Animal and human stem cell–derived three-dimensional models to study physio-pathological brain functioning are becoming a gold standard for in vitro electrophysiology, as they enable the recapitulation of complex network properties by accounting for spatial architectural features that better reflect in vivo conditions than simpler 2D models. Standard planar multielectrode arrays (MEAs), typically providing tens of recording electrodes, are commonly used to record activity from 2D neuronal cultures. However, when adapted for use with 3D models, planar 2D MEAs showed limited effectiveness. The main issues are limited specimen adhesion to the chip, a low number of sensing elements, inability to retrieve signals from within the tissue, and reduced perfusion and vitality of the tissue in contact with sensors. To overcome these limitations, a new generation of microchip-based 3D high-density MEAs (3D HD-MEA) has been developed and validated in recent years. This technological advancement has improved the sensing capabilities and the vitality of 3D models, providing a tool tailored to maximize their potential. Here, we present an optimized protocol for neural network activity recordings in 3D models (including acute slices, brain spheroids, and organoids) from various brain regions using 3D HD-MEAs. First, we summarize the critical steps for 1) obtaining viable acute slices from the mouse cerebellum, cortico-hippocampal circuit, and prefrontal cortex, 2) establishing efficient coupling of the slices with the chip, and 3) performing recordings and analyses. We then describe the main procedures required to obtain human and animal brain spheroids and neural organoids, as well as standardized routines to perform effective recordings and analyses. For each section, we highlight the crucial steps, identify tips for specific applications, and propose troubleshooting procedures. For example, the same type of preparation (e.g., acute slices) requires different adjustments when working with different brain areas. The specific information provided here is intended to assist researchers in their daily efforts to obtain efficient and reproducible functional recordings from 3D models by using the cutting-edge technique of 3D HD-MEA.
Measuring Electrophysiological Activity in Acute Brain Slices, Spheroids, and Organoids Using 3D High-Density Multielectrode Arrays
Pali E.;Pellavio G.;Conforti M.;Mainardi F.;D'Angelo E.;Mapelli L.
2026-01-01
Abstract
Animal and human stem cell–derived three-dimensional models to study physio-pathological brain functioning are becoming a gold standard for in vitro electrophysiology, as they enable the recapitulation of complex network properties by accounting for spatial architectural features that better reflect in vivo conditions than simpler 2D models. Standard planar multielectrode arrays (MEAs), typically providing tens of recording electrodes, are commonly used to record activity from 2D neuronal cultures. However, when adapted for use with 3D models, planar 2D MEAs showed limited effectiveness. The main issues are limited specimen adhesion to the chip, a low number of sensing elements, inability to retrieve signals from within the tissue, and reduced perfusion and vitality of the tissue in contact with sensors. To overcome these limitations, a new generation of microchip-based 3D high-density MEAs (3D HD-MEA) has been developed and validated in recent years. This technological advancement has improved the sensing capabilities and the vitality of 3D models, providing a tool tailored to maximize their potential. Here, we present an optimized protocol for neural network activity recordings in 3D models (including acute slices, brain spheroids, and organoids) from various brain regions using 3D HD-MEAs. First, we summarize the critical steps for 1) obtaining viable acute slices from the mouse cerebellum, cortico-hippocampal circuit, and prefrontal cortex, 2) establishing efficient coupling of the slices with the chip, and 3) performing recordings and analyses. We then describe the main procedures required to obtain human and animal brain spheroids and neural organoids, as well as standardized routines to perform effective recordings and analyses. For each section, we highlight the crucial steps, identify tips for specific applications, and propose troubleshooting procedures. For example, the same type of preparation (e.g., acute slices) requires different adjustments when working with different brain areas. The specific information provided here is intended to assist researchers in their daily efforts to obtain efficient and reproducible functional recordings from 3D models by using the cutting-edge technique of 3D HD-MEA.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
