Characterization of spontaneous and evoked activity of mouse embryonic stem cell derived motor neurons using optogenetics and micro electrode array electrophysiology

Multi-electrode array (MEA);microelectrode array;mES cells, embryoid bodies;electrophysiology

Pagán-Díaz, Gelson J ; Sengupta ; Parijat ; Bahisr ; Rashid

关键词: Multi-electrode array (MEA);    microelectrode array;    mES cells, embryoid bodies;    electrophysiology;   

Others  :  https://www.ideals.illinois.edu/bitstream/handle/2142/90854/PAGAN-DIAZ-THESIS-2016.pdf?sequence=1&isAllowed=y

美国|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

Thorough characterization of distinct neuronal lineages derived from progenitor cells is essential for the development of biological models that can recreate native function of neurons of the central nervous system while permitting researchers to have an easily accessible source of cellular building blocks. These models are particularly important for studies on neurological disorders aimed at understanding disease pathways and for high throughput testing of drug candidates. Furthermore, developing a model system with stem cell-derived spinal cord motor neurons (MN) is attractive for designing control systems for soft-tissue robotics. The critical first step towards building a useful model with MNs require a detail investigation of spontaneous and stimulation-evoked electrical activity of developing MN networks. The following work uses multi-electrode array (MEA) electrophysiology and optogenetics to characterize electrical activity of MNs differentiated from mouse embryonic stem (mES) cells. A customizable substrate integrated 60-electrode MEA chip was designed for this study. MEA chips were fabricated with platinum deposited on borofloat glass for detection of small changes in electrical field potentials resulting from neuronal activity that causes small change in ionic currents. A Multi-Channel Systems amplifier was used for recording. MN embryoid bodies (MEBs) were grown in these MEA chips, and differentiation of mES cells into MNs was monitored by the expression of eGFP with a MN specific promoter, Hb9. The mES cell line was transfected with channelrhodopsin-2 (ChR2) tagged with Td-Tomato which allowed for optogenetic stimulation of the networks with a 470 nm LED. Network firing patterns were evaluated for bursting activity and spectral content using analysis algorithms developed in MATLAB. Data presented here demonstrated that MEBs are spontaneously active, they develop a robust network synchronization, and optogenetic stimulation increased the firing rate and affected the firing patterns. This work established a model system with mES cell derived MNs. These findings are a milestone in the efforts of developing neural circuits that can be used to potentially control higher order soft-tissue robotics.