Objective: Parkinson's disease (PD) is one of the most prevalent neurodegenerative diseases, characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. The treatment of PD aims to alleviate motor symptoms by replacing the reduced endogenous dopamine. Currently, there are no disease-modifying agents for the treatment of PD. Zebrafish (Danio rerio) have emerged as an effective tool for new drug discovery and screening in the age of translational research. The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to cause a similar loss of dopaminergic neurons in the human midbrain, with corresponding Parkinsonian symptoms. L-type calcium channels (LTCCs) have been implicated in the generation of mitochondrial oxidative stress, which underlies the pathogenesis of Parkinson's disease. Therefore, we investigated the neuro-restorative effect of LTCC inhibition in an MPTP-induced zebrafish PD model and suggested a possible drug candidate that might modify the progression of PD.
Methods: All experiments were conducted using a line of transgenic zebrafish, Tg (dat:EGFP), in which green fluorescent protein (GFP) is expressed in dopaminergic neurons. The experimental groups were exposed to 500µ㏖ MPTP from 1 to 3 days post fertilization (dpf). The drug candidates: Levodopa 1m㏖, Nifedipine 10µ㏖, Nimodipine 3.5 µ㏖, Diethylstilbestrol 0.3 µ㏖, Luteolin 100 µ㏖, Cacitriol 0.25 µ㏖ were exposed from 3 to 5 dpf. Locomotor activity was assessed by automated tracking and dopaminergic neurons were visualized in vivo by confocal microscopy.
Results: Levodopa, Nimodipine, Diethylstilbestrol, and Calcitriol had significant positive effects on the restoration of motor behavior, which was damaged by MPTP. Nimodipine and Calcitriol have significant positive effects on the restoration of dopaminergic neurons, which were reduced by MPTP. Through locomotor analysis and dopaminergic neuron quantification, we identified the neuro-restorative effects of Nimodipine and Calcitriol in Zebrafish MPTP-induced PD model.
Conclusion: The present study identified the neuro-restorative effects of nimodipine and calcitriol in an MPTP-induced zebrafish model of Parkinson's disease. They restored dopaminergic neurons which were damaged due to the effects of MPTP and normalized the locomotor activity. LTCCs have potential pathological roles in neurodevelopmental and neurodegenerative disorders. Zebrafish are highly amenable to high-throughput drug screening and might, therefore, be a useful tool to work towards the identification of disease-modifying treatment for PD. Further studies including zebrafish genetic models to elucidate the mechanism of action of the disease-modifying candidate by investigating Ca2+ influx and mitochondrial function in dopaminergic neurons, are needed to reveal the pathogenesis of PD and develop disease-modifying treatments for PD.
Keywords: Dopaminergic neuron; Drug screening; L-type Ca2+ channel; MPTP; Parkinson’s disease; Zebrafish.