New therapeutic agents developed for treating neurological disorders are often tested successfully on rodents, and yet 80–90% of these prove ineffective when transitioning to clinical application. Testing in an appropriate large-animal model should improve translational success and is frequently expected by regulatory bodies. In this project, we aimed to establish a novel sheep model of Parkinson’s disease as a large-brained experimental model for translational research. Our objective was to create a sheep model of Parkinson’s disease by unilaterally infusing the neurotoxin 6-hydroxydopamine into the substantia nigra pars compacta. This approach, previously used to induce parkinsonism in rat and non-human primate models, causes dopaminergic imbalance and induces rotational behaviour in quadrupeds challenged with dopaminergic receptor agonists. In the present sheep study, the mixed dopamine receptor agonist apomorphine, 0.25 mg/kg, and dopamine D2 agonist ropinirole, 0.16 mg/kg, were used to induce rotational behaviour and confirm dopamine depletion. Behavioural signs were then measured and characterised in the field using automated movement tracking with simultaneous video recordings. Post-mortem, the extent of the 6-hydroxydopamine lesions were evaluated through tyrosine hydroxylase immunohistochemistry and quantifying levels of catecholamines (dopamine, 3,4-dihydroxyphenylacetic acid, and homovanilic acid) quantified using high-performance liquid chromatography. Our new sheep model of Parkinson’s disease using 6-hydroxydopamine is safe and offers a number of regulatory, ethical, and financial advantages over non-human primate 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine models. It provides a platform to evaluate novel anti-parkinsonian agents and medical devices with the promise of greater success for translation into clinical application than has been achieved using small animal models.