The striatal cholinergic system in L-dopa-induced dyskinesias.

Cholinergic signaling plays a key role in regulating striatal function. The principal source of acetylcholine in the striatum is the cholinergic interneurons which, although low in number, densely arborize to modulate striatal neurotransmission. This modulation occurs via strategically positioned nicotinic and muscarinic acetylcholine receptors that influence striatal dopamine, GABA and other neurotransmitter release. Cholinergic interneurons integrate multiple striatal synaptic inputs and outputs to regulate motor activity under normal physiological conditions. Consequently, an imbalance between these systems is associated with basal ganglia disorders. Here, we provide an overview of how striatal cholinergic interneurons modulate striatal activity under normal and pathological conditions. Numerous studies show that nigrostriatal damage such as that occurs with Parkinson's disease affects cholinergic receptor-mediated striatal activity. This altered cholinergic signaling is an important contributor to Parkinson's disease as well as to the dyskinesias that develop with L-dopa therapy, the gold standard for treatment. Indeed, multiple preclinical studies show that cholinergic receptor drugs may be beneficial for the treatment of L-dopa-induced dyskinesias. In this review, we discuss the evidence indicating that therapeutic modulation of the cholinergic system, particularly targeting of nicotinic cholinergic receptors, may offer a novel approach to manage this debilitating side effect of dopamine replacement therapy for Parkinson's disease.

Evidence for a role for α6(∗) nAChRs in l-dopa-induced dyskinesias using Parkinsonian α6(∗) nAChR gain-of-function mice.

l-Dopa-induced dyskinesias (LIDs) are a serious side effect of dopamine replacement therapy for Parkinson's disease. The mechanisms that underlie LIDs are currently unclear. However, preclinical studies indicate that nicotinic acetylcholine receptors (nAChRs) play a role, suggesting that drugs targeting these receptors may be of therapeutic benefit. To further understand the involvement of α6ß2(∗) nAChRs in LIDs, we used gain-of-function α6(∗) nAChR (α6L9S) mice that exhibit a 20-fold enhanced sensitivity to nAChR agonists. Wildtype (WT) and α6L9S mice were lesioned by unilateral injection of 6-hydroxydopamine (6-OHDA, 3µg/ml) into the medial forebrain bundle. Three to 4wk later, they were administered l-dopa (3mg/kg) plus benserazide (15mg/kg) until stably dyskinetic. l-dopa-induced abnormal involuntary movements (AIMs) were similar in α6L9S and WT mice. WT mice were then given nicotine in the drinking water in gradually increasing doses to a final 300µg/ml, which resulted in a 40% decline AIMs. By contrast, there was no decrease in AIMs in α6L9S mice at a maximally tolerated nicotine dose of 20µg/ml. However, the nAChR antagonist mecamylamine (1mg/kg ip 30min before l-dopa) reduced l-dopa-induced AIMs in both α6L9S and WT mice. Thus, both a nAChR agonist and antagonist decreased AIMs in WT mice, but only the antagonist was effective in α6L9S mice. Since nicotine appears to reduce LIDs via desensitization, hypersensitive α6ß2(∗) nAChRs may desensitize less readily. The present data show that α6ß2(∗) nAChRs are key regulators of LIDs, and may be useful therapeutic targets for their management in Parkinson's disease.

Compensation in pre-synaptic dopaminergic function following nigrostriatal damage in primates.

Clinical symptoms of Parkinson's disease only become evident after 70-80% reductions in striatal dopamine. To investigate the importance of pre-synaptic dopaminergic mechanisms in this compensation, we determined the effect of nigrostriatal damage on dopaminergic markers and function in primates. MPTP treatment resulted in a graded dopamine loss with moderate to severe declines in ventromedial striatum (approximately 60-95%) and the greatest reductions (approximately 95-99%) in dorsolateral striatum. A somewhat less severe pattern of loss was observed for striatal nicotinic receptor, tyrosine hydroxylase and vesicular monoamine transporter expression. Declines in striatal dopamine uptake and transporter sites were also less severe than the reduction in dopamine levels, with enhanced dopamine turnover in the dorsolateral striatum after lesioning. The greatest degree of adaptation occurred for nicotine-evoked [(3)H]dopamine release from striatal synaptosomes, which was relatively intact in ventromedial striatum after lesioning, despite > 50% declines in dopamine. This maintenance of evoked release was not due to compensatory alterations in nicotinic receptor characteristics. Rather, there appeared to be a generalized preservation of release processes in ventromedial striatum, with K(+)-evoked release also near control levels after lesioning. These combined compensatory mechanisms help explain the finding that Parkinson's disease symptomatology develops only with major losses of striatal dopamine.

Enhanced striatal opioid receptor-mediated G-protein activation in L-DOPA-treated dyskinetic monkeys.

Long-term l-3,4-dihydroxyphenylalanine (L-DOPA) treatment in Parkinson's disease leads to dyskinesias in the majority of patients. The underlying molecular mechanisms for L-DOPA-induced dyskinesias (LIDs) are currently unclear. However, the findings that there are alterations in opioid peptide mRNA and protein expression and that opioid ligands modulate dyskinesias suggest that the opioid system may be involved. To further understand its role in dyskinesias, we mapped opioid receptor-stimulated G-protein activation using [35S]guanylyl-5'-O-(gamma-thio)-triphosphate ([35S]GTPgammaS) autoradiography in the basal ganglia of normal and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned squirrel monkeys administered water or L-DOPA. Subtype-selective opioid receptor G-protein coupling was investigated using the mu-opioid agonist [D-Ala, N-Me-Phe, Gly-ol]-enkephalin, delta-agonist SNC80 and kappa-agonist U50488H. Our data show that mu-opioid receptor-mediated G-protein activation is significantly enhanced in the basal ganglia and cortex of L-DOPA-treated dyskinetic monkeys, whereas delta- and kappa-receptor-induced increases were limited to only a few regions. A similar pattern of enhancement was observed in both MPTP-lesioned and unlesioned animals with LIDs suggesting the effect was not simply due to a compromised nigrostriatal system. Opioid receptor G-protein coupling was not enhanced in non-dyskinetic L-DOPA-treated animals, or lesioned monkeys not given L-DOPA. The increases in opioid-stimulated [35S]GTPgammaS binding are directly correlated with dyskinesias. The present data demonstrate an enhanced subtype-selective opioid-receptor G-protein coupling in the basal ganglia of monkeys with LIDs. The positive correlation with LIDs suggests this may represent an intracellular signaling mechanism underlying these movement abnormalities.

Selective recovery of striatal 125I-alpha-conotoxinmii nicotinic receptors after nigrostriatal damage in monkeys.

Evidence suggests that nicotinic receptors play a role in nigrostriatal function, a finding that may be relevant to Parkinson's disease. Knowledge of the conditions that regulate nicotinic receptor expression is therefore important. Previous studies showed that several different nicotinic receptors, including alpha-conotoxinMII (alpha-CtxMII)-sensitive receptors, are decreased after nigrostriatal damage. Nigrostriatal dopaminergic terminals also demonstrate a capacity for recovery after lesioning. The present experiments were therefore done to determine whether there were changes in striatal nicotinic receptors with recovery. To address this, we used two well-characterized animal models of nigrostriatal damage produced using the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Studies in mice showed that striatal 125I-alpha-CtxMII, as well as 125I-epibatidine and 125I-A85380 binding sites significantly recovered 1 month after lesioning, suggesting that alpha6* and most likely alpha4* receptors are increased. Experiments were next done in monkeys since striatal 125I-alpha-CtxMII receptors constitute a large percentage of nicotinic receptors and are more vulnerable to nigrostriatal damage in this model that closely mirrors Parkinson's disease. In monkeys allowed to recover from the toxic effects of MPTP for a 1-2 year period, there was a significant improvement in the Parkinson disability score. There was also a reversal in lesion-induced declines in striatal alpha-CtxMII-sensitive receptors, but no significant change in 125I-epibatidine and 125I-A85380 receptors. These findings suggest that alpha3*/alpha6* sites are selectively increased in monkey striatum with recovery. The present data show that recovery of 125I-alpha-CtxMII receptors occurs in parallel with the dopamine transporter, indicating that these nicotinic receptors sites are localized to presynaptic dopamine terminals in both species.

Loss of alpha-conotoxinMII- and A85380-sensitive nicotinic receptors in Parkinson's disease striatum.

Multiple nicotinic receptors are present in rodent and monkey striatum, with a selective localization of alpha-conotoxinMII-sensitive sites in the striatum and preferential declines in their numbers after nigrostriatal damage. Here we report the presence of 125I-alpha-conotoxinMII and alpha-conotoxinMII-sensitive 125I-epibatidine nicotinic receptors in human control and Parkinson's disease striatum. 125I-alpha-ConotoxinMII bound to control striatum with the characteristics of a nicotinic receptor ligand although the number of sites was approximately fivefold lower than in rodent and monkey. Competition analyses of alpha-conotoxinMII with 125I-epibatidine showed that toxin-sensitive sites comprised approximately 15% of nicotinic receptors in human striatum. In Parkinson's disease caudate, there was a approximately 50% decline in 125I-alpha-conotoxinMII sites with a similar decline in the dopamine transporter. In putamen, there were substantially greater losses of the dopamine transporter (80-90%) but only 50-60% decreases in 125I-alpha-conotoxinMII sites with corresponding declines in alpha-conotoxinMII-sensitive 125I-epibatidine sites, 125I-epibatidine (multiple) sites and 125I-A85380 (beta2-containing) nicotinic receptors. The greater loss of the transporter compared with nicotinic sites suggests that only a subpopulation of nicotinic receptors is located pre-synaptically on striatal dopaminergic neurons in man. Correlation analyses between changes in nicotinic receptors and the dopamine transporter in Parkinson's disease striatum suggest that alpha-conotoxinMII-sensitive 125I-epibatidine sites (low-affinity sites), 125I-A85380 and 125I-epibatidine sites are localized in part to dopaminergic terminals. In summary, these results show that alpha-conotoxinMII-sensitive sites are present in human striatum and that there are high- and low-affinity subtypes which are both decreased in Parkinson's disease.

Increases in striatal preproenkephalin gene expression are associated with nigrostriatal damage but not L-DOPA-induced dyskinesias in the squirrel monkey.

Changes in preproenkephalin expression in the caudate and putamen have been linked to the development of L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesias in primate models of Parkinson's disease, although not all investigators have been able to confirm this association. Because nigrostriatal damage per se is associated with increases in striatal preproenkephalin mRNA levels, it is difficult to know if changes in transcript levels are a result of lesioning or concurrent L-DOPA treatment and resulting dyskinesias. To circumvent these difficulties, we measured striatal preproenkephalin mRNA levels in monkeys with L-DOPA-induced dyskinesias both with and without lesions of the nigrostriatal system. The latter model is not confounded by morphological and biochemical changes resulting from nigrostriatal damage. Monkeys were gavaged with L-DOPA (15 mg/kg) twice daily for a 2-week period and killed 3 days after treatment. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment alone resulted in an increase in preproenkephalin mRNA levels as previously shown. However, striatal transcript levels were similarly elevated in dyskinetic MPTP-lesioned animals treated with L-DOPA. In unlesioned animals, preproenkephalin mRNA levels were also similar in control and L-DOPA-treated dyskinetic monkeys. Because drug-induced changes in mRNA may not be sustained for a prolonged period after treatment, a second series of experiments were done in which animals were killed 3-4 h after the last dose of L-DOPA, but the results were similar to those obtained after 3 days. These data show that, while elevations in striatal preproenkephalin mRNA levels are associated with nigrostriatal damage, they are not linked to the development of L-DOPA-induced dyskinesias. These results thus question the importance of preproenkephalin mRNA in the pathogenesis of this disabling complication of L-DOPA therapy in Parkinson's disease.

Differential nicotinic receptor expression in monkey basal ganglia: effects of nigrostriatal damage.

Our previous work showed that there were marked declines in (125)I-alpha-conotoxin MII labeled nicotinic receptors in monkey basal ganglia after nigrostriatal damage, findings that suggest alpha3/alpha6 containing nicotinic receptors sites may be of relevance to Parkinson's disease. We now investigate whether there are differential changes in the distribution pattern of nicotinic receptor subtypes in the basal ganglia in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned animals compared to controls to better understand the changes occurring with nigrostriatal damage. To approach this we used (125)I-alpha-conotoxin MII, a marker for alpha3/alpha6 nicotinic receptors, and (125)I-epibatidine, a ligand that labels multiple nicotinic subtypes. The results demonstrate that there were medial to lateral gradients in nicotinic receptor distribution in control striatum, as well as ventromedial to dorsolateral gradients in the substantia nigra, which resembled those of the dopamine transporter in these same brain regions. Treatment with MPTP, a neurotoxin that selectively destroys dopaminergic nigrostriatal neurons, led to a relatively uniform decrease in nicotinic receptor sites in the striatum, but a differential effect in the substantia nigra with significantly greater declines in the ventrolateral portion. Competition analysis in the striatum showed that alpha-conotoxin MII sensitive sites were primarily affected after lesioning, whereas multiple nicotinic receptor populations were decreased in the substantia nigra. From these data we suggest that in the striatum alpha3/alpha6 nicotinic receptors are primarily localized on dopaminergic nerve terminals, while multiple nicotinic receptor subtypes are present on dopaminergic cell bodies in the substantia nigra. Thus, if activation of striatal nicotinic receptors is key in the regulation of basal ganglia function, alpha3/alpha6-directed nicotinic receptor ligands may be more relevant for Parkinson's disease therapy. However, nicotinic receptor ligands with a broader specificity may be more important if receptors in the substantia nigra play a dominant role in controlling nigrostriatal activity.

Stimulation of non-alpha7 nicotinic receptors partially protects dopaminergic neurons from 1-methyl-4-phenylpyridinium-induced toxicity in culture.

Previous work has shown that nicotine treatment protects against nigrostriatal degeneration in rodents, findings that may be of relevance to the decreased incidence of Parkinson's disease in cigarette smokers. In the present studies, we investigated the effect of nicotine against 1-methyl-4-phenylpyridinium-induced toxicity in dopaminergic ventral mesencephalic cultures to identify the nicotinic receptor population that may be involved. [3H]Epibatidine, a ligand that binds to receptors containing alpha2-alpha6 subunits, bound to at least two populations of sites that were up-regulated by nicotine in a time and dose dependent manner. We next examined the effect of nicotine on cultures treated with 1-methyl-4-phenylpyridinium, a neurotoxin that selectively damages nigrostriatal dopaminergic neurons. Pre-treatment with nicotine, at 10(-7)-10(-4) M, partially prevented the toxin-induced decline in dopaminergic cells. Pre-exposure to nicotine for 24 h resulted in optimal protection, suggesting that receptor up-regulation may contribute to the observed neuroprotective effect. Nicotine-mediated protection was blocked by pre-incubation with the nicotinic receptor antagonist d-tubocurarine (10(-4) M), but not the alpha7 receptor-selective antagonist alpha-bungarotoxin (10(-7) M). Our results show that nicotinic receptor activation partially protects nigral dopaminergic neurons from 1-methyl-4-phenylpyridinium-induced toxicity in culture and that this appears to occur through an interaction at non-alpha7 containing receptors.

Nicotine administration reduces striatal MPP+ levels in mice.

Nicotine administration has previously been shown to attenuate nigrostriatal damage in animal models of Parkinson's disease, including the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mouse. The present experiments were done to determine whether nicotine may be exerting its effects by altering striatal levels of 1-methyl-4-phenylpyridinium (MPP+), the active metabolite of MPTP. Mice were injected with nicotine (0.33-1 mg/kg i.p.) 10 min prior to MPTP (30 mg/kg s.c.) followed by three subsequent doses of nicotine at 15-min intervals according to a dose schedule previously shown to be neuroprotective. The mice were sacrificed 1.5, 4 and 8 h after MPTP administration and striatal MPP+ levels measured. Nicotine administration (0.33-1.0 mg/kg) resulted in a time-dependent decline in striatal MPP+ levels that was significantly enhanced over that in saline injected animals. Experiments done to examine the effect of age showed that the decrease was observed in older (8-10 months) but not young (6-8 weeks) mice, a finding which may explain some of the variability in the effect of nicotine in the MPTP-induced model of nigrostriatal degeneration. In summary, these results suggest that nicotine may exert its neuroprotective action against nigrostriatal degeneration, at least in part, by decreasing striatal MPP+ levels.

Levodopa induces dyskinesias in normal squirrel monkeys.

This study assessed whether or not levodopa induces dyskinesias in normal (ie, unlesioned) squirrel monkeys. All six animals treated twice daily with levodopa (15 mg/kg with carbidopa by oral gavage) for two weeks developed choreoathetoid dyskinesias, whereas none of the vehicle-treated animals displayed any abnormal movements. These dyskinesias did not merely reflect a generalized motor activation as locomotion was actually suppressed. The present data demonstrate that preexisting nigrostriatal damage is not necessary for the development of levodopa-induced dyskinesias.

Vulnerability of 125I-alpha-conotoxin MII binding sites to nigrostriatal damage in monkey.

Parkinson's disease, a neurodegenerative movement disorder characterized by selective degeneration of nigrostriatal dopaminergic neurons, affects approximately 1% of the population over 50. Because nicotinic acetylcholine receptors (nAChRs) may represent an important therapeutic target for this disorder, we performed experiments to elucidate the subtypes altered with nigrostriatal damage in parkinsonian monkeys. For this purpose we used (125)I-alpha-conotoxin MII (CtxMII), a relatively new ligand that identifies alpha3 and/or alpha6 subunits containing nAChR subtypes. In brain from untreated monkeys, there was saturable (125)I-alpha-CtxMII binding to a single population of high-affinity nicotinic sites (K(d) = 0.9 nm), primarily localized in the visual, habenula-interpeduncular, and nigrostriatal-mesolimbic pathways. Administration of the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine resulted in damage to the nigrostriatal system and parkinsonism. Autoradiographic analysis showed that (125)I-alpha-CtxMII sites were selectively reduced (>/=99%) in the basal ganglia and that the lesion-induced decreases correlated well with declines in the dopamine transporter, a marker of dopaminergic neuron integrity. These findings may indicate that most or all of (125)I-alpha-CtxMII-labeled nAChR subtypes in the basal ganglia are present on nigrostriatal dopaminergic neurons, in contrast to (125)I-epibatidine sites. These data suggest that the development of ligands directed to nAChR subtypes containing alpha3 and/or alpha6 subunits may yield a novel treatment strategy for parkinsonian patients with nigrostriatal dopaminergic degeneration.

Reliability and validity of a new global dyskinesia rating scale in the MPTP-lesioned non-human primate.

Behavioral rating scales for dyskinesia in the non-human primate are frequently used to assess the efficacy of new treatments and to provide a clinical correlative with neurochemical and neuropathological changes. Although a large variety of different scales have been used in non-human primate studies, there is no single standardized scale, and none have been evaluated for reliability and validity. We are reporting a new global non-human primate dyskinesia rating scale (GPDRS) for the squirrel monkey, developed in the context of an independent study of dyskinesia. In this report we demonstrate the reliability and validity of this scale. The GPDRS is a single-item scale with well-defined points and brevity allowing for rapid and easy application for assessing the overall degree of dyskinesia. In this study, seven MPTP-lesioned and four non-lesioned (control) non-human primates were videotaped following treatment with either levodopa or water. To test inter- and intra-rater reliability, three examiners rated the videotape independently at two different time points and these assessments were compared. The validity of the scale was tested in two phases. First, examiners rated the videotape using the GPDRS and the Abnormal Involuntary Movement Scale (AIMS), a scale commonly used to rate dyskinesia in the non-human primate, and the ratings from each scale were compared. Second, validity was tested in the context of an independent dyskinesia study, in which the scale was used to distinguish between two treatment groups. The GPDRS was shown to have high inter- and intra-rater reliability and to be valid for the assessment of dyskinesia in the squirrel monkey. In this report we also demonstrate the inter- and intra-rater reliability of the AIMS.

Autoradiographic analysis of dopamine receptor-stimulated [(35)S]GTPgammaS binding in rat striatum.

Autoradiographic analysis of [(35)S]GTPgammaS binding was used to investigate functional activation of dopamine receptors in rat striatum. Dopamine-stimulated [(35)S]GTPgammaS binding was observed with a maximal increase of 38% over basal activity. A similar stimulatory response was obtained with the D(2) agonist quinpirole, but not SKF-238393, a D(1) agonist. The effect of dopamine was blocked by the D(2) antagonist raclopride, but was unaffected by SCH-23990, a D(1) antagonist. There appeared to be a differential distribution of dopamine-stimulated [(35)S]GTPgammaS binding, with the lowest activity obtained in the medial portion of the caudal striatum. These results demonstrate, using an autoradiographic approach, (i) that dopamine stimulated [(35)S]GTPgammaS binding in the rat striatum occurs through activation of D(2) receptors, and (ii) that the effects of dopamine activation vary in different areas of the rat striatum.

Differential alterations in nicotinic receptor alpha6 and beta3 subunit messenger RNAs in monkey substantia nigra after nigrostriatal degeneration.

Our previous studies showed that alpha4, alpha6, alpha7, beta2, beta3 and beta4 nicotinic acetylcholine receptor messenger RNAs are present in monkey substantia nigra, with a particularly intense and localized labelling of the alpha6 and beta3 subunit messenger RNAs to this brain region. Because loss of nigrostriatal neurons is a central feature of Parkinson's disease and evidence suggests that nicotinic agonists potentiate antiparkinsonian effects of L-dopa, experiments were done to determine whether nicotinic receptor subunit messenger RNAs and binding sites were altered in the basal ganglia after nigrostriatal degeneration. Squirrel monkeys (Saimiri sciureus) were rendered parkinsonian by systemic injection of the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine. Behavioral studies showed that this treatment decreased baseline motor activity to 36+/-11% of control. One month after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine administration, caudate and putamen dopamine levels were reduced to 51+/-15% and 43+/-6% of control, respectively, while the number of tyrosine hydroxylase-positive neurons in the substantia nigra was 75+/-6% of control. Despite the reduction in nigral cell number after nigrostriatal degeneration, there were no changes in alpha4, alpha7, beta2 and beta4 messenger RNA levels in the substantia nigra. In contrast, alpha6 mRNA levels were significantly increased (143+/-10%) and the beta3 transcript decreased (62+/-6%) in the substantia nigra after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Declines were also observed in [125I]epibatidine binding in both the caudate-putamen and substantia nigra, with no change in alpha7 receptor binding. These results may suggest a dissociation in the regulation of receptor messenger RNA and binding sites, and/or that there are differential alterations in the different receptor subtypes measured using [125I]epibatidine. The changes in the two nicotinic receptor subunit messenger RNAs, alpha6 and beta3, which exhibit a selective localization to the substantia nigra, may indicate that nicotinic receptors containing these subunits are altered after nigrostriatal degeneration.

Nitric oxide is involved in acetylcholinesterase inhibitor-induced myopathy in rats.

Excess activation of muscle nicotinic acetylcholine receptors due to genetic mutations, as seen in slow channel congenital myasthenic syndrome, or acetylcholinesterase (AChE) inhibition results in muscle cell degeneration. Our recent work showed that nitric oxide synthase (NOS) inhibitors prevent nicotine-induced muscle cell death in culture. In the present study, we examined the effects of NOS inhibition on nicotinic receptor-mediated myopathy in vivo. Rats injected with the AChE inhibitor paraoxon demonstrate a 90-fold increase in the number of dying muscle cells compared with control as evidenced histologically by centralized nuclei and the presence of degenerating profiles. Coadministration of the nonspecific NOS inhibitor nitro-L-arginine methyl ester or the neuronal NOS-specific inhibitor 7-nitroindazole dramatically reduced the presence of such degenerating profiles to approximately 20% of that seen with paraoxon alone. These results show that inhibition of NOS, as well as neuronal NOS, significantly reduces AChE inhibitor-induced muscle cell degeneration, suggesting that increased nitric oxide production mediates such myopathy.

Autoradiographic analysis of N-methyl-D-aspartate receptor binding in monkey brain: effects of 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine and levodopa treatment.

The anatomic distribution of N-methyl-D-aspartate receptors was investigated in the squirrel monkey brain using quantitative autoradiography with [125I]MK-801 as the radioligand. A heterogeneous distribution of [125I]MK-801 binding sites was observed, with the most intense expression in the outer cortex, hippocampus, olfactory tubercle, caudate and putamen. High levels were also observed in the thalamus, nucleus accumbens and inner cortex, with moderate levels in the claustrum. Relatively low expression levels were detected in the subthalamic nucleus with no apparent binding in the globus pallidus and the substantia nigra. Characterization of striatal [125I]MK-801 binding yielded a B(max) of 63.5 fmol/mg tissue and K(d) of 0.53 nM in the caudate, with similar values for the putamen. Experiments were subsequently performed to compare striatal [125I]MK-801 binding in the following four experimental groups: (i) control animals injected with saline; (ii) monkeys treated with levodopa; (iii) animals rendered parkinsonian after exposure to the neurotoxicant 1-methyl-4-phenyl-1, 2,3,6-tetrahydropyridine; and (iv) dyskinetic monkeys treated with both 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and levodopa. No changes were observed in 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine-lesioned animals compared with the saline control group. However, administration of levodopa to either unlesioned or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated monkeys resulted in a significant decrease in [125I]MK-801 binding in both the caudate and putamen. The data indicate that levodopa exerts a modulatory effect on the striatal glutamatergic system and suggest that a down-regulation of N-methyl-D-aspartate receptors by levodopa, combined with a deficiency in nigrostriatal dopamine function, may play a role in the development of levodopa induced dyskinesias.

Localization of nicotinic receptor subunit mRNAs in monkey brain by in situ hybridization.

Nicotinic receptors are implicated in memory, learning, locomotor activity, and addiction. Identification of the specific receptor subtypes that mediate these behaviors is essential for understanding their role in central nervous system (CNS) function. Although expression of nicotinic receptor transcript has been studied in rodent brain, their localization in the monkey CNS, which may be a better model for the human brain, is not yet known. We therefore investigated the distribution of alpha4, alpha6, alpha7, beta2, beta3, and beta4 receptors subunit mRNAs in the monkey brain by using in situ hybridization. alpha4 and alpha7 mRNAs were very widely expressed, with a substantial degree of overlap in their distribution, except for the reticular nucleus of the thalamus in which alpha7 mRNA was much more prominently expressed. beta2 and beta4 mRNA were also widely distributed, although beta4 was more prominently localized in thalamic nuclei than beta2. The distribution of alpha6 and beta3 mRNA was very distinct from that of the other transcripts, being restricted to catecholaminergic nuclei, the cerebellum, and a few other areas. Although there were similarities in distribution of the nicotinic receptor subunit mRNAs in monkey and rodent brain, there were prominent differences in areas such as the caudate, putamen, locus coeruleus, medial habenula, and cerebellum. In fact, the distribution of alpha4 and alpha7 mRNAs in the monkey caudate and putamen was more similar to that reported in the human than rodent brain. These findings have implications for the development of drug therapies for neurological disorders, such as Alzheimer's and Parkinson's disease, in which nicotinic receptors are decreased.

Relationship among nigrostriatal denervation, parkinsonism, and dyskinesias in the MPTP primate model.

Presynaptic denervation is likely to play an important role in the pathophysiology of dyskinesias that develop after levodopa administration to patients with Parkinson's disease. In this study, the thresholds of nigrostriatal damage necessary for the occurrence of parkinsonism and levodopa-induced involuntary movements were compared in squirrel monkeys lesioned with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Animals treated with a regimen of MPTP that caused parkinsonism displayed > or =95% striatal dopamine depletion, 90% reduction of striatal dopamine uptake sites, and 70% nigral neuronal loss. Levodopa administration ameliorated the parkinsonian signs of these monkeys but also induced dyskinesias. A separate group of animals was treated with a milder MPTP regimen that caused 60%-70% striatal dopamine depletion, a 50% decrease in dopamine transporter, and 40% loss of dopaminergic nigral neurons. While these monkeys displayed no behavioral signs of parkinsonism, they all became dyskinetic after levodopa administration. The priming effect of levodopa, that is, the recurrence of dyskinesias in animals previously exposed to the drug, was compared in severely versus mildly lesioned monkeys. When severely injured parkinsonian animals underwent a second cycle of levodopa treatment, they immediately and consistently developed involuntary movements. In contrast, the recurrence of dyskinesias in primed monkeys with a partial nigrostriatal lesion required several levodopa administrations and remained relatively sporadic. The data indicate that moderate nigrostriatal damage which does not induce clinical parkinsonism predisposes to levodopa-induced dyskinesias. Once dyskinesias have been induced, the severity of denervation may enhance the sensitivity to subsequent levodopa exposures.

Expression of D(3) receptor messenger RNA and binding sites in monkey striatum and substantia nigra after nigrostriatal degeneration: effect of levodopa treatment.

D(3) receptors are prominently localized in the primate caudate-putamen, and D(3) receptor agonist properties may offer an advantage in Parkinson's disease therapy. In the present experiments, we investigated the relationship between D(3) receptor mRNA, D(3) receptor sites and the dopamine transporter in monkey basal ganglia by comparing their distribution in the brain of control and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys (Samirai sciureus). In control monkeys, D(3) receptor mRNA appears to be widely expressed throughout the brain, with a distribution similar to that observed in both man and rodent. D(3) receptors are present in areas which express mRNA but also in some which do not, an observation which suggests they may be both pre- and postsynaptic in the monkey brain. Chronic MPTP administration, which selectively destroys the nigrostriatal system, resulted in a 70 to 99% depletion of the dopamine transporter in the basal ganglia. Autoradiographic analysis showed that after MPTP treatment there was a significant decline in D(3) receptors in the caudate, but not putamen, globus pallidus, substantia nigra or other dopaminergic regions. D(3) receptor mRNA expression was not changed in any region after nigrostriatal lesioning. Two weeks of L-3,4-dihydroxyphenylalanine (levodopa, L-DOPA) treatment, which alleviated Parkinsonism but also induced dyskinesias, reversed the MPTP-induced decline in caudate D(3) receptors. These results show that there is a selective decline in D(3) receptors in the caudate after nigrostriatal degeneration, which is reversed by L-DOPA treatment. Since the majority of dopaminergic nerve terminals were destroyed after MPTP lesioning, the reversal in D(3) receptors after L-DOPA treatment may represent an increase in caudate postsynaptic receptors, which could conceivably contribute to an imbalance in striatal circuitry and the development of dyskinesias.