Long-Term Developmental Process of the Human Cortex Revealed In Vitro by Axon-Targeted Recording Using a...

Area of Science:

• Neuroscience
• Stem Cell Biology
• Bioengineering

Background:

• Studying human neuronal development requires reliable methods to monitor neural network activity over time.
• Existing methods using human induced pluripotent stem cells (hiPSCs) often rely on feeder astrocytes, which can influence neuronal activity.
• Long-term evaluation of synchronized neuronal activity in a more natural cellular environment is crucial for understanding development and disease.

Purpose of the Study:

• To develop and validate a microelectrode array (MEA) method for assessing the developmental changes in synchronized activity of hiPSC-derived neurons.
• To enable long-term monitoring of neuronal network maturation without the confounding influence of extrinsic signals from feeder astrocytes.

Main Methods:

• Fabrication of microelectrode arrays (MEAs) and microtunnels using photolithography and soft lithography.
• Induction of differentiation of hiPSCs into cortical neurons and seeding onto conventional and microtunnel MEAs.
• Recording and analysis of spontaneous spiking and bursting activities over extended periods.

Main Results:

• Conventional MEAs showed limited synchronized activity beyond 300 days due to cell detachment.
• Microtunnel MEAs successfully maintained neuronal networks, enabling synchronized activity recording up to 450 days post-induction.
• The microtunnel MEA demonstrated a higher proportion of active electrodes and a steady-state activity pattern around 330 days, indicating network maturation.

Conclusions:

• Microtunnel MEAs provide a robust platform for monitoring the long-term development of human neuronal networks in a more natural state.
• This method facilitates the study of neuronal network maturation and provides a valuable tool for investigating human cortical development and disease mechanisms.