Massively scalable 3D electrophysiology and two-photon imaging in freely-moving animals

Purdue University, West Lafayette IN

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Abstract

SUMMARY Revealing how neural circuits encode and enable behavioral experiences is a fundamental problem in neuroscience. Decoding and dissecting the mechanisms of signal flow across such circuits necessitates the ability to record millisecond electrical dynamics and simultaneously map the spatial organization of cellular and sub-cellular circuit motifs, in awake behaving animals. During natural behavior, animals actively acquire sensory information as they move through the environment and use this information to guide ongoing actions. In this context, unconstrained movement-related signals could allow sensory systems to efficiently predict self- generated motion and extract additional information about the environment, thereby forming a stable internal representation of the external world. However, a majority of recordings are performed in head-fixed animals which imposes severe restrictions on how movement related signals shape ongoing sensory and memory processing in the brain. Performing high-density electrophysiology and concomitant two-photon calcium imaging is -at present- not feasible due to technical limitations and are therefore performed separately in both head fixed and freely moving preparations. There is a great need for technology platforms that can combine high-resolution electrical recordings across entire volumes of brain tissue and two-photon calcium imaging in freely behaving animals. In this proposal we introduce a new paradigm for high-density electrophysiology across 3D volume with capabilities to simultaneously perform two-photon calcium imaging. Our innovation termed NET-2P, comprises of 3D Nanoelectrodes of variable height integrated onto the baseplate of a head-mounted mini two-photon microscope. The Nanoelectrode array is integrated with custom-designed CMOS electronics and will in total weigh 3.5 grams. The transparent and flexible nature of the nanoelectrode array allows for easy two-photon access whilst facilitating rapid electrical mapping across large cortical sections. We propose to use the head- mounted setup to 1) assay cortical travelling waves under tactile processing whilst mapping the underlying cellular scale ensemble map via two-p imaging and 2) unravel how cortical ensembles and travelling waves that emerge after a learning task enhance memory consolidation during sleep. In preliminary experiments performed in head-fixed awake animals under passive whisker touch, we used planar transparent electrode array recordings and conventional two-photon calcium imaging, and discovered microscopic travelling waves upon whisker touch, a late reverberatory wave 100ms post touch, and sparse yet stable cellular ensemble structure that supports wave propagation. We hypothesize that spontaneous travelling waves, including late reverberatory components that emerge hundreds of milliseconds post stimulus, carry movement related, head-direction, and volitional control signals, which will enhance travelling wave dynamics in freely-moving mice. By combining electrophysiology and imaging-based ensemble mapping during natural sleep we will assay how spiking and synaptic changes across cortical layers strengthen and enable robust functional cellular activity landscapes.