LUHMES-derived 3D organoids as an enhanced platform for modelling dopaminergic neurodegeneration in Parkinson's...

Parkinson's disease (PD) research traditionally relies on animal models and two-dimensional (2D) culture models. These models fail to recapitulate the complex cellular architecture and network interactions that are possible with three-dimensional (3D) organoid models. While 3D models allow improved cell-cell interactions, spatial organization, and metabolic microenvironments, however, their utility for modelling dopaminergic neurodegeneration remains underexplored.

We developed and characterized 3D organoids from LUHMES cells which are human embryonic neuronal precursor cells and a well-established human dopaminergic neuronal cell line. We directly compare their responses to the mitochondrial toxin MPP+ against conventional 2D cultures. Functional readouts included cell viability, ATP production, dopaminergic marker expression (tyrosine hydroxylase, MAP2, β-III tubulin), reactive oxygen species (ROS) generation, electrophysiological activity via multi-electrode arrays (MEA), and calcium signalling dynamics.

3D LUHMES organoids had 9-fold higher synapsin expression compared to 2D cultures, indicating enhanced synaptic maturity and network complexity. Following acute MPP+ exposure (0.25 mM, 24 h), 3D organoids were significantly more sensitive than 2D cultures, with greater reductions in cell viability (35% vs. 31%), ATP production (36% vs. 31%), and dopaminergic marker expression (TH: 60% reduction in both; MAP2: 70% vs. 54%; TUJ1: 62% vs. 20%). ROS production increased uniformly in 3D organoids (85% positive cells) compared to heterogeneous accumulation in 2D cultures (78% positive cells). Functional assessments revealed that 3D organoids displayed higher baseline electrophysiological activity that was sensitive to impairment following MPP+ treatment, spike amplitude, and calcium signalling responses to various stimuli (ATP, glutamate, GABA).

LUHMES-derived 3D organoids demonstrate greater physiological relevance for modelling PD-related dopaminergic neurodegeneration than 2D cultures. The enhanced sensitivity to mitochondrial toxins, combined with more sophisticated network architecture and functional properties, suggests this model is a valuable platform for mechanistic studies of neurodegeneration and preclinical drug screening. These findings also support the broader adoption of 3D culture systems in neurodegenerative disease research.