Primate MPTP model

The 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication protocol, chronic L-Dopa treatment, and the clinical assessments were conducted in four male macaques (Macaca fascicularis, Xierxin, Beijing, PR of China). The macaques were first rendered parkinsonian with MPTP-hydrochloride (0.2 mg/kg, i.v., Sigma) dissolved in saline. Daily (at 9 a.m.) assessment of parkinsonism in home cages for 30 min by two blinded observers was done using a validated rating scale assessing tremor, general level of activity, body posture (flexion of spine), vocalization, freezing and frequency of arm movements (for each upper limb), and rigidity. Once parkinsonism was stable, levodopa (Madopar, Roche, Levodopa/carbidopa, ratio 4:1) was administered twice daily for 4–5 months at an individually tailored dose (18–22 mg/kg) designed to produce a full reversal of the parkinsonian condition (p.o. by gavage). Over this period, animals developed severe and reproducible dyskinesia, presenting choreic–athetoid (characterized by constant writhing and jerking motions), dystonic, and sometimes ballistic movements (large-amplitude flinging, flailing movements) as seen in long-term L-Dopa-treated Parkinson’s disease patients. SAG (3, 9, 27 mg/kg) was dissolved in 10% DMSO, 45% HPCD in 0.9% saline and administered i.v. Within-subject escalation was performed with a washout period of 3 days between escalating doses.

Immediately after drug administration, monkeys were transferred to an observation cage (dimensions—1.1 m × 1.5 m × 1.1 m) as per guidelines⁸⁴. The total duration of observation was 240 min of drug exposure. We performed a battery of behavioral observations as previously described⁸⁵. Experts blinded to the treatment observed 10-min video recordings taken every 30 min throughout the duration of the experiment and scored the severity of the parkinsonian condition using the parkinsonian disability score. The parkinsonian disability score is a combination of four different scores: (1) the range of movement score, (2) bradykinesia score, (3) posture score, and (4) tremor score. These four scores are combined using formula: (4 − range of movement) + bradykinesia + postural abnormality + tremor. We rated the severity of dyskinesia using the Dyskinesia Disability Scale⁸⁴: 0, dyskinesia absent; 1, mild, fleeting, and rare dyskinetic postures and movements; 2, moderate, more prominent abnormal movements, but not interfering with normal behavior; 3, marked, frequent and, at times, continuous dyskinesia intruding on the normal repertoire of activity; or, 4, severe, virtually continuous dyskinetic activity replacing normal behavior and disabling to the animal.

Primate behavior was rated using most recent guidelines⁸⁴ which include the recommendation to use nonparametric tests (nature of the rating scale and number of individuals). Nonparametric analysis is less sensitive compared to parametric analysis and reduces the risk of identifying false-positive responses. We presented the time course of parkinsonian disability and dyskinesia scores in 30 min time bins over the 4 h observation period. We also presented the median of the total scores of disability, dyskinesia, chorea, and dystonia at 0–2 h following treatment. We statistically compared the parkinsonian and dyskinesia scores between different conditions using a Friedman’s test followed by Dunn’s multiple comparison test.