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Design, Surface Treatment, Cellular Plating, and Culturing of Modular Neuronal Networks Composed of Functionally Inter-connected Circuits
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Nanoscale Patterning of In Vitro Neuronal Circuits
José C Mateus1, Sean Weaver2, Dirk van Swaay3
1Neuroengineering and Computational Neuroscience Laboratory, i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, 4200-135 Porto, Portugal.
ACS Nano
|April 11, 2022
Summary
This study introduces a novel in vitro method for precise neuronal circuit construction. It enables the study of neuronal connectivity at both circuit and synapse levels, aiding biophysical model validation.
Area of Science:
- Neuroscience
- Biophysics
- Bioengineering
Background:
- Current in vitro methods for patterning neurons offer limited control over multiscale neuronal connectivity.
- Studying neuronal circuits in vivo at synaptic resolution presents significant challenges.
- There is a need for in vitro alternatives to validate biophysical and computational models of neuronal networks.
Purpose of the Study:
- To develop an in vitro technique for multiscale control of neuronal connectivity, from circuits to synapses.
- To create an in vitro system for validating biophysical and computational models of neuronal circuits.
- To enable the study of neuronal circuits with synaptic resolution in a controlled environment.
Main Methods:
- Utilized electron beam lithography and photolithography to fabricate polydimethylsiloxane (PDMS) structures with feature sizes from 150 nm to millimeters.
- Engineered nanochannels to restrict axon growth while permitting dendritic spine passage, guiding synapse formation between neuronal nodes.
- Employed genetically encoded calcium indicators and fluorescently tagged postsynaptic protein PSD-95 to confirm functional synapse formation.
Main Results:
- Successfully generated large numbers of isolated feed-forward neuronal circuits in vitro.
- Demonstrated that neuronal connections were precisely restricted to regions defined by nanochannels.
- Confirmed the formation of functional synapses within the engineered nanochannel-connected regions.
Conclusions:
- The developed lithography-based technique enables precise, multiscale control over neuronal connectivity in vitro.
- This method provides a valuable platform for validating biophysical and computational models of neural circuits.
- The approach facilitates the study of synaptic formation and function in a highly controlled experimental setting.
