Real-Time Neurotransmitter Sensing in Midbrain Organoids Using 3D Carbon Cup Electrodes

Real-time chemically selective neurotransmitter detection in brain organoids poses difficulties due to their intricate three-dimensional structure and limited optical access. Researchers have developed 3D printed carbon cup electrodes (CCEs), which are pyrolyzed multi-level pillar structures. These CCEs are intended to enclose a single organoid, increase electroactive surface area, and facilitate amperometric recordings of oxidizable neurotransmitters.

The CCEs demonstrate excellent electrochemical performance and reproducibility, allowing diffusion-controlled dopamine redox behavior without interference from the culture medium.

By utilizing ventral midbrain (vMB) organoids at two maturation stages (Day 30 and Day 90), the researchers were able to show real-time detection of potassium-evoked dopamine release directly from the organoids. The mature organoids displayed higher peak currents, which correlated with increased tyrosine hydroxylase (TH) expression, as confirmed by immunocytochemistry.

Embedding the organoids in Matrigel significantly dampened dopamine signals, indicating a diffusion barrier that restricts stimulant access and/or dopamine transport to the electrode.

Repetitive stimulations revealed a progressive depletion of dopamine stores, consistent with reduced vesicular release. Chemical perturbations using 6-hydroxydopamine and verapamil further validated the platform by producing significant reductions in dopamine release accompanied by a loss of TH neurons. These findings establish the CCEs as a powerful tool for label-free, real-time monitoring of neurotransmission in human organoids.

This platform has the potential for multiplexed sensing and integration of enzymatic detection strategies for non-electroactive neurotransmitters.