Demonstration of the nigrostriatal projection by silver staining after nigral injections of 6-hydroxydopamine

Lesions of the substantia nigra of rats were made by local injections of 6-hydroxydopamine. The degenerative changes in the brain were studied by means of the Fink-Heimer technique. Survival times of 2 or 3 days were optimal for the demonstration of degeneration. Extensive necrotic changes were observed in neurons of the substantia nigra pars compacta following 6-hydroxydopamine injections; control injections did not produce necrosis. Dense fields of terminal degeneration were observed in the caudate of the 6-hydroxydopamine treated rats but not in the caudates of the control animals. A comparison was made on the amount of degeneration present in the ventral nucleus of the thalamus after three types of lesions: (a) nigral injection of 6-hydroxydopamine; (b) nigral injection of carrier solution (control); and (c) electrolytic lesion of the substantia nigra. Lesions (a) and (b) produced only small amounts of thalamic degeneration, while lesion (c) produced extensive terminal degeneration in the ventral thalamic nucleus. The toxic effects of nigral injections of 6-hydroxydopamine were confined to the dopaminergic neurons of the substantia nigra pars compacta; nondopaminergic neurons of the substantia nigra pars reticulata, and their thalamic projection field, appeared normal in silver stains.     

Distribution of tritium label in the neonate rat brain following intracisternal or subcutaneous administration of [H]6-OHDA. An autoradiographic study

The present report describes the distribution of tritium label after injection of newborn rats with [³H]6-hydroxydopamine ([³H]6-OHDA). The animals were injected either intracisternally (i.c.) or subcutaneously (s.c.), with or without pretreatment with nomifensine, which blocks the high-affinity uptake of both noradrenaline (NA) and dopamine (DA), and sacrificed at intervals from 40 min to 24 h post-injection (p.i.). In i.c. injected animals, tritium label is demonstrable as early as 40 min p.i. in neurons of all known NA and DA cell groups. In NA neurons, it is taken up into cell body, dendrites, preterminal and terminal axons. The intensity of neuronal labeling is highest within the first 4 h p.i. and decreases in most neurons with longer postinjection intervals. A significant proportion of both NA and DA neurons degenerate beginning 6 h p.i., the majority show morphological signs of the axon reaction 24 h p.i. Uptake of [³H]6-OHDA into serotonergic and non-catecholaminergic neurons is not demonstrable.[³H]6-OHDA is accumulated by the following extraneuronal cells of the CNS: ependymal cells, epithelial cells of the choroid plexus, subependymal macrophages, smooth muscle cells in the wall of large intraparenchymal blood vessels, meningeal cells and glial cells. The time course of accumulation and disappearance of the label varies among these extraneuronal elements. The meningeal cells show the highest labeling intensity and degenerate within 24 h p.i.After pretreatment of the animals with nomifensine, the uptake of [³H]6-OHDA into NA and DA neurons is totally blocked; by contrast uptake of the labeled drug into extraneuronal cells is not prevented.These findings show that [³H]6-OHDA is not only accumulated by neurons possessing the high-affinity uptake for NA or DA, but by numerous other, extraneuronal cells which also participate in the metabolism of catecholamines.     

Luteinizing Hormone-Releasing Hormone and Gamma-Aminobutyric Acid Neurons in the Medial Preoptic Area are Synaptic Targets of Dopamine Axons Originating in Anterior Periventricular Areas

The aim of this study was to characterize further the transmitter content and the location of the parent cells of tyrosine hydroxylase-immunoreactive boutons terminating on luteinizing hormone-releasing hormone- and glutamic acid decarboxylase-immunoreactive neurons in the rat medial preoptic area. Electron microscopic immunostaining for luteinizing hormone-releasing hormone, tyrosine hydroxylase or glutamic acid decarboxylase was performed on desipramine-pretreated (to protect norepinephrine and epinephrine axons) rats which received a stereotaxic injection of 6-hydroxydopamine into the medial preoptic area anteroventral periventricular nucleus 48 h prior to sacrifice. This treatment induced acute degeneration of dopamine axon terminals characterized by the development of autophagous cytolysosomes, an early morphological sign of catecholamine axon degeneration. To further define the cells of origin of these dopamine boutons, the anterograde marker Phaseolus vulgaris leucoagglutinin was iontophoretically applied to the zona incerta. Six days later, rats received a 6-hydroxydopamine injection into the zona incerta or the lateral ventricle, and 48 h later, double immunostaining was performed for Phaseolus vulgaris leucoagglutinin and tyrosine hydroxylase, luteinizing hormone-releasing hormone, or glutamic acid decarboxylase on preoptic area vibratome sections. Following the 6-hydroxydopamine injection into the anteroventral periventricular nucleus, autophagous cytolysosome-containing degenerated axons were found in synaptic contact with both luteinizing hormone-releasing hormone and GABA neurons in the medial preoptic area, confirming that these are dopaminergic connections. Following the double injection treatment, 6-hydroxydop-amine-induced degenerated, Phaseolus vulgaris leucoagglutinin-labeled dopamine axons originating in the zona incerta were not found to contact luteinizing hormone-releasing hormone-containing or GABA cells. Instead, many degenerated, Phaseolus vulgaris leucoagglutinin-immunopositive boutons were observed in the dorsomedial and paraventricular nuclei of the hypothalamus. These observations indicate that tyrosine hydroxylase-immunoreactive fibers terminating on the medial preoptic area luteinizing hormone-releasing hormone and GABA cells are dopaminergic and most probably originate from dopamine neurons located in anterior periventricular areas of the hypothalamus.     

Striatal membrane-bound and soluble catechol-O-methyl-transferase after selective neuronal lesions in the rat

Summary Activities of the two forms of catechol-O-methyltransferase (COMT), viz. the soluble (S-COMT) and the membrane-bound (MB-COMT), have been studied in the rat striatum to characterize their localization in relation to the nigrostriatal dopaminergic neurons. Selective unilateral nigrostriatal dopaminergic lesions were produced by an intranigral injection of 6-hydroxydopamine (6-OHDA; 8g/site). 6-OHDA caused an extensive lesion of the dopaminergic neurons as revealed by non-detectable concentrations of dopamine in the striata of the lesioned sites. In spite of that neither S-COMT nor MB-COMT activities were altered in comparison with the intact control striata. The intrastriatal injection of kainic acid significantly increased S-COMT activity but to some extent decreased MB-COMT activity. Kainic acid did not alter the striatal concentration of dopamine.These results suggest that both S-COMT and MB-COMT reside postsynaptically the nigrostriatal dopaminergic neurons. S-COMT seems to be found mainly in striatal glial cells, whereas striatal MB-COMT might be located both in postsynaptic neuronal and extraneuronal cells.     

Effect of the additional noradrenergic neurodegeneration to 6-OHDA-lesioned rats in levodopa-induced dyskinesias and in cognitive disturbances

Parkinson’s disease is a motor and cognitive disorder characterised by a progressive loss of the substantia nigra pars compacta (SNc) dopaminergic neurons as well as of the locus coeruleus (LC) noradrenergic neurons. It has been suggested that LC neurodegeneration might influence levodopa-induced motor disturbances and cognitive performance. We investigated the influence of dopaminergic and noradrenergic lesions on levodopa-induced dyskinesias and on working memory in rats. Two groups of animals were used: (1) rats with a dopaminergic lesion induced by a unilateral administration of the neurotoxin 6-hydroxydopamine (6-OHDA), and (2) rats with a combined lesion of the dopaminergic and noradrenergic systems induced by 6-OHDA and N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), respectively. Dyskinesias were evaluated on days 1, 8, 15 and 22 of chronic levodopa treatment (6 mg/kg, twice at day, i.p.). Working memory was evaluated by a radial-arm maze (1) before lesions, (2) before levodopa administration and (3) after 22 days of levodopa treatment. Total, axial, limb and orofacial dyskinesias not differed significantly between both groups. Working memory tasks worsened in both lesioned groups reaching significance in terms of time of performance (P < 0.05). The number of repeated entries in the same arm (errors) was only significant in the double-lesioned group (P < 0.05). This behaviour was not different from the one observed after chronic levodopa treatment. These results suggest that levodopa-induced dyskinesias in the 6-OHDA-lesioned rats were not affected by the additional noradrenergic lesion, whereas this last condition was sufficient to worse the cognitive performance deficit produced by the dopaminergic lesion.     

Norepinephrine in the interpeduncular nucleus of the rat: normal distribution and the effects of deafferentation

We used correlative biochemical and histochemical methods to examine (1) the norepinephrine (NE) projection from the paired locus coeruleus (LC) to the midline interpeduncular nucleus (IPN) of the adult rat and (2) the ability of the LC to respond to denervation of their target following removal of noradrenergic afferents (6-hydroxydopamine lesions of the LC) or non-noradrenergic afferents (lesion of the paired fasciculi retroflexi(FR]. Histofluorescence revealed that the NE innervation from the two LC to the IPN is symmetric and overlapping. This projection is confined to rostral, central, and intermediate subnuclei and is absent from lateral and dorsal subnuclei. We found no evidence for homotypic collateral sprouting of undamaged LC neurons into the IPN following unilateral LC lesion. Bilateral LC lesions also did not induce sprouting by NE-containing neurons from other systems (e.g. the superior cervical ganglion or the lateral tegmental group) or from those LC neurons that survived the 6-hydroxydopamine lesion. Histofluorescence following bilateral FR lesions confirmed an earlier observation that apparent hyperinnervation of the IPN by LC afferents is elicited following removal of non-noradrenergic afferents. Measurements of the turnover rate of NE in the IPN of control animals and those that received bilateral FR lesions indicate an increased NE content and increased turnover rate of NE in the IPN of lesioned animals. Taken together these results suggest an increased number of NE terminals and an increase in the activity of tyrosine hydroxylase. No change in NE content or turnover rate was seen in the frontal cortex from these same animals. This is consistent with a target-dependent regulation of heterotypic collateral sprouting.     

Purification of mouse schwann cells using neurite-induced proliferation in serum-free monolayer culture

We have recently reported that neonatal mouse dorsal root ganglionic Schwann cells will (i) survive and assume characteristic morphologies in a serum-free, fully defined culture medium (N1 medium), (ii) proliferate extensively in the same N1 medium if neurons are also present and maintained by nerve growth factor, and (iii) display a strong proliferative response to serum even in the absence of neuronal elements, while also undergoing marked changes in their morphology and their associative behavior toward neurites. In this report, we present a detailed procedure, based upon these earlier observations, which yields purified cultures of either neurons plus associated Schwann cells or Schwann cells in the absence of neurons. The procedure utilizes the neuritic mitogen for selective expansion of Schwann cell numbers in serum-free primary cultures, and a secondary culture step involving neuronal removal and additional Schwann cell expansion using the serum mitogen. The procedure requires 9 days for the generation of3−4 × 10⁶ Schwann cells from 12 newborn mice (with a Schwann cell to neuron ratio of 10) and an additional 6–7 days for the generation of a neuron-free secondary population of40 × 10⁶ Schwann cells with less than 3% contamination by identifiable ganglionic fibroblasts.     

Dopamine D2 receptors exert tonic regulation over discrete neurotensin systems of the rat brain

Blockade of dopamine D₂ receptors with either the selective antagonist, sulpiride, or the non-selective antagonist, haloperidol, induces 2- to 3-fold increases in the content of neurotensin-like immunoreactivity in the striatum and the nucleus accumbens of the rat brain. Quantitatively similar increases were also observed (a) in the striatum following selective degeneration of more than 85% of the nigrostriatal dopamine pathway with 6-hydroxydopamine and (b) in both the striatum and the nucleus accumbens after non-selective depletion of brain dopamine using reserpine plus α-methyl-p-tyrosine. Interestingly, treatment of animals with sulpiride or haloperidol, following the depletion of dopamine by either 6-hydroxydopamine or reserpine plus α-methyl-p-tyrosine, did not add to the elevation in neurotensin content of either structure caused by the dopamine depletion alone. These data suggest that an intact dopamine system is required for the neuroleptics to exert effects on individual neurotensin systems. In addition, the same mechanism appears to underlie the responses of the neurotensin pathways to treatments with the neuroleptics or dopamine-depleting drugs. A likely explanation for the effects of neuroleptics and dopamine-depleting drugs is that they eliminate tonic activity on D₂ receptors by basally released dopamine in the striatum and the nucleus accumbens. Supportive evidence for this hypothesis is that concurrent administration of the D₂ receptor agonist, LY 171555, with reserpine, completely blocked the effects of reserpine-induced dopamine depletion on neurotensin systems of the striatum and the nucleus accumbens.     

The effects of drugs which destroy the sympathetic nervous system on the retrograde transport of nerve growth factor

It has been proposed that the drugs (6-hydroxydopamine, guanethidine, vinblastine) which are known to destroy sympathetic neurons in neonatal animals do so by preventing the accumulation of retrogradely transported nerve growth factor (NGF). It was found, consistent with the proposal, that administration of 6-hydroxydopamine (100 mg/kg s.c.) or vinblastine (0.4 mg/kg s.c.) 16 h prior to the administration of [¹²⁵I]NGF completely prevented the accumulation of retrogradely transported [¹²⁵I]NGF in superior cervical ganglia of neonatal rats. Administration of 6-hydroxydopamine or vinblastine to adult rats (where it does not cause sympathetic neuron cell death) didnot completely prevent the retrograde transport of NGF, although 6-hydroxydopamine produced an alteration of the time course of accumulation (early times unaffected, later times depressed). The administration of guanethidine to adult rats (50 mg/kg/day) produced a modest decrease in the accumulation of NGF (40–60%). It would appear, however, that this decrease cannot account for the cytotoxic effects of guanethidine since: (1) sub-cytotoxic doses of guanethidine and non-cytotoxic guanidinium blocking agents also produce modest decreases in the retrograde transport in NGF; and (2) the retrograde transport of [¹²⁵I]NGF is not affected in neonatal animals until after the neurons are clearly damaged. Hence, the data are entirely consistent with the hypothesis that NGF deprivation caused by 6-hydroxydopamine and vinblastine is the mechanism of the cytotoxic effects of these drugs on sympathetic neurons in neonatal animals. Guanethidine destroys sympathetic neurons by some other mechanism.     

Dopaminergic Neuronal Sprouting and Behavioral Recovery in Hemi-Parkinsonian Rats after Implantation of Amnion Cells

Cells obtained from human, monkey, or rat term amnion membrane produce an activity which, in vitro, increases process outgrowth front rat sympathetic neurons and from dopaminergic neurons of the rat ventral mesencephalon. To determine if these cells could induce sprouting of dopaminergic nerve fibers in vivo, the substantia nigra of rats was lesioned unilaterally with 6-hydroxydopamine and live-rat-term amnion cells, or killed-rat-term amnion cells were implanted into the denervated striata. A control group of rats received saline injections into the denervated striata. Rats implanted with live amnion cells had a significant decrease in turning in response to amphetamine. The lesioned and implanted striata of live-amnion-cell-implanted rats contained significantly greater areas of tyrosine hydroxylase-immunoreactive fibers that the lesioned and implanted striatum of rats in the killed-amnion-cell or saline groups. Differences in the area of tyrosine hydroxylase-immunoreactive fibers in the implanted striata or in amphetamine-induced rotation between killed amnion cell-implanted and saline-injected rats did not reach significance. Implants of live amnion cells into the striatum of a parkinsonian animal model can evoke the de novo appearance of dopaminergic fibers in the denervated striatum and behavioral recovery, most likely through a trophic mechanism.     

Persistence of the releasable pool of CCK in the rat nucleus accumbens and caudate-putamen following lesions of the midbrain

Previous studies have identified populations of dopamine neurons in the midbrain that colocalize cholecystokinin some of which project to the nucleus accumbens and caudate-putamen. The contribution of dopamine-colocalized peptide to the total releasable pool of cholecystokinin in these brain regions was investigated using microdialysis. Dopamine, dihydroxyphenylacetic acid and cholecystokinin immunoreactive levels in dialysates of the posterior medial nucleus accumbens and medial caudate-putamen were determined following 6-hydroxydopamine lesions of the ventral tegmental area and substantia nigra or transection of the medial forebrain bundle. An 89–99% depletion in basal extracellular dihydroxyphenylacetic acid and an 87–99% decrease in veratridine-evoked extracellular dopamine levels was observed in the nucleus accumbens and caudate-putamen, 4 weeks after 6-hydroxydopamine lesion. No statistically significant difference was observed between lesioned and control animals in the basal or veratridine-evoked extracellular level of cholecystokinin immunoreactivity in either region. Similarly, transection of the medial forebrain bundle failed to significantly deplete the releasable pool of cholecystokinin immunoreactivity in the nucleus accumbens or caudate nucleus despite 89–99% depletions of dopamine and its metabolite. These data suggest that midbrain dopamine or non-dopaminergic cells are not the primary source of releasable cholecystokinin in the posterior medial nucleus accumbens and medial caudate-putamen measured by microdialysis.     

Differential regulation of glutamate decar☐ylase and preproenkephalin mRNA levels in the rat striatum

The mRNA levels encoding for the enzyme glutamate decar☐ylase (GAD67) and the peptide enkephalin were measured in the striatum of adult and 15 day-old rats by in situ hydridization histochemistry and radioautography after neonatal injections of 6-hydroxydopamine or after acute pharmacological blockade of dopamine receptors with haloperidol or sulpiride. In adult rats injected as neonates with 6-hydroxydopamine or treated with the D₁/D₂ dopamine receptors antagonist, haloperidol, an increase in preproenkephalin and GAD67 mRNA levels was measured in the striatum. The D₂ dopamine receptor antagonist, sulpiride, did not change the mRNA levels of either GAD67 or PPE in the striatum. In 15-day-old rats, neonatal 6-hydroxydopamine or haloperidol treatment resulted in increased preproenkephalin but unchanged GAD67 mRNA levels compared to controls. In these 15-day-old rats, however, sulpiride produced an increase in GAD67 but not preproenkephalin mRNA levels. Intrastriatal injections to adult rats of pertussis toxin which uncouples Gi/Go proteins from their receptors resulted in a dramatic increase in preproenkephalin without concomitant change in GAD67 mRNA levels. Altogether, these results show that GAD67 and preproenkephalin mRNA levels are modulated in parallel in adult but not in 15 day-old rats after 6-hydroxydopamine injections or dopaminergic blockade. In keeping with evidence of a co-localization of GAD67 and preproenkephalin mRNAs in some striatal neurons, the results indicate that these two mRNAs can be differentially regulated in the same neurons. In addition, the differential effect of haloperidol, sulpiride or pertussis toxin on GAD67 and preproenkephalin mRNA levels suggests that these two mRNAs are regulated through different dopamine receptor subtypes.     

New striatal neurons form projections to substantia nigra in adult rat brain after stroke

Previous studies have demonstrated that newborn striatal neurons can functionally integrate with local neural networks in adult rat brain after injury. In the present study, we determined whether these newly generated striatal neurons can develop projections to the substantia nigra, a target of striatal projection neurons. We used 5′-bromodeoxyuridine (BrdU) and a retroviral vector expressing green fluorescent protein (GFP) combined with multiple immunostaining labels of newborn striatal neurons, and nigral microinjection of fluorogold (FG) to trace the striatonigral projection in adult rat brain at different weeks following a transient middle cerebral artery occlusion (MCAO). We found that FG positive (FG⁺) cells could be detected in newly generated neurons (BrdU⁺-NeuN⁺ and GFP⁺-NeuN⁺) in ipsilateral striatum clearly at 12, but not 2 weeks after MCAO. The data suggest that ischemia-induced newborn striatal projection neurons could form long axons that targeted the substantia nigra (striatonigral projection pathway) and that have intact axonal transport from the nerve terminal to cell body. These new striatal neurons express glutamate NR2 and dopamine D2L receptors, which form the molecular basis for responding to the inputs from cortical glutamatergic and nigral dopaminergic projection neurons. Our data provide the first morphological evidence that newborn neurons in the striatum, a non-neurogenic region, can establish new striatonigral neural circuits, important pathways for the maintenance of motor function. These results help us to understand endogenous cellular mechanisms of brain repair, and suggest that increasing adult neurogenesis could be a practical strategy for enhancing the efficacy of rehabilitative therapy in stroke patients.     

Neuroprotective effects of delayed administration of growth/differentiation factor-5 in the partial lesion model of Parkinson's disease

Neurotrophic factors have the potential for therapeutic use in Parkinson's disease (PD) to support the remaining dopaminergic neurons and protect them against the ongoing disease process. We have examined the effects of the neurotrophin growth and differentiation factor-5 (GDF-5) in a rat model of Parkinson's disease, the intrastriatal 6-hydroxydopamine (6-OHDA) lesion. GDF-5 (25 microg) was injected into either the striatum or substantia nigra (SN) of adult rats at 1 or 2 weeks after 6-hydroxydopamine administration. The behavioral effects of GDF-5 treatment were examined in vivo by amphetamine-induced rotational testing. Injection of GDF-5 into the nigra at either 1 or 2 weeks, or into the striatum at 1 week, after the lesion induced significant decreases in rotations. Post-mortem immunocytochemistry after 6 weeks showed that GDF-5 administration into either site protected dopaminergic cell bodies of the nigra when injected at 1 but not 2 weeks after 6-hydroxydopamine. However, no significant protection of striatal dopaminergic fiber density was observed after GDF-5 treatment. This study shows that the delayed administration of a single dose of GDF-5 has significant protective effects on the damaged adult rat nigrostriatal pathway, reinforcing its therapeutic potential for Parkinson's disease.     

Motor stimulant effects of caffeine in 6‐hydroxydopamine‐lesioned rats are dependent on previous stimulation of dopamine receptors: a different role of D1 and D2 receptors

Caffeine has been reported to induce contralateral rotational behaviour in rats bearing a unilateral 6-hydroxydopamine lesion of the dopaminergic nigrostriatal pathway. In order to define the role of dopamine receptors in the mediation of this behaviour, we have evaluated the influence of previous exposure to a dopamine receptor agonist and the importance of the time elapsed from the 6-hydroxydopamine lesion on the rotational behaviour induced by caffeine. Separate groups of rats lesioned with 6-hydroxydopamine 2 weeks previously were exposed to four administrations of the D₁/D₂ receptor agonist apomorphine (0.3 mg/kg s.c.) (primed) or vehicle (drug-naive). Three days later, all rats received caffeine (30 mg/kg s.c.). Drug-naive 6-hydroxydopamine-lesioned rats did not rotate in response to caffeine, while rats primed with apomorphine rotate contralaterally in response to caffeine. When apomorphine priming was paired to the same environment (hemispherical bowls) where rats received caffeine, rotational behaviour was significantly higher than that obtained in rats primed in an unpaired environment (cylinders). Repeated priming with the D₂/D₃ receptor agonist quinpirole (0.2 mg/kg s.c.) induced a totally context-dependent contralateral rotation in response to caffeine, while caffeine contralateral rotation was not dependent from the context after repeated priming with the D₁ agonist SKF 38393 [1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol hydrochloride, 3 mg/kg s.c.]. Caffeine-mediated contralateral rotation was also evaluated in rats lesioned with 6-hydroxydopamine 12 weeks previously and exposed to four administrations of apomorphine or vehicle. As for rats repeatedly exposed to vehicle or apomorphine 2 weeks after 6-hydroxydopamine lesioning, caffeine failed to induce contralateral rotation in drug-naive rats, while it did induce a partially context-dependent contralateral rotation in apomorphine-primed rats. Different from rats receiving apomorphine priming 2 weeks after 6-hydroxydopamine lesioning, in 12 week-lesioned rats, caffeine also induced contralateral rotation after one priming with apomorphine (0.3 mg/kg s.c.), a condition which fails to induce context-dependent rotation. Administration of selective antagonists of A₁ (8-cyclopentyl-1,3-dipropylxanthine), (DPCPX) or A_2A (5-amino-2-(2-furyl)-7-(3-phenylpropyl)-pyrazolo[4,3-e]-1,2,4-triazolo[5c]pirimidine), (SCH 58261) adenosine receptors failed to induce contralateral rotation either alone or in combination in 12 week-6-hydroxydopamine-lesioned rats repeatedly primed with apomorphine. All together, the results indicate that: (i) caffeine does not induce any contralateral rotation in drug-naive 6-hydroxydopamine-lesioned rats; (ii) priming with a dopamine agonist enables caffeine to induce contralateral rotation, this rotation is, however, context independent only after priming with a selective D₁ agonist; (iii) contralateral rotation in response to caffeine is dependent on the time from the 6-hydroxydopamine lesion; (iv) blockade of A₁ and A_2A adenosine receptors with selective antagonists does not induce contralateral rotational behaviour in 6-hydroxydopamine-lesioned rats.     

L-DOPA-induced dyskinesia in the rat is associated with striatal overexpression of prodynorphin- and glutamic acid decarboxylase mRNA

Rats sustaining unilateral near-complete 6-hydroxydopamine lesions of the mesostriatal dopamine pathway received daily injections of 3,4 dihydroxyphenyl-l -alanine (L-DOPA, 8 mg/kg plus 15 mg/kg benserazide) for 3 weeks. During this period, about 50% of the rats gradually developed abnormal involuntary movements, lasting for 2–3 h following each L-DOPA dose. Rats were killed 3 days after the last L-DOPA injection, and sections through the striatum were processed for in situ hybridization histochemistry. Within the L-DOPA-treated group, levels of preproenkephalin (PPE) mRNA, glutamic acid decarboxylase (GAD67) mRNA, and prodynorphin (PDyn) mRNA in the dopamine-denervated caudate-putamen, as well as GAD67 mRNA expression in the globus pallidus ipsilateral to the 6-hydroxydopamine (6-OHDA) lesion, were higher in dyskinetic than non-dyskinetic animals, and positively correlated with the rats' dyskinesia scores. By contrast, striatal preprotachykinin mRNA expression and D2 receptor-radioligand binding were not significantly associated with dyskinesia. Among all these markers, PDyn mRNA levels showed the most pronounced treatment-dependence (three times higher in the L-DOPA-treated group than in saline-injected lesion-only controls), and the strongest correlation with the rats' dyskinesia scores (r² = 0.82). However, a multiple regression equation including the three factors, GAD67 mRNA levels in the GP, GAD67 mRNA in the lateral CPu, and striatal PDyn mRNA, gave a better fit for dyskinesia scores than PDyn mRNA alone (r² = 0.92). The results show that L-DOPA-induced dyskinesia is associated with overexpression of PDyn and GAD67 mRNA in the striatal projection neurons, and GAD67 mRNA levels in the globus pallidus. Due to its treatment-dependent expression, and strong correlation with the associated dyskinetic symptoms, striatal PDyn mRNA, in particular, may play a role in the mechanisms of behavioural sensitization brought about by the drug.     

Neuronal and nonneuronal COX‐2 expression confers neurotoxic and neuroprotective phenotypes in response to excitotoxin challenge

Treating acute brain injuries with COX‐2 inhibitors can produce both neuroprotective and neurotoxic effects. This study investigated the role of COX‐2 in modulating acute brain injury induced by excitotoxic neural damage. Intrastriatal injection of excitotoxin (RS)‐(tetrazole‐5yl) glycine elicited COX‐2 expression in two distinct groups of cells. cortical neurons surrounding the lesion and vascular cells in the lesion core. The vascular COX‐2 was expressed in two cell types, endothelial cells and monocytes. Selective deletion of COX‐2 in vascular cells in Tie2Cre Cox‐2^flox/flox mice did not affect the induction of COX‐2 in neurons after the excitotoxin injection but resulted in increased lesion volume, indicating a neuroprotective role for the COX‐2 expressed in the vascular cells. Selective deletion of monocyte COX‐2 in LysMCre Cox‐2^flox/flox mice did not reduce COX‐2‐dependent neuroprotection, suggesting that endothelial COX‐2 is sufficient to confer neuroprotection. Pharmacological inhibition of COX‐2 activity in Tie2Cre Cox‐2^flox/flox mice reduced lesion volume, indicating a neurotoxic role for the COX‐2 expressed in neurons. Furthermore, COX‐2‐dependent neurotoxicity was mediated, at least in part, via the activation of the EP1 receptor. These results show that Cox‐2 expression induced in different cell types can confer opposite effects. © 2013 Wiley Periodicals, Inc.     

Chronic lesions differentially decrease tyrosine hydroxylase messenger RNA in dopaminergic neurons of the substantia nigra

Long-term effects of lesions were analyzed in terms of gene expression. Nine months after unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta (s. nigra), the remaining dopaminergic (DAergic) neurons (tyrosine hydroxylase (TH) cells determined by immunocytochemistry (ICC] on the lesioned side were atrophic with smaller nucleoli. By in situ hybridization, the DAergic neurons on the lesioned side had a 50% smaller TH-mRNA concentration than on the contralateral non-lesioned side. However, beta-tubulin mRNA concentration in DAergic neurons was unaffected by the lesion. The lesions did not alter TH-mRNA concentration in the contralateral non-lesioned side by comparison with unoperated controls. We propose that chronic lesions have long-term effects on gene expression because of damage sustained during compensatory hyperactivity after the lesion, or because of decreased trophic support from other neurons.