropic receptors, reduces excess dopamine release. Synapse 28:220–226, 1998. © 1998 Wiley-Liss, Inc.     

Glutamate inhibition of NMDA-induced hydroxyl radical release: an ontogenic study in rat.

Hydroxyl radicals (.OH) are frequently associated with glutamate excitotoxicity and may be critical in the occurrence of perinatal brain damage. We thus investigated the mechanisms regulating the glutamate-induced release of toxic.OH during development, using microdialysis and salicylate as an.OH trap. Glutamate inhibited.OH release until post-natal day 14, but stimulated this release from day 21 onwards. DHPG [(RS)-3,5-dihydroxyphenylglycine], a group-I metabotropic glutamate receptor agonist, similarly reduced the.OH release at day 14, but was ineffective afterwards. DHPG also completely blunted the tremendous NMDA-induced.OH release at day 14 but not at day 21. Glutamate itself therefore tonically inhibited a possible free radical release through NMDA channel activation during early development.     

The salicylate hydroxylation assay to measure hydroxyl free radicals induced by local application of glutamate in vivo or induced by the Fenton reaction in vitro

The direct measurement of hydroxyl radicals in vivo is extremely difficult. Therefore, the indirect determination of hydroxyl radicals using salicylate (2-hydroxybenzoate) is widely accepted. Reverse microdialysis with glutamate led to a dose-dependent production of hydroxyl free radicals indicated by the hydroxylation adduct of salicylate, namely 2,3-dihydroxybenzoic acid. The local stimulation of hydroxyl free radical formation seems to be suitable to investigate a radical-scavenging property of potential neuroprotective drugs. In vitro experiments using the Fenton reaction may be a helpful tool to assess whether or not a substance is able to act as a radical scavenger in a cell free environment, which is easy to handle and a simple screening method before in vivo experiments were performed. In the present study we present an in vivo approach using local application of glutamate into the striatum and an in vitro screening using the Fenton reaction to induce hydroxyl radical formation. The main goal is to reliable measure hydroxyl free radicals, which are the most reactive oxygen radicals in biology and medicine.     

Effect of Adenosine A2A Receptor Antagonists and l-DOPA on Hydroxyl Radical, Glutamate and Dopamine in the Striatum of 6-OHDA-Treated Rats

A(2A) adenosine receptor antagonists have been proposed as a new therapy of PD. Since oxidative stress plays an important role in the pathogenesis of PD, we studied the effect of the selective A(2A) adenosine receptor antagonists 8-(-3-chlorostyryl)caffeine (CSC) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) on hydroxyl radical generation, and glutamate (GLU) and dopamine (DA) extracellular level using a microdialysis in the striatum of 6-OHDA-treated rats. CSC (1 mg/kg) and ZM 241385 (3 mg/kg) given repeatedly for 14 days decreased the production of hydroxyl radical and extracellular GLU level, both enhanced by prior 6-OHDA treatment in dialysates from the rat striatum. CSC and ZM 241385 did not affect DA and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) extracellular levels in the striatum of 6-OHDA-treated rats. L-DOPA (6 mg/kg) given twice daily for two weeks in the presence of benserazide (3 mg/kg) decreased striatal hydroxyl radical and glutamate extracellular level in 6-OHDA-treated rats. At the same time, L-DOPA slightly but significantly increased the extracellular levels of DOPAC and HVA. A combined repeated administration of L-DOPA and CSC or ZM 241385 did not change the effect of L-DOPA on hydroxyl radical production and glutamate extracellular level in spite of an enhancement of extracellular DA level by CSC and elevation of extracellular level of DOPAC and HVA by ZM 241385. The data suggest that the 6-OHDA-induced damage of nigrostriatal DA-terminals is related to oxidative stress and excessive release of glutamate. Administration of L-DOPA in combination with CSC or ZM 241385, by restoring striatal DA-glutamate balance, suppressed 6-OHDA-induced overproduction of hydroxyl radical.     

NMDA antagonist effects on striatal dopamine release: microdialysis studies in awake monkeys.

Brain imaging studies have suggested that the NMDA antagonist ketamine is as potent a releaser of striatal dopamine as amphetamine. This conclusion contradicts microdialysis findings in the rodent that NMDA antagonists, in contrast to amphetamine, have little or no effect on striatal dopamine release. The present study addressed two mechanisms that could account for this discrepancy: 1) whether there is a species difference, i.e., rodents vs. primates, in the responsivity of striatal dopamine to NMDA antagonists, and 2) whether rapid uptake of dopamine prevents reliable measures of synaptic dopamine release by microdialysis in response to NMDA antagonists. MRI-directed in vivo microdialysis was used to compare the effects of psychotomimetic NMDA antagonists phencyclidine (PCP), ketamine, and amphetamine on extracellular striatal dopamine levels in awake rhesus monkeys. The effect of PCP was also investigated in the presence of intrastriatally applied nomifensine, a dopamine uptake blocker. Amphetamine (0.1 or 0.4 mg/kg) produced robust and dose-dependent increases in dopamine release ranging 2-10-fold above baseline. PCP at 0.1 mg/kg had no effect and at 0.3 mg/kg produced a small 50% increase over baseline. Ketamine, at the relatively high dose of 5 mg/kg, produced only a 30% increase in dopamine release. Intrastriatal application of nomifensine did not influence the effect of PCP, suggesting that rapid uptake of dopamine is not preventing the detection of a PCP-induced increase in dopamine release. These findings suggest that in the primate, ketamine and PCP are not effective dopamine releasers, as has been suggested by previous imaging studies.     

Glutamate-induced cell death and formation of radicals can be reduced by lisuride in mesencephalic primary cell culture

Summary. Oxidative stress evoked by excitotoxicity is considered an important factor for the loss of dopaminergic neurons in Parkinson’s disease. In vitro, protective effects of the dopamine agonist lisuride on complex I inhibition in primary dopaminergic cell culture have been reported. However, little is known about the effects of lisuride on glutamate-induced radical formation. Here, effects of lisuride on the formation of nitric oxide (NO) and superoxide radicals following glutamate exposure were studied on primary cell cultures prepared from mouse mesencephala. Glutamate treatment resulted in doubling of NO and superoxide radical formation, increased dopaminergic cell degeneration and extensively altered neuronal appearance. Pretreatment with lisuride significantly lowered the levels of either reactive species and increased the survival of dopaminergic neurons compared to glutamate-treated cultures. Moreover, the beneficial effect of lisuride could be completely inhibited by the D2/D3 receptor antagonist sulpiride when co-treated in cultures.     

Partial characterization of kainic acid-induced striatal dopamine release using in vivo microdialysis

The aim of this study was to characterize interactions between striatal kainate (KA) receptors and dopamine (DA) release using in vivo microdialysis. After insertion of a microdialysis probe and establishment of baseline DA release, each preparation was standardized with a pulse of an iso-osmotic solution of 100 mM KCl in Ringer's solution. DA release following pharmacological manipulation was compared to potassium-induced release and expressed as a percent value. In one group of animals, KA (12.5 mM in Ringer's solution) was administered via the microdialysis probe in 2, 3, 5 or 10 min pulses 30 min following standardization with potassium resulting in release of DA which was15.7 ± 3.9, 30.3 ± 11.3, 67.5 ± 15.0and92.9 ± 19.8% of potassium-induced DA release, respectively. Perfusion of CdCl₂ (0.6 mM in Ringer's solution) 30–45 min prior to a 10 min KA pulse significantly reduced KA-induced DA release compared to control values. Intrastriatal administration of kynurenate (Kyn) attenuated KA-induced DA release in a dose-dependent manner. Levels of DA metabolites in striatal perfusates were significantly reduced following KA administration. This effect was partially reversed by cadmium pretreatment but not affected by Kyn pretreatment. Findings of this study indicate that KA induces striatal DA release in a dose-dependent manner, and this effect is at least partially dependent upon activation of calcium channels. Results also indicate dose-dependent inhibition of KA-induced striatal DA release by the excitatory amino acid receptor antagonist, Kyn, suggesting that this compound interacts with striatal KA receptors and that these receptors are involved with modulating striatal DA release in vivo.     

Partial depletion of dopamine in substantia nigra impairs motor performance without altering striatal dopamine neurotransmission

Previous data indicate that the release of somatodendritic dopamine in substantia nigra influences motor activity and coordination, but the relative importance of somatodendritic dopamine release vs. terminal striatal dopamine release remains to be determined. We utilized simultaneous measurement of dopamine neurotransmission by microdialysis and motor performance assessment by rotarod test to investigate the effects of local dopamine depletion in rats. The vesicular monoamine transporter inhibitor tetrabenazine (100 microm) was administered locally in substantia nigra as well as in striatum. Nigral tetrabenazine administration decreased nigral dopamine dialysate concentrations to 7% of baseline and whole-tissue dopamine content by 60%. Nigral dopamine depletion was associated with a reduction in motor performance to 73 +/- 6% of pretreatment value, but did not alter dialysate dopamine concentrations in the ipsilateral striatum. Striatal tetrabenazine administration decreased striatal dopamine dialysate concentrations to 5% of baseline and doubled the somatodendritic dopamine response to motor activity, but it was not associated with changes in motor performance or dopamine content in striatal tissue. Simultaneous treatment of substantia nigra and striatum reduced motor performance to 58 +/- 5% of the pretreatment value. The results of this study indicate that partial depletion of nigral dopamine stores can significantly impair motor functions, and that increased nigral dopamine release can counteract minor impairments of striatal dopamine transmission.