In vitro neuronal culture is an important research platform in cellular and network neuroscience. However, neurons cultured on a homogeneous scaffold form dense, randomly connected networks and display excessively synchronized activity; this phenomenon has limited their applications in network-level studies, such as studies of neuronal ensembles, or coordinated activity by a group of neurons. Herein, we develop polydimethylsiloxane-based microfluidic devices to create small neuronal networks exhibiting a hierarchically modular structure resembling the connectivity observed in the mammalian cortex. The strength of intermodular coupling was manipulated by varying the width and height of the microchannels that connect the modules. Using fluorescent calcium imaging, we observe that the spontaneous activity in networks with smaller microchannels (2.2−5.5 μm2) had lower synchrony and exhibit a threefold variety of neuronal ensembles. Optogenetic stimulation demonstrates that a reduction in intermodular coupling enriches evoked neuronal activity patterns and that repeated stimulation induces plasticity in neuronal ensembles in these networks. These findings suggest that cell engineering technologies based on microfluidic devices enable in vitro reconstruction of the intricate dynamics of neuronal ensembles, thus providing a robust platform for studying neuronal ensembles in a well-defined physicochemical environment.
@article{arxiv.2405.19159,
title = {Precision microfluidic control of neuronal ensembles in cultured cortical networks},
author = {Hakuba Murota and Hideaki Yamamoto and Nobuaki Monma and Shigeo Sato and Ayumi Hirano-Iwata},
journal= {arXiv preprint arXiv:2405.19159},
year = {2024}
}