Advancing a MEMS-Based 3D Cell Culture System for in vitro Neuro-Electrophysiological Recordings

Alex J Bastiaens,Teun Van Nunen,Erik F G A Homburg,Jean-Philippe Frimat,Bart Schurink,Regina Luttge

doi:10.3389/fmech.2018.00021

Abstract

In this work we present advances in three dimensional (3D) neuronal cell culture systems based on a reversible assembly of a microbioreactor with a microelectrode array (MEA) to create a MEMS-based 3D cell culture system for in vitro neuro-electrophysiological recordings. A batch of six molds were milled in poly (methyl methacrylate). The molds were used for soft lithography of polydimethylsiloxane (PDMS). In the center of the PDMS shape, a porous polyethersulfone (PES) cylindrical tube was press-fitted to form a growth barrier between the culture chamber inside the PES tube and the microfluidic channel surrounding the PES tube. A thin layer of partially cured PDMS was used to seal the bottom of the microbioreactor and provide reversible adhesion with the glass surface of a MEA. SH-SY5Y cells were successfully differentiated inside the microbioreactors in Matrigel and demonstrated extended neuronal networks over a height of at least 184 micrometers within the system. In previous microbioreactor designsvisibility was limited due to the closed top with the dispensing holes. The new open top design allows for a better evaluation of the cell culture by optical detection methods during the experiment. . Electrophysiological activity was recorded within the microbioreactor using human induced pluripotent stem cell-derived cortical neurons cultured in Matrigel, in 3D, up until 21 days in vitro In summary, we present advances made in the design, the fabrication process and integration of microbioreactors with MEAs. Optical imaging capabilities improved significantly with an open top and the culture time was further extended from 7 to 21 DIV without leakage or degradation thanks to introducing PES as a barrier material and an enhanced assembly procedure. The latter facilitated a sufficient long-term culture for neurons to mature in an environment free from flow-induced stress and provided a proof of principle for. the recording of electrophysiological activity of cortical neurons cultured in 3D.

Highlights

The advance of cell culture models employing threedimensional (3D) matrices for cells has proven to be of vital importance in studying cellular physiological and pathological responses
The PES tube locked into the PDMS firmly and the thin layer of partially cured PDMS underneath the microbioreactor ensured the PES was locked in place and when inserted into the microbioreactor, no fluids would circumvent the PES tube
In this work we present advances in 3D neuronal cell culture systems based on a reversible assembly of a microbioreactor with a microelectrode array (MEA) to create a microelectromechanical system (MEMS)-based 3D cell culture system for in vitro neuro-electrophysiological recordings

Summary

Introduction

The advance of cell culture models employing threedimensional (3D) matrices for cells has proven to be of vital importance in studying cellular physiological and pathological responses. Hydrogels are often employed to create these 3D microenvironments (Tibbitt and Anseth, 2009; Verhulsel et al, 2014). 3D cell cultures are performed in culture platforms with microelectromechanical system (MEMS) features and often microfluidic components to compensate for such issues and create small, sustainable culture volumes typically in the microliter range that are accessible for optical, chemical, or electrical analysis. We contribute to this research field with the advances made on our own microbioreactor design (Schurink and Luttge, 2013)

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