Protocol for the enhanced analysis of electrophysiological data from high-density multi-electrode arrays with nicespike and spikeNburst

High-density multi-electrode arrays enable the recording of in vitro neuronal activity with exceptional spatial and temporal resolution. Here, we describe a protocol for analyzing these extensive datasets by using two complementary tools. The nicespike tool implements a full electrophysiological data analysis pipeline featuring graphics processing unit–accelerated spike sorting via template matching with Kilosort , enabling accurate identification of neuronal units across multiple electrodes. The spikeNburst tool incorporates advanced techniques for spike train filtering, burst and network burst detection, and synchronization analysis. • Instructions for setting up graphical user interfaces for HD-MEA data analysis • Instructions for analyzing voltage traces from HD-MEA recordings using spike sorting • Procedures for characterizing spike-sorted units and neural network dynamics • Guidance on extending the analysis for HD-MEA devices from various manufacturers Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. High-density multi-electrode arrays enable the recording of in vitro neuronal activity with exceptional spatial and temporal resolution. Here, we describe a protocol for analyzing these extensive datasets by using two complementary tools. The nicespike tool implements a full electrophysiological data analysis pipeline featuring graphics processing unit–accelerated spike sorting via template matching with Kilosort, enabling accurate identification of neuronal units across multiple electrodes. The spikeNburst tool incorporates advanced techniques for spike train filtering, burst and network burst detection, and synchronization analysis.