Poster BB4 - Investigation of microfluidic generated neuron-collagen spheroids for use as building blocks

Introduction: Better understanding of the development, connectivity, and network dynamics of neurons is critical for finding therapeutic treatments for neurodegenerative diseases. In vitro neuronal models have been widely used for this purpose. Creation of 3D in vitro neuronal structures has the potential to model cell-cell and cell-matrix interactions found in the intact brain. However, traditional bioprinting strategies for creating neural tissue do not produce robust and dense networks of neurons. To overcome this limitation, we propose using matrix-infused neural spheroids as a building block and potentially bioink for bioprinting brain tissue on a dish in the future.

Materials and

Methods: In this study, we used a microfluidic platform to generate encapsulated droplets of cortical neurons in a collagen matrix. The cellular model used in this study consisted of dissociated primary rat cortical neurons. After exploring the interaction of neurons with different types of nature-driven hydrogels, we used collagen type I (5 mg/ml) to make a mixture with dissociated neurons. The mixture was then used as the aqueous phase fed to the microfluidic droplet generator. Neural spheroids expressed genetically encoded calcium indicator jRGECO1a for optical recording of neuronal activity. Cell staining assays were employed for live/dead staining and visualization of neuronal and astrocyte structures in the spheroids.

Results, Conclusions, and Discussions: We successfully generated densely packed spheroids of neurons in collagen. After releasing the encapsulated spheroids, we monitored them for few weeks to track neuronal aggregation in the gel. These spheroids exhibited normal neuronal maturation and functional behavior, demonstrating the potential of this method for a matrix-infused neuronal model.
Microfluidic droplet generation offers a high-throughput and efficient method of forming neural spheroids to better study cell-cell and cell-extracellular matrix (ECM) interactions in an in vivo like environment. This model could be utilized for fundamental studies of neuronal structures in neurodegenerative diseases and traumatic brain injury (TBI). Furthermore, we are investigating the feasibility of using these spheroids as bioink to form a cortical tissue and assess its functionality.

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