MPTP-treated Common Marmosets Research Articles

The adenosine A2A receptor antagonist/inverse agonist, KW-6356 enhances the anti-parkinsonian activity of L-DOPA with a low risk of dyskinesia in MPTP-treated common marmosets

The adenosine A2A receptor antagonist/inverse agonist, KW-6356 has been shown to be effective in Parkinson's disease (PD) patients as monotherapy and as an adjunct therapy to L-3,4-dihydroxyphenylalanine (L-DOPA)/decarboxylase inhibitor. However, the effects of KW-6356 combined with L-DOPA on anti-parkinsonian activity and established dyskinesia has not been investigated in preclinical experiments. We examined the effects of combination of KW-6356 with L-DOPA in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmosets. Oral administration of KW-6356 (1 mg/kg) enhanced the anti-parkinsonian activities of various doses of L-DOPA (2.5–10 mg/kg). In MPTP-treated common marmosets primed with L-DOPA to show dyskinesia, KW-6356 (1 mg/kg) also enhanced the anti-parkinsonian activities of various doses of L-DOPA (1.25–10 mg/kg) but not dyskinesia. Chronic co-administration of KW-6356 (1 mg/kg) with a low dose of L-DOPA (2.5 mg/kg) for 21 days increased the degree of dyskinesia induced by the low dose of L-DOPA, but the amplitude of dyskinesia induced by combined administration of KW-6356 (1 mg/kg) with L-DOPA (2.5 mg/kg) was lower than that induced by an optimal dose of L-DOPA (10 mg/kg). These results suggest that KW-6356 can be used to potentiate the effects of a wide range of L-DOPA doses with a low risk of dyskinesia for the treatment of PD.

Anti-parkinsonian activity of the adenosine A2A receptor antagonist/inverse agonist KW-6356 as monotherapy in MPTP-treated common marmosets

KW-6356 is a novel adenosine A2A receptor antagonist/inverse agonist that not only blocks binding of adenosine to adenosine A2A receptor but also inhibits the constitutive activity of adenosine A2A receptor. The efficacy of KW-6356 as both monotherapy and an adjunct therapy to L-3,4-dihydroxyphenylalanine (L-DOPA)/decarboxylase inhibitor in Parkinson's disease (PD) patients has been reported. However, the first-generation A2A antagonist istradefylline, which is approved for use as an adjunct treatment to L-DOPA/decarboxylase inhibitor in adult PD patients experiencing OFF episodes, has not shown statistically significant efficacy as monotherapy. In vitro pharmacological studies have shown that the pharmacological properties of KW-6356 and istradefylline at adenosine A2A receptor are markedly different. However, the anti-parkinsonian activity and effects on dyskinesia of KW-6356 in PD animal models and the differences in the efficacy between KW-6356 and istradefylline are unknown. The present study investigated the anti-parkinsonian activity of KW-6356 as monotherapy in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmosets, and its efficacy was directly compared with that of istradefylline. In addition, we investigated whether or not repeated administration of KW-6356 induced dyskinesia. Oral administration of KW-6356 reversed motor disability in a dose-dependent manner up to 1 mg/kg in MPTP-treated common marmosets. The magnitude of anti-parkinsonian activity induced by KW-6356 was significantly greater than that of istradefylline. Repeated administration of KW-6356 induced little dyskinesia in MPTP-treated common marmosets primed to exhibit dyskinesia by prior exposure to L-DOPA. These results indicate that KW-6356 can be a novel non-dopaminergic therapy as monotherapy without inducing dyskinesia in PD patients.

The effect of Banisteriopsis caapi (B.caapi) on the motor deficits in the MPTP-treated common marmoset model of Parkinson's disease.

Banisteriopsis caapi (B.caapi) contains harmine, harmaline, and tetrahydroharmine, has monoamine oxidase inhibitory activity, and has reported antiparkinsonian activity in humans when imbibed as a tea; however, its effects are poorly documented. For this reason, motor function was assessed in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated common marmosets following administration of B.caapi extract (28.4-113.6mg/kg; po), harmine (0.1 and 0.3mg/kg; sc), and selegiline (10mg/kg; sc), alone or with a submaximal dose of L-3,4-dihydroxyphenylalanine (L-DOPA; 4-7mg/kg). L-DOPA reversed motor disability, increased locomotor activity, and induced moderate dyskinesia. B.caapi did not increase locomotor activity or induce dyskinesia but at 56.8 and 113.6mg/kg improved motor disability. The L-DOPA response was unaltered by co-administration of B.caapi. Harmine (0.1 and 0.3mg/kg) produced a mild improvement in motor disability without affecting locomotor activity or dyskinesia but had no effect on the L-DOPA-induced antiparkinsonian response. Selegiline (10mg/kg) alone improved motor function to the same extent as L-DOPA, but with only mild dyskinesia, and did not alter the response to L-DOPA, although dyskinesia was reduced. The findings suggest that B.caapi alone has a mild antiparkinsonian effect but does not enhance the L-DOPA response or reduce dyskinesia.

Altered detrusor contractility in MPTP-treated common marmosets with bladder hyperreflexia

Bladder hyperreflexia is a common non-motor feature of Parkinson’s disease. We now report on the contractility of the isolated primate detrusor strips devoid of nerve input and show that following MPTP, the amplitude and frequency of spontaneous contraction was increased. These responses were unaffected by dopamine D1 and D2 receptor agonists A77636 and ropinirole respectively. Contractions by exogenous carbachol, histamine or ATP were similar and no differences in the magnitude of noradrenaline-induced relaxation were seen in detrusor strip obtained from normal and MPTP-treated common marmosets (Callithrix jacchus). However, the neurogenic contractions following electrical field stimulation of the intrinsic nerves (EFS) were markedly greater in strips obtained from MPTP treated animals. EFS evoked non-cholinergic contractions following atropine were also greater but the contribution of the cholinergic innervation as a proportion of the overall contraction was smaller in the detrusor strips of MPTP treated animals, suggesting a preferential enhancement of the non-cholinergic transmission. Although dopaminergic mechanism has been proposed to underlie bladder hyperreflexia in MPTP-treated animals with intact bladder, the present data indicates that the increased neurogenically mediated contractions where no extrinsic innervation exists might be due to long-term adaptive changes locally as a result of the loss of the nigrostriatal output.

The adenosine A2A receptor antagonist, istradefylline enhances anti-parkinsonian activity induced by combined treatment with low doses of L-DOPA and dopamine agonists in MPTP-treated common marmosets

The adenosine A2A receptor antagonist, istradefylline improves motor function in patients with advanced Parkinson's disease (PD) optimally treated with a combination of l-DOPA and a dopamine agonist without increasing the risk of troublesome dyskinesia. However, the effects of istradefylline on motor function when administered in combination with low dose of l-DOPA and dopamine agonists as occurs in early PD are unknown. We investigated whether istradefylline enhances the combined anti-parkinsonian effects of a suboptimal dose of l-DOPA and a threshold dose of either the non-ergot dopamine agonist, ropinirole or the ergot dopamine agonist, pergolide in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmoset.Threshold doses of ropinirole (0.025–0.075mg/kg p.o.) and pergolide (0.01mg/kg p.o.) produced a weak anti-parkinsonian effect. Co-administration of a suboptimal dose of l-DOPA (2.5mg/kg p.o.) with threshold doses of the dopamine agonists enhanced their anti-parkinsonian effect that led to increased ‘ON’ time without dyskinesia appearing. Administering istradefylline (10mg/kg p.o.) with the threshold doses of dopamine agonists and the suboptimal dose of l-DOPA in a triple combination caused a further enhancement of the anti-parkinsonian response but dyskinesia was still absent.In early PD, dopamine agonists are often used as first-line monotherapy, but efficacy is usually lost within a few years, at which time l-DOPA is added but with the risk of dyskinesia appearance. These results show that istradefylline is effective in improving motor function in combination with low dose dopaminergic drug treatment without provoking dyskinesia.

The adenosine A2A receptor antagonist, istradefylline enhances the anti-parkinsonian activity of low doses of dopamine agonists in MPTP-treated common marmosets

The adenosine A2A receptor antagonist, istradefylline, enhances anti-parkinsonian activity in patients with advanced Parkinson׳s disease (PD) already treated with combinations of L-DOPA and dopamine agonist drugs but who are still exhibiting prolonged ‘OFF’ periods. In contrast, the effects of istradefylline on motor function when administered in combination with low dose dopamine agonist therapy in early PD are unknown. We now investigate whether istradefylline administered with a threshold dose of either the non-ergot dopamine agonist, ropinirole or the ergot dopamine agonist, pergolide enhances anti-parkinsonian activity in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmoset.Both ropinirole (0.01–0.1mg/kg p.o.) and pergolide (0.003–0.1mg/kg p.o.) administered alone produced dose dependent increases in locomotor activity, a reduction in motor disability. Threshold doses of ropinirole (0.025–0.075mg/kg p.o.) and pergolide (0.01–0.075mg/kg p.o.) were then selected that in individual animals caused a small but non-significant anti-parkinsonian effect. Administration of istradefylline (10mg/kg p.o.) alone resulted in a decrease in motor disability and increase in ‘ON’ time but dyskinesia was not observed. Combined administration of pergolide or ropinirole with istradefylline resulted in an increase in the reversal of motor disability and increase in ‘ON’ time compared to that produced by either treatment alone but dyskinesia was still not observed.These results show that istradefylline is effective in improving motor function when combined with low dose dopamine agonist treatment. In early PD, this may avoid dose escalation or allow a reduction in dopamine agonist dosage without a loss of efficacy and prevent dopaminergic side-effects from becoming treatment limiting.

L-DOPA-induced behavioral sensitization of motor activity in the MPTP-treated common marmoset as a Parkinson's disease model

l-DOPA is the gold standard for treatment of Parkinson's disease (PD). However, the drug produces some serious side effects, including dyskinesia, which is characterized by repetitive involuntary movements—including chorea. In the present preclinical study using a nonhuman primate model, dyskinesia caused by repeated l-DOPA administration was investigated in the context of behavioral sensitization by objectively quantifying motor activity in the common marmoset of PD model (the Parkinsonian marmoset). Twelve male Parkinsonian marmosets previously treated with MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) and six intact male marmosets were used. The motor activity of each marmoset was measured using infrared sensors attached to each individual living cage. Parkinsonian marmosets (n=6) exhibited behavioral sensitization (enhanced motor activity) in 10weeks upon oral administration of l-DOPA (10mg/kg per day on 3days/week). These animals also exhibited dyskinesia characterized by repetitive rapid movements including chorea in 6–10weeks. Neither behavioral sensitization nor dyskinesia was observed in Parkinsonian marmosets given vehicle and in intact marmosets given l-DOPA at the same dose (both n=6 each). Behavioral sensitization was detected sensitively and objectively on motor activity only in Parkinsonian marmosets given repeated l-DOPA at a similar dose used in PD patients. The behavioral feature of the marmosets was dyskinesia similar to that of PD patients but appeared earlier than would be manifested in humans. In spite of statistically significant behavioral sensitization, some marmosets did not exhibit dyskinesia in the present limited l-DOPA administration period. Although both commonalities and differences may exist between behavioral sensitization and dyskinesia, behavioral sensitization is considered to be an objective, quantitative, sensitive and predictive measure of behavioral mechanism underlying dyskinesia in preclinical studies in evaluating compounds.

The H3 receptor agonist immepip does not affect l-dopa-induced abnormal involuntary movements in 6-OHDA-lesioned rats

The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have been suggested as a novel pharmacological approach. We examined the effects of the H3 agonist, immepip, in 6-OHDA-lesioned rats exhibiting AIMs (abnormal involuntary movements), a rat analogue of dyskinesia, in response to l-dopa compared to the known anti-dyskinetic agents amantadine, MK-801 and 8-OHDPAT. We then attempted to extend these studies in to dyskinetic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated common marmosets.Amantadine, MK-801 and 8-OHDPAT all dose-dependently reduced l-dopa-induced axial, lingual and oral (ALO) AIMs in 6-OHDA-lesioned animals accompanied by a reduction in contralateral rotation with higher doses of amantadine and MK-801. By contrast, immepip had no effect on AIMs expression or contralateral rotation. In the MPTP-treated common marmoset exhibiting dyskinesia to l-dopa, immepip alone induced retching and in combination with l-dopa administered subcutaneously or orally induced the rapid onset of retching and vomiting which was not controlled by pretreatment with domperidone. Administration of the unrelated H3 agonist, imetit had the same effect. Despite causing negative side-effects, it appears that both agonists reduced the antiparkinsonian response to l-dopa resulting in reduced dyskinesia.H3 agonists appear unlikely candidates for the treatment of dyskinesia in PD based on lack of evidence of efficacy and potential adverse effects.

The Adenosine A2A-Receptor Antagonist Istradefylline Enhances the Motor Response of l-DOPA Without Worsening Dyskinesia in MPTP-Treated Common Marmosets

The adenosine A2A-receptor antagonist istradefylline decreases OFF time in patients with Parkinson’s disease who are already treated with optimal doses of dopaminergic medication but can cause an increase in non-troublesome dyskinesia. Preclinical experiments have shown that A2A antagonists are most effective in potentiating motor function when combined with submaximal doses of l-DOPA. However, the effects of combining istradefylline with sub-optimal l-DOPA treatment on established dyskinesia have not been studied. We now examine the effects of acute and repeated administration of istradefylline on dyskinesia in MPTP-treated common marmosets previously primed to exhibit involuntary movements by prior exposure to l-DOPA. In these animals, single dose acute oral administration of istradefylline (10 mg/kg) enhanced and prolonged the anti-parkinsonian effects of a sub-optimal dose of l-DOPA (2.5 mg/kg). The chronic co-administration of istradefylline (10 mg/kg) with l-DOPA (2.5 mg/kg) for 21 days did not worsen the severity of existing dyskinesia. Rather, the severity of dyskinesia tended to be reduced over the 21-day treatment period. These results suggest that istradefylline can be used to potentiate the effects of sub-optimal doses of l-DOPA in the treatment of Parkinson’s disease without causing or worsening dyskinesia.

PET Analysis of Dopaminergic Neurodegeneration in Relation to Immobility in the MPTP-Treated Common Marmoset, a Model for Parkinson’s Disease

BackgroundPositron Emission Tomography (PET) measurement was applied to the brain of the common marmoset, a small primate species, treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The marmoset shows prominent Parkinson’s disease (PD) signs due to dopaminergic neural degeneration. Recently, the transgenic marmoset (TG) carrying human PD genes is developing. For phenotypic evaluations of TG, non-invasive PET measurement is considered to be substantially significant. As a reference control for TG, the brain of the MPTP-marmoset as an established and valid model was scanned by PET. Behavioral analysis was also performed by recording locomotion of the MPTP-marmoset, as an objective measure of PD signs.Methodology/Principal FindingsMarmosets received several MPTP regimens (single MPTP regimen: 2 mg/kg, s.c., per day for 3 consecutive days) were used for PET measurement and behavioral observation. To measure immobility as a central PD sign, locomotion of marmosets in their individual living cages were recorded daily by infrared sensors. Daily locomotion counts decreased drastically after MPTP regimens and remained diminished for several months or more. PET scan of the brain, using [11C]PE2I as a ligand of the dopamine (DA) transporter, was performed once several months after the last MPTP regimen. The mean binding potential (BPND) in the striatum (putamen and caudate) of the MPTP-marmoset group was significantly lower than that of the MPTP-free control group (n = 5 for each group). In the MPTP-marmosets, the decrease of BPND in the striatum closely correlated with the decrease in locomotion counts (r = 0.98 in putamen and 0.91 in caudate).Conclusion/SignificanceThe present characterization of neural degeneration using non-invasive PET imaging and of behavioral manifestation in the MPTP marmoset mimics typical PD characteristics and can be useful in evaluating the phenotype of TG marmosets being developed.

Pardoprunox reverses motor deficits but induces only mild dyskinesia in MPTP‐treated common marmosets

Long-acting full dopamine D(2) agonists produce less dyskinesia in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated primates and in Parkinson's disease than effective antiparkinsonian doses of levodopa. They do not however, prevent priming for dyskinesia expression on subsequent levodopa exposure. In contrast, the effects of partial D(2) receptor agonists on dyskinesia are unclear. We now examine the ability of the partial D(2) agonist pardoprunox (SLV308) to improve motor function and its propensity to prime for dyskinesia in drug naïve, MPTP-treated common marmosets. Previously, drug naïve, MPTP-treated common marmosets were treated with equivalent doses of either pardoprunox (SLV308) (0.1 mg/kg po), ropinirole (0.18 mg/kg po), or levodopa (10 mg/kg po BID) for 28 days. All treatments induced a similar reduction of motor disability. Dyskinesia induced by levodopa was of greater intensity than that following administration of either pardoprunox (SLV308) or ropinirole. Administration of pardoprunox (SLV308) resulted in dyskinesia that was less intense and of shorter duration than either ropinirole or levodopa. At the end of drug treatment, acute challenge with levodopa resulted in the expression of marked dyskinesia in animals that had previously received chronic levodopa or ropinirole treatment. However, animals previously treated with pardoprunox (SLV308) showed only mild dyskinesia in response to the levodopa challenge. These results suggest that the partial D(2) agonist pardoprunox (SLV308) is less likely to prime for dyskinesia or to lead to the expression of dyskinesia than either levodopa or full dopamine agonists.

Synthetic studies on selective adenosine A2A receptor antagonists. Part II: Synthesis and structure–activity relationships of novel benzofuran derivatives

Based on the previously reported lead compound, a series of benzofuran derivatives were prepared to study their antagonistic activities to A2A receptor. The replacement of the phenyl group at the 4-position with a heterocyclic ring improved the PK profile and aqueous solubility. From these studies, we discovered a potent new A2A antagonist, 12a, which has both a good oral bioavailability and in vivo efficacy on motor disability in MPTP-treated common marmosets.

The timing of administration, dose dependence and efficacy of dopa decarboxylase inhibitors on the reversal of motor disability produced by L-DOPA in the MPTP-treated common marmoset

Dopa decarboxylase inhibitors are routinely used to potentiate the effects of L-DOPA in the treatment of Parkinson's disease. However, neither in clinical use nor in experimental models of Parkinson's disease have the timing and dose of dopa decarboxylase inhibitors been thoroughly explored. We now report on the choice of dopa decarboxylase inhibitors, dose and the time of dosing relationships of carbidopa, benserazide and L-α-methyl dopa (L-AMD) in potentiating the effects of L-DOPA in the 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP)-treated common marmoset. Pre-treatment with benserazide for up to 3 h did not alter the motor response to L-DOPA compared to simultaneous administration with L-DOPA. There was some evidence of a relationship between carbidopa and benserazide dose and increased locomotor activity and the reversal of motor disability. But in general, commonly used dose levels of dopa decarboxylase inhibitors appeared to produce a maximal motor response to L-DOPA. In contrast, dyskinesia intensity and duration continued to increase with both carbidopa and benserazide dose. The novel dopa decarboxylase inhibitor, L-AMD, increased locomotor activity and improved motor disability to the same extent as carbidopa or benserazide but importantly this was accompanied by significantly less dyskinesia. This study shows that currently, dopa decarboxylase inhibitors may be routinely employed in the MPTP-treated primate at doses which are higher than those necessary to produce a maximal potentiation of the anti-parkinsonian effect of L-DOPA. This may lead to excessive expression of dyskinesia in this model of Parkinson's disease and attention should be given to the dose regimens currently employed.

Pramipexole combined with levodopa improves motor function but reduces dyskinesia in MPTP‐treated common marmosets

Reduced expression of dyskinesia is observed in levodopa-primed MPTP-treated common marmosets when dopamine agonists are used to replace levodopa. We now investigate whether a combination of the D-2/D-3 agonist pramipexole and levodopa also reduces dyskinesia intensity while maintaining the reversal of motor disability. Drug naïve, non-dyskinetic MPTP-treated common marmosets were treated daily for up to 62 days with levodopa (12.5 mg/kg plus carbidopa 12.5 mg/kg p.o. BID) or pramipexole (0.04-0.3 mg/kg BID) producing equivalent reversal of motor disability and increases in locomotor activity. Levodopa alone resulted in marked dyskinesia induction but little or no dyskinesia resulted from the administration of pramipexole. From day 36, some animals were treated with a combination of levodopa (3.125-6.25 mg/kg plus carbidopa 12.5 mg/kg p.o. BID) and pramipexole (0.1-0.2 mg/kg p.o. SID). This improved motor disability to a greater extent than occurred with levodopa alone. Importantly, while dyskinesia was greater than that produced by pramipexole alone, the combination resulted in less intense dyskinesia than produced by levodopa alone. These results suggest that pramipexole could be administered with a reduced dose of levodopa to minimize dyskinesia in Parkinson's disease while maintaining therapeutic efficacy.

Zonisamide Attenuates MPTP Neurotoxicity in Marmosets

MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) induces parkinsonism in humans and animals. The effects of zonisamide on dopamine neurons were studied in MPTP-treated common marmosets (Callithrix jacchus). Groups of animals (n = 3) were treated with MPTP (2.5 mg/kg, every 24 h x 3); MPTP plus zonisamide (40 mg/kg administered 1 h before each MPTP dose); MPTP plus selegiline (a known MAO-B inhibitor) (2 mg/kg administered 1 h before each MPTP dose); and saline controls. An immunohistochemical study of the substantia nigra was performed 14 days after MPTP treatment in each group. MPTP reduced the mean number of tyrosine hydroxylase (TH)-positive neurons to 10% of the normal control group and mean cell size was significantly (P < 0.001) reduced from 424 to 159 µm². In the group pre-treated with zonisamide, the mean number of TH-positive neurons was reduced to 26% of that in the normal control group and the mean neuron size was significantly (P < 0.05) increased from 159 to 273 µm² compared with the group treated with MPTP alone. Moreover, in the group pre-treated with selegiline, the mean number of TH-positive neurons was 47% of that in the normal control group and the mean neuron size was increased significantly (P < 0.01) from 159 to 319 µm² compared to the group treated with MPTP alone. This observation suggests that zonisamide reduces MPTP toxicity.

Continuous rotigotine administration reduces dyskinesia resulting from pulsatile treatment with rotigotine or l-DOPA in MPTP-treated common marmosets

We previously showed that continuous infusion of rotigotine resulted in less dyskinesia than repeated pulsatile rotigotine administration or repeated oral administration of l-DOPA in MPTP-treated marmosets. Now we investigate whether continuous rotigotine delivery modifies established dyskinesia induced by prior exposure to repeated pulsatile administration of l-DOPA or rotigotine in MPTP-treated common marmosets. Repeated oral administration of l-DOPA or subcutaneous bolus administration of rotigotine over 28 days improved motor deficits but resulted in the onset of dyskinesia of moderate intensity. When these animals were switched to a 28-day continuous infusion of rotigotine, the reversal of motor disability was maintained. In those animals initially treated with l-DOPA, there was a small reduction in dyskinesia intensity but a significant reduction in the duration of dyskinesia. However, in animals initially treated with repeated bolus administration of rotigotine, dyskinesia intensity was significantly reduced. Initial treatment with a continuous infusion of rotigotine for 28 days reversed motor disability and resulted in a low incidence of dyskinesia. On switching to repeated oral administration of l-DOPA, the improvement in motor disability was maintained but the propensity of l-DOPA to provoke dyskinesia was not affected. In addition, while the continuous delivery of rotigotine prevented the expression of dyskinesia, the previously demonstrated ability of dopamine agonists to prime for dyskinesia could not be avoided. These data suggest that dyskinesia induced by pulsatile drug treatment may be improved by switching to continuous rotigotine delivery. In addition, while continuous delivery of rotigotine may prime for dyskinesia, it does not lead to its expression.

Expression of catechol-O-methyltransferase in the brain and periphery of normal and MPTP-treated common marmosets

Catechol-O-methyltransferase (COMT) inhibition is widely used to potentiate the effects of levodopa in Parkinson's disease but the effects of nigral dopaminergic cell loss and levodopa treatment on COMT activity are not known. The present study investigated the expression of COMT in the brain and liver of normal common marmosets, and animals treated with MPTP and those treated with levodopa to induce dyskinesia. Reverse transcript PCR demonstrated the expression of COMT mRNA in the liver, cortex and striatum of normal marmosets. Using Western blotting, the presence of two subunits of COMT protein, membrane bound COMT (MB-COMT) and soluble COMT (S-COMT), was shown in the liver, cortex and striatum of normal marmosets. Quantitative analysis of the MB-COMT and S-COMT subunit bands showed that there was no significant difference in the density of bands in MPTP treated marmosets or those exposed to levodopa compared to normal animals. COMT immunoreactivity was expressed in many brain regions including the cortex and striatum. No difference in COMT staining intensity was observed between normal, MPTP exposed or MPTP plus levodopa treated animals. COMT immunostaining was present in most striatal neurones and it was occasionally seen in glial cells. The data from present study demonstrated the expression of COMT mRNA and protein in the brain of common marmoset contrary to a previous report that it is not expressed in this species. COMT activity appears unaffected by loss of the dopaminergic nigro-striatal pathway and levodopa treatment.

Continuous administration of rotigotine to MPTP-treated common marmosets enhances anti-parkinsonian activity and reduces dyskinesia induction

Rotigotine is a novel, non-ergoline dopamine D 3/D 2/D 1-receptor agonist for the treatment of Parkinson's disease that can be continuously delivered by the transdermal route to provide stable plasma levels. Continuous drug delivery should reduce the risk of dyskinesia induction in comparison to pulsatile dopaminergic treatment. Thus the aim of the study was to compare the reversal of motor disability and induction of dyskinesia produced by continuous compared to pulsatile rotigotine administration in MPTP-treated common marmosets. The study also investigated whether pulsatile or continuous rotigotine administration in combination with l-DOPA prevented l-DOPA-induced dyskinesia. Animals were treated for 28 days with vehicle or pulsatile (twice daily) or continuous delivery of rotigotine (via an osmotic minipump). Subsequently, l-DOPA was then co-administered for a further 28 days. Animals were assessed for locomotor activity, motor disability and dyskinesia induction. The study showed that both continuous and pulsatile administration of rotigotine improved motor deficits and normalized motor function in MPTP-treated monkeys. However, continuous rotigotine delivery reduced dyskinesia expression compared to pulsatile treatment. Both pulsatile and continuous rotigotine administration produced less dyskinesia than administration of l-DOPA alone. The addition of l-DOPA to either pulsatile or continuous rotigotine treatment resulted in the induction of marked dyskinesia similar to that produced by treatment with l-DOPA alone. These data further support the hypothesis that continuous delivery of a dopaminergic agent reduces the risk of dyskinesia induction. However, continuous rotigotine administration did not prevent l-DOPA from inducing dyskinesia suggesting that l-DOPA may induce dyskinesia by mechanisms different from dopamine agonist drugs.

Striatal Alterations of Secretogranin-1, Somatostatin, Prodynorphin, and Cholecystokinin Peptides in an Experimental Mouse Model of Parkinson Disease

The principal causative pathology of Parkinson disease is the progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta projecting to the striatum in the brain. The information regarding the expression of neuropeptides in parkinsonism is very limited. Here we have elucidated striatal neuropeptide mechanisms in experimental parkinsonism using the unilateral 6-hydroxydopamine model to degenerate dopamine neurons. A thoroughly controlled sample preparation technique together with a peptidomics approach and targeted neuropeptide sequence collections enabled sensitive detection, identification, and relative quantitation of a great number of endogenous neuropeptides. Previously not recognized alterations in neuropeptide levels were identified in the unilateral lesioned mice with or without subchronic 3,4-dihydroxy-L-phenylalanine administration, the conventional treatment of Parkinson disease. Several of these peptides originated from the same precursor such as secretogranin-1, somatostatin, prodynorphin, and cholecystokinin. Disease-related biotransformation of precursors into individual peptides was observed in the experimental model of Parkinson disease. Several previously unreported potentially biologically active peptides were also identified from the striatal samples. This study provides further evidence that neuropeptides take part in mediating the central nervous system failure associated with Parkinson disease.

Continuous delivery of ropinirole reverses motor deficits without dyskinesia induction in MPTP-treated common marmosets

L-DOPA treatment of Parkinson's disease induces a high incidence of motor complications, notably dyskinesia. Longer acting dopamine agonists, e.g. ropinirole, are thought to produce more continuous dopaminergic stimulation and less severe dyskinesia. However, standard oral administration of dopamine agonists does not result in constant plasma drug levels, therefore, more continuous drug delivery may result in both prolonged reversal of motor deficits and reduced levels of dyskinesia. Therefore, we compared the effects of repeated oral administration of ropinirole to constant subcutaneous infusion in MPTP-treated common marmosets. Animals received oral administration (0.4 mg/kg, BID) or continuous infusion of ropinirole (0.8 mg/kg/day) via osmotic minipumps for 14 days (Phase I). The treatments were then switched and continued for a further 14 days (Phase II). In Phase I, locomotor activity was similar between treatment groups but reversal of motor disability was more pronounced in animals receiving continuous infusion. Dyskinesia intensity was low in both groups however there was a trend suggestive of less marked dyskinesia in those animals receiving continuous infusion. In Phase II, increased locomotor activity was maintained but animals switched from oral to continuous treatment showing an initial period of enhanced locomotor activity. The reversal of motor disability was maintained in both groups, however, motor disability tended towards greater improvement following continuous infusion. Importantly, dyskinesia remained low in both groups suggesting that constant delivery of ropinirole neither leads to priming nor expression of dyskinesia. These results suggest that a once-daily controlled-release formulation may provide improvements over existing benefits with standard oral ropinirole in Parkinson's disease patients.