A multimodal 3D neuro-microphysiological system with neurite-trapping microelectrodes

Abstract

Three-dimensional cell technologies as pre-clinical models are emerging tools for mimicking the structural and functional complexity of the nervous system. The accurate exploration of phenotypes in engineered 3D neuronal cultures, however, demands morphological, molecular and especially functional measurements. Particularly crucial is measurement of electrical activity of individual neurons with millisecond resolution. Current techniques rely on customized electrophysiological recording set-ups, characterized by limited throughput and poor integration with other readout modalities. Here we describe a novel approach, using multiwell glass microfluidic microelectrode arrays, allowing non-invasive electrical recording from engineered 3D neural tissues. We demonstrate parallelized studies with reference compounds, calcium imaging and optogenetic stimulation. Additionally, we show how microplate compatibility allows automated handling and high-content analysis of human induced pluripotent stem cell–derived neurons. This microphysiological platform opens up new avenues for high-throughput studies on the functional, morphological and molecular details of neurological diseases and their potential treatment by therapeutic compounds.

Microengineered 2D and 3D modular neuronal networks represent structure-function relationship

“…Even though in our 3D modular model the readout efficiency was 10 times higher than in random 2D networks, planar electrodes allow recording only from the bottom side of the network [53]. This challenge could be resolved by recently developed 3D MEA systems that offer multisite recording from 3D neural culture microsystems and organoids [55,59,78,79]. Nevertheless, the feasibility of constructing functional 3D modular networks on currently available models of 3D MEAs needs to be tested.…”

“…Self-assembled 100 µm thick 3D structure in each node was composed of neurons and astrocytes that was supported by PDL-coated microwells. Previous models were limited to engineering uniform and randomly connected 3D neural circuits on MEA substrates [53,57,59]. To mimic modular brain circuits we designed, for the first time, a 3D modular network on planar MEA electrodes with long-term access to network electrophysiology data.…”

“…To probe the activity dynamics of 3D networks, most groups have assembled a bulk 3D structure covering the MEA sensing areas and spaces between electrodes [53,56,57]. Regardless of their many advantages over 2D models, these 3D networks were made of randomly distributed and interconnected neurons [58,59]. Lately, some groups engineered modular 2D networks on MEAs by changing the degree of cellular aggregation or using PDMS microstructures to adjust the spatial arrangement of neurons [44,60].…”

Fully Integrated 3D Microelectrode Arrays with Polydopamine-Mediated Silicon Dioxide Insulation for Electrophysiological Interrogation of a Novel 3D Human, Neural Microphysiological Construct

“…In recent decades, the complexity of in vitro biological systems has increased dramatically through the development of “Organ-on-a-Chip” models, which replicate specific organ- or tissue-level systems through benchtop data collection. Similarly, microphysiological systems (MPS) aim to incorporate highly specific cellular models that replicate precise biological functions in a benchtop form factor with integrated sensing modalities. − Importantly, the emphasis of MPS modeling in vitro is motivated by several factors. First, physiologically representative laboratory models could equate to more robust preclinical data for human trials (e.g., drug candidate screening and mechanism of action data).…”

Microfluidics for Neuronal Cell and Circuit Engineering

“…Microfluidics and microfabrication technologies have been extensively used to develop intricate devices with integrated neural cell-sized microchannels. − These devices operate with volumes in the micro- and nanoliter scales and incorporate pumps, valves, and electrokinetic elements. − Thereby, not only are they compatible with rapid and directed transport of fluids but also support the straightforward automation and parallel execution of multiple operational steps. − Further, by depositing chemical cues in the physically confined spaces of these devices, it is also possible to control neural circuit architecture and function in vitro. ,, In addition, many microfluidic devices are also compatible with optical and electrophysiological tools that enable individual neurons to be monitored, manipulated, and examined. − In the past decade, the use of microfluidics has deepened our understanding of neurons and the circuits they form by enabling the isolation and molecular profiling of single cells from primary tissues − by supporting the in vitro engineering of neural cells − and the construction of 2- and 3-dimensional neural circuits with defined spatial orientations. ,− …”