Modulating electrophysiology of motor neural networks via optogenetic stimulation during neurogenesis and synaptogenesis

Abstract

control of electrical activity in neural circuits through network training is a grand challenge for biomedicine and engineering applications. Past efforts have not considered evoking long-term changes in firing patterns of in-vitro networks by introducing training regimens with respect to stages of neural development. Here, we used Channelrhodopsin-2 (ChR2) transfected mouse embryonic stem cell (mESC) derived motor neurons to explore short and long-term programming of neural networks by using optical stimulation implemented during neurogenesis and synaptogenesis. not only did we see a subsequent increase of neurite extensions and synaptophysin clustering, but by using electrophysiological recording with micro electrode arrays (MEA) we also observed changes in signal frequency spectra, increase of network synchrony, coordinated firing of actions potentials, and enhanced evoked response to stimulation during network formation. our results demonstrate that optogenetic stimulation during neural differentiation can result in permanent changes that extended to the genetic expression of neurons as demonstrated by RNA Sequencing. To our knowledge, this is the first time that a correlation between training regimens during neurogenesis and synaptogenesis and the resulting plastic responses has been shown in-vitro and traced back to changes in gene expression. This work demonstrates new approaches for training of neural circuits whose electrical activity can be modulated and enhanced, which could lead to improvements in neurodegenerative disease research and engineering of in-vitro multi-cellular living systems. Inducing neuronal plasticity is one of the grand challenges in neuroengineering. Understanding and controlling nerve connectivity and their plasticity could have profound impacts in regenerative medicine 1-4 , as studies have shown that engrafting motor neuron containing embryoid bodies (MEBs) can improve recovery in motor nerve injuries 1,3. The ability to enhance or control the electrophysiological functions of such MEBs could be used for improvement of motor function recovery. Furthermore, in the emerging field of engineering biohybrid neuronal-driven biological machines, it would be highly advantageous to forward-engineer programmable neural networks that could be installed within in vivo or in vitro systems in order to achieve targeted functional behaviors 5-7. While the mechanisms responsible for synaptic modulation and circuitry formation (i.e. memory and learning) have been studied in invertebrate and mammals, the complexity of neuronal plasticity pathways,

3D high-density microelectrode array with optical stimulation and drug delivery for investigating neural circuit dynamics

“…Also, we observed a higher firing rate in the first than the second somatic region due to active burst activity recorded from electrodes (E5 and E6) around the stimulation site. This result could be attributed to enhanced local network formation in the first somatic region by repetitive neural activation through daily optical stimulation 55 . With increasing connectivity between the two somatic regions via the neurite region from DIV 11 to DIV 13 (Supplementary Figs.…”

A human stem cell-derived test system for agents modifying neuronal N-methyl-d-aspartate-type glutamate receptor Ca2+-signalling

“…The hitherto most robust MEA data have been generated with rat primary neurons. More recently, murine and human-stem cell-derived neurons have been demonstrated to be principally suitable for the analysis of network signalling on MEA (Pagan-Diaz et al 2020 ; Tukker et al 2020 ). However, the production of cultures at a reasonable price, time effort and robustness has proven challenging.…”

Astrocyte-mediated transduction of muscle fiber contractions synchronizes hippocampal neuronal network development

“…The synchrony of spike trains between electrodes was assessed through cross-correlation for discrete functions (Pagan-Diaz et al, 2020). There are total nine electrodes and spike train data for each electrode was cross-correlated with every other electrode.…”