L-DOPA modulation of corpus striatal dopamine and dihydroxyphenylacetic acid output from intact and 6-OHDA lesioned rats

Summary In the present report we examined the differences in in vitro dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) efflux from the corpus striatum (CS) of intact versus 6-hydroxydopamine (6-OHDA) lesioned (in substantia nigra) male rats in response to different doses of two pulse infusions of L-dihydroxyphenylalanine (L-DOPA). In the first experiment, we tested the effects of two 20-min infusions of 5 uM L-DOPA. In the second experiment we repeated this protocol using 50 uM L-DOPA. There was an overall significantly greater output of DA for intact versus 6-OHDA lesioned rats for both doses. Moreover, in Experiment 1, the 5 uM L-DOPA produced a peak DA response to the second infusion which was significantly higher than that of the first infusion in the intact, but not lesioned rats. In Experiment 2, the 50 uM L-DOPA group showed no significant differences in DA output between the two infusions for both intact and lesioned rats. In contrast to DA responses, there were no overall significant differences in DOPAC output between intact and 6-OHDA lesioned rats for both doses. However, for both doses tested, the peak DOPAC output from the second infusion was significantly increased in lesioned, but not intact rats. These data demonstrate that L-DOPA evoked DA and DOPAC output are differentially modulated in intact and 6-OHDA lesioned striatum. The lesions of the striatal dopaminergic system may alter these responses through changes in intraneuronal storage and metabolism of DA following L-DOPA infusion.     

Differential regulation of striatal preproenkephalin mRNA by D1 and D2 dopamine receptors.

The effect of administration of subtype selective dopamine (DA) agonists on the 6-hydroxydopamine (6-OHDA) lesion-induced increase of striatal preproenkephalin (PPE) mRNA was examined by dot-blot hybridization. Eight days following a unilateral 6-OHDA lesion of the substantia nigra pars compacta (SNc), PPE mRNA levels in the ipsilateral striatum were increased approximately two-fold. Administration of the D2 DA agonist, quinpirole, dose-dependently attenuated the 6-OHDA lesion-induced increase in striatal PPE mRNA. The effect of quinpirole was blocked by coadministration of the D2 DA antagonist eticlopride. In contrast, administration of the D1 DA agonist, SKF 38393, either dose-dependently augmented or had no effect on the 6-OHDA lesion-induced increase in striatal PPE mRNA. In the contralateral striatum, administration of quinpirole decreased PPE mRNA, while administration of SKF 38393 increased PPE mRNA compared to sham lesioned control levels. These data suggest the action of DA at D1 and D2 DA receptors differentially regulates striatal PPE mRNA levels and the apparent inhibition of ENK biosynthesis by DA is mediated via an interaction with D2 DA receptors.     

L-DOPA-induced activation of striatal p38MAPK and CREB in neonatal dopaminergic denervated rat: relevance to self-injurious behavior

The destruction of nigrostriatal dopaminergic neurons with 6-hydroxydopamine (6OHDA) during the neonatal period results in dopamine (DA) loss and susceptibility for self-injurious behavior (SIB) when challenged with L-dihydroxyphenylalanine (L-DOPA), via a supersensitive D1 receptor-mediated mechanism. However, there are no changes in D1 receptor binding or mRNA levels, suggesting a potential postreceptor signaling mechanism(s). Here, we examined whether L-DOPA-induced SIB is associated with altered MAPK signaling (p38MAPK, ERK1/2, and JNK) and their nuclear target, CREB. Neonatal dopaminergic lesioned animals were challenged, as adults, with L-DOPA, observed for SIB for 6 hr, and then sacrificed. The data were grouped as follows: control, lesioned rats without SIB (SIB(-)), and lesioned rats that were positive for SIB (SIB(+)). HPLC analysis of striatal extracts revealed a more significant loss of DA and an increase of serotonin in the SIB(+) than in the SIB(-) group. The striatal levels of TH protein were severely decreased, but D1 receptor levels were unaltered in the lesioned groups. These results confirm and extend previous studies indicating that SIB is associated with a near-total loss of DA and TH, an increase in serotonin, and no change in D1 receptor levels. The present studies further revealed that the levels of active phosphorylated forms of p38MAPK and CREB were significantly higher in the SIB(+) group than in the SIB(-) group in the striatum, but not in cortex or olfactory tubercle. The results indicate an induction of striatal p38MAPK and an activation of its nuclear target, CREB, as additional mechanisms in the genesis of L-DOPA-induced SIB.     

偏侧帕金森病大鼠模型的行为,免疫组化和脑微透析的实验研究

评价6-羟多巴胺(6-OHDA)损毁大鼠单侧黑质制备的偏侧帕金森病动物模型。应用6-羟多巴胺损毁SD大鼠单侧黑质制备偏侧PD鼠模型。3周后根据药物诱发试验,TH免疫组化证实模型制作成功。进一步用脑微透析技术结合HPLC-ECD在体检测PD鼠纹状体多巴胺及代谢产物含量。结果:82只大鼠中有36只阿朴吗啡(APO)诱发的旋转次数>7转/min。6-OHDA注射侧黑质DA神经末稍已绝大多数被损毁。6-OHDA损毁侧纹状体多巴胺及代谢产物明显低于健侧(P<0.05,P<0.01)。应用6-OHDA制备的偏侧PD鼠模型是PD研究的理想模型之一。     

Simultaneous microdialysis in striatum and substantia nigra suggests that the nigra is a major site of action of L-dihydroxyphenylalanine in the "hemiparkinsonian" rat.

L-DOPA is frequently used to relieve symptoms of Parkinson's disease (PD), but its use in patients with more advanced PD is complicated by on-off phenomena. We used simultaneous microdialysis of striatum and ipsilateral substantia nigra to characterize changes in extracellular fluid (ecf) levels of dopamine (DA) following systemic treatment with L-DOPA (25 mg/kg as methylester) in awake, normal rats and those with partial (less than 99%) or complete (greater than 99%) DA depleting unilateral lesions of the nigrostriatal pathway (nsp). In normal rats, nigral ecf DA rose 17-fold above baseline after L-DOPA, compared to a 2.6-fold increase in normal striata. Striatal ecf DA rose equally after L-DOPA in all three groups, whereas peak nigral ecf DA in completely lesioned rats was three times that in normal or partially lesioned animals. Peak nigral ecf DA in completely lesioned rats exceeded striatal ecf DA in all groups by almost 2-fold. Activity after L-DOPA was biphasic ("hyperkinetic/bradykinetic") in completely lesioned but not in normal or partially lesioned animals, and the reduced activity occurred 2.5-4 h after L-DOPA at a time when both nigral and striatal ecf DA levels were still elevated. L-DOPA-induced increases in activity were predictable by greater elevations in nigral compared to striatal ecf DA in animals with complete lesions of the nigrostriatal pathway. Post-DOPA reduced activity might result from desensitization of synaptic events mediated by DA receptors; this may underlie DOPA-related on-off phenomena in patients with advanced PD.     

Dopamine receptors: effects of chronic L-dopa and bromocriptine treatment in an animal model of Parkinson's disease

Male albino rats received a stereotaxic injection of 6-hydroxydopamine (6-OHDA) into the right substantia nigra. Animals demonstrating contralateral rotations 2 weeks postoperatively with apomorphine (0.5 mg/kg i.p.) were treated with L-Dopa (55 mg/kg i.p.), bromocriptine (2 mg/kg i.p.), or polyethylene glycol (vehicle) every 12 h for 30 days. Striatal dopamine (DA) receptors were analyzed by Scatchard plot using 3H-spiroperidol (3H-SP) as ligand. 3,4-Dihydroxyphenylacetic acid (DOPAC) and DA were measured by use of high pressure liquid chromatography. 6-OHDA lesions produced a supersensitivity in striatal DA receptors. Chronic L-Dopa or bromocriptine treatment reversed this supersensitivity. Neither lesion nor drug treatment alone or together produced a significant change in affinity (KD) for 3H-SP. Drug treatment alone also had no effect on Bmax. DA and DOPAC levels were reduced by greater than 98% in lesioned striata. Neither drug treatment affected DA or DOPAC levels as compared with controls. These results indicate that chronic administration of either bromocriptine or L-Dopa will reverse the DA receptor denervation supersensitivity in striatum seen following 6-OHDA lesion. This reversal may play a role in the clinical changes seen in Parkinson's disease patients following chronic use of these drugs.     

Vitamin D(3) attenuates 6-hydroxydopamine-induced neurotoxicity in rats

Previous reports have demonstrated that exogeneous administration of glial cell line-derived neurotrophic factor (GDNF) reduces ventral mesencephalic (VM) dopaminergic (DA) neuron damage induced by 6-hydroxydopamine (6-OHDA) lesioning in rats. Recent studies have shown that 1,25-dihydroxyvitamin D(3) (D3) enhances endogenous GDNF expression in vitro and in vivo. The purpose of present study was to investigate if administration of D3 in vivo and in vitro would protect against 6-OHDA-induced DA neuron injury. Adult male Sprague-Dawley rats were injected daily with D3 or with saline for 8 days and then lesioned unilaterally with 6-OHDA into the medial forebrain bundle. Locomotor activity was measured using automated activity chambers. We found that unilateral 6-OHDA lesioning reduced locomotor activity in saline-pretreated animals. Pretreatment with D3 for 8 days significantly restored locomotor activity in the lesioned animals. All animals were sacrificed for neurochemical analysis 6 weeks after lesioning. We found that 6-OHDA administration significantly reduced dopamine (DA), 3,4-dihydroxy-phenylacetic acid (DOPAC) and homovanilic acid (HVA) levels in the substantia nigra (SN) on the lesioned side in the saline-treated rats. D3 pretreatment protected against 6-OHDA-mediated depletion of DA and its metabolites in SN. Using primary cultures obtained from the VM of rat embryos, we found that 6-OHDA or H(2)O(2) alone caused significant cell death. Pretreatment with D3 (10(-10) M) protected VM neurons against 6-OHDA- or H(2)O(2)-induced cell death in vitro. Taken together, our data indicate that D3 pretreatment attenuates the hypokinesia and DA neuronal toxicity induced by 6-OHDA. Since both H(2)O(2) and 6-OHDA may injure cells via free radical and reactive oxygen species, the neuroprotection seen here may operate via a reversal of such a toxic mechanism.     

D1-dopamine receptor agonists selectively activate striatal c-fos independent of rotational behaviour

L-Dopa and dopaminergic agonists selective for the D1- or D2-dopamine receptor subtype induce contraversive rotation in rats which have been unilaterally lesioned with injections of 6-hydroxydopamine (6-OHDA) into the substantia nigra. D-Amphetamine, which releases dopamine from neurones on the unlesioned side of the animal, causes ipsiversive rotation. These increases in rotational behaviour are mediated, at least in part, by dopamine receptors in the striatum. In unilaterally lesioned animals, L-dopa and the D1-selective agonists SKF 38393 and CY 208-243 produce contralateral rotation and induction of the nuclear proto-oncogene c-fos in the lesioned striatum. D-Amphetamine induces both ipsilateral rotation and c-fos activation in the intact striatum. Three lines of evidence, however, dissociate fos induction and rotation. First, LY 171555, a selective D2-dopamine receptor agonist, also induces contraversive rotation but this rotation is not accompanied by c-fos activation in striatum. Second, D1-dopamine agonists produce activation of striatal c-fos even if rotation is prevented by an anaesthetic. Third, rotation induced by injection of SKF 38393 into substantia nigra is not accompanied by c-fos induction. These results suggest a mechanism by which D1-dopamine receptor mechanisms may regulate long-term changes in dopaminergic systems.     

Neurotrophic effects of bone morphogenetic protein-7 in a rat model of Parkinson's disease

Previous studies have demonstrated that pretreatment with bone morphogenetic protein-7 (BMP7) reduces ischemic neuronal injury in vivo. Moreover, exogenous application of BMP7 increases both the number of tyrosine hydroxylase (+) cells and dopamine (DA) uptake in rat mesencephalic cell cultures. The purpose of this study was to investigate the in vivo effects of BMP7 on 6-hydroxydopamine (6-OHDA) induced lesioning of midbrain DA neurons. Adult Fischer 344 rats were anesthetized and injected with BMP7 or vehicle into the left substantia nigra, followed by local administration of 9 microg of 6-OHDA into the left medial forebrain bundle. The lesioned animals that received BMP7 pretreatment, as compared to vehicle/6-OHDA controls, had a significant reduction in methamphetamine-induced rotation 1 month after the surgery. BMP7-pretreatment partially preserved KCl-induced dopamine release in the lesioned striatum and significantly increased TH immunoreactivity in the lesioned nigra and striatum. In summary, our data suggest that BMP7 has neuroprotective and/or neuroreparative effects against 6-OHDA lesioning of the nigrostriatal DA pathway in an animal model of Parkinson's disease (PD).     

Serotonergic sprouting is induced by dopamine-lesion in substantia nigra of adult rat brain

We have previously extracted a serotonin (5-HT) neurotrophic supernatant from the 5,7-DHT lesioned hippocampus. The current study shows that a new 5-HT neurotrophic signal was monitored in the striatum and nigra after DA-denervation. Such a signal may be involved in the heterotypic sprouting. Dopaminergic neurotoxin, 6-hydroxydopamine (6-OHDA), was injected directly into the substantia nigra of adult rats. Two months after surgery, immunocytochemical staining showed that tyrosine hydroxylase (TH)-positive cell bodies had mostly disappeared in the substantia nigra, and TH-positive terminals in the striatum were almost completely depleted. Meanwhile, the 5-HT fibers, which exist in the same areas with low density, sprouted in the nigra as well as in the striatum and became dense. Normally 5-HT fibers innervate the striatum sparsely and the globus pallidus densely with sharp delineation (in the control side), and become dense across both areas with no appreciable delineation (in the lesion side). The increase of 5-HT fibers was more prominent in the posterior than in the anterior striatum. A significant increase in 5-HT and 5-HIAA levels was also evident in the posterior striatum when the decrease in DA level exceeded 90% in the nigra and striatum. In addition, we found that induction of 5-HT sprouting requires a greater than 90% decrease of DA level. Current data support that 6-OHDA injection in the substantia nigra of adult rats triggered a trophic signal or removed an inhibition for the growth of 5-HT neurons which responded with sprouting in the nigra as well as in the striatum.     

Vitamin D3 attenuates 6-hydroxydopamine-induced neurotoxicity in rats

Previous reports have demonstrated that exogeneous administration of glial cell line-derived neurotrophic factor (GDNF) reduces ventral mesencephalic (VM) dopaminergic (DA) neuron damage induced by 6-hydroxydopamine (6-OHDA) lesioning in rats. Recent studies have shown that 1,25-dihydroxyvitamin D₃ (D3) enhances endogenous GDNF expression in vitro and in vivo. The purpose of present study was to investigate if administration of D3 in vivo and in vitro would protect against 6-OHDA-induced DA neuron injury. Adult male Sprague–Dawley rats were injected daily with D3 or with saline for 8 days and then lesioned unilaterally with 6-OHDA into the medial forebrain bundle. Locomotor activity was measured using automated activity chambers. We found that unilateral 6-OHDA lesioning reduced locomotor activity in saline-pretreated animals. Pretreatment with D3 for 8 days significantly restored locomotor activity in the lesioned animals. All animals were sacrificed for neurochemical analysis 6 weeks after lesioning. We found that 6-OHDA administration significantly reduced dopamine (DA), 3,4-dihydroxy-phenylacetic acid (DOPAC) and homovanilic acid (HVA) levels in the substantia nigra (SN) on the lesioned side in the saline-treated rats. D3 pretreatment protected against 6-OHDA-mediated depletion of DA and its metabolites in SN. Using primary cultures obtained from the VM of rat embryos, we found that 6-OHDA or H₂O₂ alone caused significant cell death. Pretreatment with D3 (10⁻¹⁰ M) protected VM neurons against 6-OHDA- or H₂O₂-induced cell death in vitro. Taken together, our data indicate that D3 pretreatment attenuates the hypokinesia and DA neuronal toxicity induced by 6-OHDA. Since both H₂O₂ and 6-OHDA may injure cells via free radical and reactive oxygen species, the neuroprotection seen here may operate via a reversal of such a toxic mechanism.     

Effect of repeated administration of dopamine agonists on striatal neuropeptide mRNA expression in rats with a unilateral nigral 6-hydroxydopamine lesion

Summary Striatal mRNA expression for preproenkephalin (PPE) and preprotachykinin (PPT) was studied in unilateral 6-OHDA lesioned rats treated subchronically with a range of selective and non-selective D-1 or D-2 dopamine (DA) agonists. Apomorphine (5mg/kg sc), pergolide (0.5mg/kg sc), SKF 38393 (5mg/kg sc), SKF 80723 (1.5mg/kg sc), and quinpirole (5mg/ kg sc), or 0.9% saline (150l sc) were all given twice daily (except pergolide: once daily) for 7 days. The abundance of PPE mRNA was not altered by any of these DA agonists in the intact striatum contralateral to the 6-OHDA lesion. Only apomorphine and quinpirole increased the abundance of PPT mRNA in the intact striatum. In saline treated 6-OHDA lesioned animals PPE mRNA was elevated (+160%, p < 0.005) and PPT mRNA decreased (–36%, p < 0.005) in the denervated striatum. The up-regulation of striatal PPE mRNA in the lesioned striatum was reversed only by pergolide. The downregulation of striatal PPT mRNA in the lesioned striatum was reversed only by apomorphine. The differential sensitivity of the striatal PPE message to the long-acting DA agonist pergolide, and of the striatal PPT message to the mixed D-1/D-2 DA agonist apomorphine suggests that the striatopallidal enkephalinergic pathways are mainly regulated by prolonged DA receptor stimulation, whereas the striatonigral substance P pathways are mainly regulated by mixed D-1/D-2 DA receptor stimulation.     

Differential modulation of striatonigral dynorphin and enkephalin by dopamine receptor subtypes

Previous studies indicate that dopaminergic transmission inhibits the biosynthesis of enkephalin and stimulates that of dynorphin in the striatonigral pathway of intact rat. The purpose of this study was to determine which dopamine (DA) receptor subtype(s) mediate the modulatory actions of DA. We measured striatal and nigral levels of enkephalin and dynorphin in: (1) intact rats repeatedly injected with D1 (SKF-38393, 5 mg/kg, i.p.) or D2 (LY-171555, 1 mg/kg, i.p.) agonists, alone or in combination, (2) 6-hydroxydopamine (6-OHDA)-lesioned rats repeatedly injected with the same D1 or D2 agonists, and (3) intact rats repeatedly injected with D1 (SCH-23390, 0.05 mg/kg, s.c.) or D2 (sulpiride, 100 mg/kg, s.c.) antagonists, given alone or in combination with the mixed D1/D2 agonist apomorphine (5 mg/kg, i.p.). Repeated injections of the D1 agonist to intact rats (twice daily for 7 days) produced a small but not statistically significant increase in striatal levels of dynorphin; similar treatment with the D2 agonist did not affect dynorphin levels at all. Combined treatments with D1 and D2 agonists did not potentiate the effect of the D1 agonist. 6-OHDA lesions of the nigrostriatal DA pathway alone decreased the level of dynorphin in both the striatum and substantia nigra. However, repeated D1 agonist, but not D2, injections not only reversed the decrease in dynorphin levels, but caused a significant increase above control levels. In intact rats, repeated injections of the D1 or D2 antagonist alone failed to alter the levels of dynorphin, but the D1 antagonist, not the D2 antagonist, attenuated the apomorphine-induced increase in striatal dynorphin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Substantia nigra 6-hydroxydopamine lesions alter dopaminergic synaptic markers in the nucleus basalis magnocellularis and striatum of rats

Recent findings have demonstrated the existence of dopaminergic (DA) markers in the nbM of the human brain and a reduction of these markers in both the nbM and the striatum of patients suffering from Alzheimer's disease (AD). To investigate the source of the DA synaptic markers found in the nbM, rats received unilateral 6-OHDA lesions of the substantia nigra pars compacta (SNc). The SNc lesions caused significant reductions in DA and DOPAC but not HVA in the nbM and the striatum; 3H-sulpiride binding to D2 receptors ipsilateral to the SNc lesion was significantly increased in the striatum (16%), consistent with denervation supersensitivity, but single-point analysis showed no significant changes in the nbM. These data suggest that the decreases in DA and 3H-spiperone binding levels observed in the nbM of AD patients may be due to partial destruction of DA nbM afferent projections from the brainstem.     

Cerebrospinal dopamine metabolites in rats after intrastriatal administration of 6-hydroxydopamine or 1-methyl-4-phenylpyridinium ion

Dopamine (DA) and its main cerebral metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were measured in striatum and cerebrospinal fluid (CSF) from cisterna magna in rats bilaterally lesioned by intrastriatal administration of 6-hydroxydopamine (6-OHDA) or 1-methyl-4-phenylpyridinium ion (MPP⁺). 6-OHDA caused a progressive lesion in striatum that is only moderately reflected in the decrease in dopamine metabolite concentration in CSF. MPP⁺ caused an acute but less selective lesion in the dopamine striatal system, as indicated by a significant reduction in striatal GABA content, followed by a slow recovery in dopamine striatal metabolism and content. The locomotor activity was dramatically reduced in both groups 48 hours after the treatment but remained significantly decreased after two months only in 6-OHDA lesioned animals. A positive correlation was found between HVA CSF concentration and striatal DA content in MPP⁺ lesioned rats, but not in 6-OHDA lesioned rats. It is concluded that the concentration of dopamine metabolites in CSF can be altered only after a severe striatal lesion: reduction of striatal dopamine content below 50% of normal values and involvement of neuronal or non-neuronal elements other than the dopaminergic system, similarly to the lesions caused by MPP⁺. These results may partly explain why CSF dopamine metabolites concentrations were significantly decreased both in advanced stages of parkinsonism and in other neurodegenerative disorders.     

Intracerebral dialysis monitoring of striatal dopamine release and metabolism in response to L-DOPA

Summary We have used intracerebral dialysis to monitor the striatal extracellular fluid (ECF) in rats with unilateral lesions of the nigrostriatal dopamine (DA) pathway. Dialysis samples were collected before and after L-dihydroxyphenylalanine (L-DOPA) administration both in the presence and absence of carbidopa, an extracerebral DOPA decarboxylase (DDC) inhibitor. The baseline ECF levels of DA, 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) were always higher in the intact than in the lesioned striata. In the normal striata, dopamine (DA) concentrations increased following L-DOPA administration. Pretreatment with carbidopa prolonged the duration of the DA increase. In the lesioned striata, DA levels increased following L-DOPA administration only in animals pretreated with carbidopa. Following L-DOPA administration, striatal HVA and DOPAC levels increased considerably more in animals not pretreated with carbidopa than they did in pretreated animals. This increase was particularly marked in the lesioned striata and leads us to conclude that extracerebrally produced HVA and DOPAC can enter the brain extracellular space.     

The adaptation of enkephalin, tachykinin and monoamine neurons of the basal ganglia following neonatal dopaminergic denervation is dependent on the extent of dopamine depletion.

This study examined whether dopamine (DA) is necessary for the normal development of striatal enkephalin and striatonigral tachykinin peptide systems. The neurotoxin, 6-hydroxydopamine (6-OHDA) was used to induce DA deficiency on the third day of the postnatal period in Sprague-Dawley rat pups. The animals were sacrificed at 60 days of age. The levels of Met5-enkephalin (ME) and substance P (SP) were determined by radioimmunoassay and preproenkephalin (PPE) and preprotachykinin (PPT) mRNA abundance in the striatum were assessed by hybridization analysis. The concentrations of DA, 5-hydroxytryptamine (5-HT) and their acid metabolites were determined by high-pressure liquid chromatography with electrochemical detection. The lesioned animals were grouped on the basis of the degree of loss of DA, and changes in ME, SP and 5-HT systems were correlated with respect to the degree of DA loss. The nature and extent of the changes in these systems were dependent on the degree of DA depletion. A loss of more than 90% DA was necessary to result in increased levels of ME and its PPE mRNA and reduced levels of SP and its PPT mRNAs; however, increased levels of 5-HT could be observed at a lower degree of DA loss. The results indicate that the normal development of enkephalin and tachykinin and 5-HT systems of basal ganglia are dependent on the availability of DA and/or the integrity of the nigrostriatal dopaminergic neurons. The results are relevant to our further understanding of the neurobiology of DA deficiency disorders.     

Intraventricular microdialysis: a new method for determining monoamine metabolite concentrations in the cerebrospinal fluid of freely moving rats

A new method is described to estimate the cerebrospinal fluid (CSF) concentrations of monoamine metabolites (dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA] in the lateral ventricle of freely moving rats by use of in vivo microdialysis. Both the baseline concentrations of these metabolites and the rate of dopamine (DA) turnover (estimated by the accumulation of total DA metabolites after 200 mg/kg probenecid) were within the range reported when other methods were used to sample CSF. A series of preliminary studies were conducted to demonstrate that this method can be used to repeatedly sample CSF, and to show that the method is sensitive to local changes in dopaminergic activity induced by lesions, drugs or grafts. (1) Unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra produced a significant decrease in the CSF concentrations of DOPAC and HVA ipsilateral to the lesion, relative to the contralateral side or to concentrations in animals without lesions. (2) When left and right lateral ventricles were sampled simultaneously in animals with a unilateral 6-OHDA lesion, haloperidol induced an increase in DOPAC and HVA concentrations in CSF on both sides of the brain. Interestingly, the haloperidol-induced increase in CSF concentrations of DA metabolites was greater adjacent to the intact striatum of rats with unilateral 6-OHDA lesions than in animals with no lesion. (3) Finally, in animals with adrenal medulla tissue grafted into the lateral ventricle there was an increase in the CSF concentration of DOPAC compared to pregraft values or to those of animals with control grafts.     

Glial cell line-derived neurotrophic factor (GDNF) gene delivery protects dopaminergic terminals from degeneration

Previously, we observed that injection of an adenoviral (Ad) vector expressing glial cell line-derived neurotrophic factor (GDNF) into the striatum, but not the substantia nigra (SN), prior to a partial 6-OHDA lesion protects dopaminergic (DA) neuronal function and prevents the development of behavioral impairment in the aged rat. This suggests that striatal injection of AdGDNF maintains nigrostriatal function either by protecting DA terminals or by stimulating axonal sprouting to the denervated striatum. To distinguish between these possible mechanisms, the present study examines the effect of GDNF gene delivery on molecular markers of DA terminals and neuronal sprouting in the aged (20 month) rat brain. AdGDNF or a control vector coding for beta-galactosidase (AdLacZ) was injected unilaterally into either the striatum or the SN. One week later, rats received a unilateral intrastriatal injection of 6-OHDA on the side of vector injection. Two weeks postlesion, rats injected with AdGDNF into either the striatum or the SN exhibited a reduction in the area of striatal denervation and increased binding of the DA transporter ligand [(125)I]IPCIT in the lesioned striatum compared to control animals. Furthermore, injections of AdGDNF into the striatum, but not the SN, increased levels of tyrosine hydroxylase mRNA in lesioned DA neurons in the SN and prevented the development of amphetamine-induced rotational asymmetry. In contrast, the level of T1 alpha-tubulin mRNA, a marker of neuronal sprouting, was not increased in lesioned DA neurons in the SN following injection of AdGDNF either into the striatum or into the SN. These results suggest that GDNF gene delivery prior to a partial lesion ameliorates damage caused by 6-OHDA in aged rats by inhibiting the degeneration of DA terminals rather than by inducing sprouting of nigrostriatal axons.     

Striatal L-dopa metabolism studied in vivo in rats with nigrostriatal lesions

Summary We have used cerebral dialysis to monitor striatal metabolism of exogenously administered L-dopa (L-dihydroxyphenylalanine) in rats with unilateral lesions of the substantia nigra. The concentration of extracellular dopamine (DA) increased in both striata following L-dopa administration but the increase was markedly attenuated in the lesioned striatum. The formation of dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), the major DA metabolites, was also reduced in the lesioned striata following L-dopa administration; however, the reduction was not as great as was that of DA formation. A significant metabolism of exogenous L-dopa to 3-O-methyldopa occurred in both striata. L-dopa administration transiently increased extracellular levels of 5-hydroxyindoleacetic acid (5 HIAA) in both the lesioned and intract striata.These results suggest that the striatum with a reduction in DA nerve terminals is deficient both in the capacity to synthesize DA and in the storage mechanisms necessary to protect the newly synthesized DA from oxidative metabolism.