Integrated Microprism and Microelectrode Array for Simultaneous Electrophysiology and Two-Photon Imaging across All Cortical Layers

Cerebral neural electronics play a crucial role in neuroscience research with increasing translational applications such as brain-computer interfaces for sensory input and motor output restoration. While widely utilized for decades, the understanding of the cellular mechanisms underlying this technology remains limited. Although two-photon microscopy (TPM) has shown great promise in imaging superficial neural electrodes, its application to deep-penetrating electrodes is technically difficult. Here, a novel device integrating transparent microelectrode arrays with glass microprisms, enabling electrophysiology recording and stimulation alongside TPM imaging across all cortical layers in a vertical plane, is introduced. Tested in Thy1-GCaMP6 mice for over 4 months, the integrated device demonstrates the capability for multisite electrophysiological recording/stimulation and simultaneous TPM calcium imaging. As a proof of concept, the impact of microstimulation amplitude, frequency, and depth on neural activation patterns is investigated using the setup. With future improvements in material stability and single unit yield, this multimodal tool greatly expands integrated electrophysiology and optical imaging from the superficial brain to the entire cortical column, opening new avenues for neuroscience research and neurotechnology development. A 16-channel transparent microelectrode array is integrated with a microprism to allow simultaneous electrophysiology and optical imaging throughout all cortical layers. The authors achieved single-unit recording, microstimulation, and simultaneous two-photon imaging of the whole cortical column in awake mice for over 4 months. Concurrent microstimulation and two-photon imaging reveal the depth dependency of various stimulation parameters. image