ABSTRACT OF THE DISSERTATION In vitro hippocampal network axonal transmission during patterned theta burst stimulation and a platform for recording in 3D

By: Ruiyi Chen Doctor of Philosophy in Chemical and Biochemical Engineering University of California, Irvine, 2022 Professor William Tang, Chair

Neuronal network dynamics by in vitro electrophysiology serves as an objective and convenient method to explore neurophysiological mechanisms and communication. Theta burst stimulation (TBS) recapitulates natural brain rhythms which is efficacious for synaptic potentiation in hippocampal circuits. TBS is typically applied to a bundle of axons to measure the immediate response in a single downstream subregion like the CA1. Yet little is known about axonal transmission between subregions as the response to TBS propagates to other subregions from upstream stimulation in the entorhinal cortex. We reverse engineered the hippocampal network on micro-electrode arrays (MEAs) contained in a four-chambered silicon rubber device with interconnecting microfluidic. Tunnels. The micro tunnels allow monitoring single axon transmission which is hard to realize in vivo or in slices. The patterned TBS was delivered to the entorhinal cortex (EC), the gateway to cortical entry into the hippocampus. Temporal Jaccard distance was used to quantify the spike pattern similarity. We recorded the network responses to stimulation patterns that produced unique response patterns on axons at three timescales. The response to short-term stimulus repeats at 0.2 s has high variability. The 10 s repeats show some retention of similar responses, especially in the axons from CA3 to CA1, suggestive of pattern completion. While in EC to DG, the repeats evoked unique responses with similarity significantly lower than random, implying pattern separation. Between-tunnel similarity in CA3-CA1 was higher for single site stimulation than from multisite stimulation. Using the temporal pattern metric, axons carried similar responses when an incomplete stimulating pattern was presented. Our design and interrogation approach offers understanding of dynamic pattern variations at the subregional level in response to TBS. To extend this 2D network model, a 3D model was designed to bridge the gap between multi-subregion electrophysiology in vitro and in vivo recordings with high complexity from the dense packing of neurons at single regions in vivo. Here, we report fabrication of a transparent 60-electrode array that forms a second layer of recording sites above a commercial MEA. This 3D culture chamber enables recording from a 400 m thick reconstruction of four hippocampal subregions. The electrodes are easily fabricated and assembled. Optical and electrical characterizations of the electrodes have been performed. Good biocompatibility enabled successful electrophysiological recording. Two hydrogel substrates and solid glass beads for the integrated 3D culture scaffold were evaluated. Only the micro glass beads supported the 3D neuronal network formation on the platform over 21 days. Building this 3D signaling system demonstrates feasibility to access the information coded in the 3D hippocampal network in vitro for comparison to the 2D network spike and burst dynamics in response to stimulation.