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Electrical Identification and Selective Microstimulation of Neuronal Compartments Based on Features of Extracellular Action Potentials

Milos Radivojevic1*, David Jäckel2, Michael Altermatt2, Jan Müller3, Vijay Viswam1, Andreas Hierlemann1 and Douglas Bakkum1
  • 1 ETH Zurich, Department of Biosystems Science and Engineering, Switzerland
  • 2 ETH Zurich, Department of Biosystems Science and Engineering, Switzerland
  • 3 ETH Zurich, Department of Biosystems Science and Engineering, Switzerland

Electrical Identification and Selective Microstimulation of Neuronal Compartments Based on Features of Extracellular Action Potentials Milos Radivojevic, David Jäckel, Michael Altermatt, Jan Müller, Vijay Viswam, Andreas Hierlemann and Douglas Bakkum ETH Zürich, Department of Biosystems Science and Engineering, Basel, Switzerland Corresponding author’s email address: milos.radivojevic@bsse.ethz.ch Keywords: cortical cultures, electrode-neuron interface, selective stimulation, patch-clamp Motivation It is generally acknowledged that complex operations performed by neuronal circuits arise from the orchestrated activities of individual neurons. Therefore, the ability to simultaneously observe and elicit the activity of selected individual neurons at cellular and subcellular resolution over extended periods of time will be pivotal for exploring some of the basic governing principles in neural circuits. Furthermore, the design of highly controllable protocols for reliable and selective extracellular stimulation of individual neurons is an important task in the development of neural prosthetics and brain-machine interfaces, where advances in electrode-neuron interfacing technology and the detailed characterization of neuronal responses to local electrical fields can lead to improved clinical outcomes. Here, we studied cultured neocortical neurons by using high-density microelectrode arrays and optical imaging, complemented by the patch-clamp technique, and with the aim to correlate morphological and electrical features of neuronal compartments with their responsiveness to extracellular stimulation. Materials and Methods We cultured primary rat cortical neurons on a high-density (HD) complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs) developed in our laboratory [1]. Comprised of 11,011 densely-packed microelectrodes, the array enables detecting action potentials arising from different compartments of a single neuron, including tiny axonal branches [2]. Each electrode can provide noninvasive recording and/or stimulation over extended periods of time (weeks), enabling bi-directional access to any neuronal site with subcellular spatial resolution. We combined HD-MEAs, patch-clamp, immunocytochemistry and live imaging of individual neurons in the culture in order to monitor suprathreshold (extracellular) and subthreshold (intracellular) electrical behavior of the cell driven by extracellular stimulation directed at different neuronal compartments. Results and Discussion We developed a set of recording and stimulation strategies to electrically visualize individual neurons in the network. Optical imaging was used to correlate extracellular electrical activity of different neuronal compartments with their morphology and to verify our electrical identification of the axon initial segment (AIS), axonal trunk, and lower order branches. Extracellular traces also enabled electrical detection of soma and proximal dendrites, however, unambiguous localization of these compartments required optical verification. In comparison to other neuronal regions, threshold stimulation near the AIS required the lowest voltage and provided immediate, selective and reliable orthodromic activation of the cell. Axonal stimulation revealed significant variations in the threshold voltages, which were discontinuously dispersed across the entire arbor. Threshold voltages of highly excitable axonal hotspots were comparable to that of the AIS and provided selective antidromic activation. Somatodendritic compartments either required high stimulation voltages to initiate an action potential (AP) or did not initiate an AP for voltages within a physiologically harmless range. Subthreshold somatic stimulation depolarized the resting membrane potential without a delay; subthreshold depolarization in the soma was not observed after axonal or AIS stimulation. Conclusion We developed a method to electrically identify and stimulate any specific neuron in isolated cortical networks noninvasively, selectively, and reliably. Threshold stimulation at the AIS replicated spontaneous action potential initiation location and propagation, whereas subthreshold somatic stimulation depolarized the somatic membrane potential. References 1. Frey, Urs, et al. "Switch-matrix-based high-density microelectrode array in CMOS technology." Solid-State Circuits, IEEE Journal of 45.2 (2010): 467-482. 2. Bakkum, Douglas J., et al. "Tracking axonal action potential propagation on a high-density microelectrode array across hundreds of sites." Nature communications 4 (2013). Acknowledgements Financial support through the ERC Advanced Grant 267351 “NeuroCMOS” and the Swiss National Science Foundation Grant 205321_157092/1 is acknowledged.

Keywords: patch-clamp, Selective stimulation, Cortical cultures, electrode-neuron interface

Conference: MEA Meeting 2016 | 10th International Meeting on Substrate-Integrated Electrode Arrays, Reutlingen, Germany, 28 Jun - 1 Jul, 2016.

Presentation Type: oral

Topic: MEA Meeting 2016

Citation: Radivojevic M, Jäckel D, Altermatt M, Müller J, Viswam V, Hierlemann A and Bakkum D (2016). Electrical Identification and Selective Microstimulation of Neuronal Compartments Based on Features of Extracellular Action Potentials. Front. Neurosci. Conference Abstract: MEA Meeting 2016 | 10th International Meeting on Substrate-Integrated Electrode Arrays. doi: 10.3389/conf.fnins.2016.93.00021

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Received: 22 Jun 2016; Published Online: 24 Jun 2016.

* Correspondence: Dr. Milos Radivojevic, ETH Zurich, Department of Biosystems Science and Engineering, Basel, Switzerland, milos.radivojevic@bsse.ethz.ch