THREE-DIMENSIONAL LIQUID METAL-BASED CONFORMAL NEURO-INTERFACES FOR HUMAN HIPPOCAMPAL ORGANOIDS

The hippocampus is a critical component of the human brain that plays a significant role in memory, learning, and spatial navigation. Due to the limited availability of human brain tissues, studies on the human hippocampus are restricted. The emergence of neural organoids has largely addressed this limitation, with some region-specific organoids exhibiting corresponding cell types and functions. However, the utilization of human hippocampal organoids (hHOs) to study hippocampal functions in vitro presents two significant challenges.  Firstly, the in vitro construction of hHOs remains in the preliminary stage. The hippocampus has a highly specific layered structure and multiple unidirectional neural projection pathways. This special structure still requires a huge boost for hHOs. Secondly, the neural activity of hHOs poses a challenge, as electrophysiological activity is critical to information transmission in neural tissue. However, there is a lack of electronics that apply to organoids. Aiming to these two challenges, this thesis reports a complex system coupling a liquid-metal polymer conductor (MPC)-based conformal neuro-interface with the hHO.

This thesis first refined the protocol derived from the dorsomedial telencephalon (DMT) organoid induction strategy to generate hHOs. This generation was accomplished by demonstrating that Wnt3a and SHH depressed the choroid plexus (ChP) growth and promoted hippocampal fate in vitro. Two types of hippocampal progenitors, HOPX⁺ and PAX6⁺ progenitors, and dentate gyrus (DG) PROX1⁺ granule neurons appeared in the hHO. The comparison between the hHO and a published dataset of the developing human hippocampus revealed a high correlation.

Then, this thesis presents a fabricating method of mesh MPC (mMPC), integrating 64 electrodes within an area of ~2*2 mm. This fabrication produced a sandwich structure with two polymer layers enclosing conductive MPC. Notably, the insulating layer accurately exposed only 15 µm in diameter electrodes. These 64 electrodes were evenly distributed on multi-points of the mesh structure in four directions. Furthermore, two layers of the mMPC were assembled in one device, allowing for the recording of the neural activities of hHOs placed between these two layers. The mesh shape allowed for the cultivation of hHOs without significant damage due to its softness and stretchability. The mMPC could be stretched up to 400% without any leakage of liquid metal and any slit in the mesh while maintaining conductivity of 4000 S/cm. The impedance and root mean square (RMS) noise were relatively low, making the mMPC an ideal neuro-interface.

Finally, the hHO was integrated into the mMPC, and its spontaneous spikes were successfully recorded. Neurons in the tissue performed more active neural firing than monolayer neurons, highlighting the importance of 3D devices for neural organoids. After building a complete system with an electrical stimulus module, temperature control module, and gas control module, the mMPC will be applied to investigate neural circuits in the future.

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