Comprehensive Imaging of Sensory-Evoked Activity of Entire Neurons Within the Awake Developing Brain Using Ultrafast AOD-Based Random-Access Two-Photon Microscopy.

TitleComprehensive Imaging of Sensory-Evoked Activity of Entire Neurons Within the Awake Developing Brain Using Ultrafast AOD-Based Random-Access Two-Photon Microscopy.
Publication TypeJournal Article
Year of Publication2020
AuthorsSakaki KDR, Podgorski K, Toth TADellaziz, Coleman P, Haas K
JournalFront Neural Circuits
Volume14
Pagination33
Date Published2020
ISSN1662-5110
KeywordsAcoustic Stimulation, Animals, Brain, Brain Chemistry, Evoked Potentials, Visual, Microscopy, Fluorescence, Multiphoton, Neurons, Optical Phenomena, Photic Stimulation, Superior Colliculi, Time Factors, Wakefulness, Xenopus laevis
Abstract

Determining how neurons transform synaptic input and encode information in action potential (AP) firing output is required for understanding dendritic integration, neural transforms and encoding. Limitations in the speed of imaging 3D volumes of brain encompassing complex dendritic arbors using conventional galvanometer mirror-based laser-scanning microscopy has hampered fully capturing fluorescent sensors of activity throughout an individual neuron's entire complement of synaptic inputs and somatic APs. To address this problem, we have developed a two-photon microscope that achieves high-speed scanning by employing inertia-free acousto-optic deflectors (AODs) for laser beam positioning, enabling random-access sampling of hundreds to thousands of points-of-interest restricted to a predetermined neuronal structure, avoiding wasted scanning of surrounding extracellular tissue. This system is capable of comprehensive imaging of the activity of single neurons within the intact and awake vertebrate brain. Here, we demonstrate imaging of tectal neurons within the brains of albino tadpoles labeled using single-cell electroporation for expression of a red space-filling fluorophore to determine dendritic arbor morphology, and either the calcium sensor jGCaMP7s or the glutamate sensor iGluSnFR as indicators of neural activity. Using discrete, point-of-interest scanning we achieve sampling rates of 3 Hz for saturation sampling of entire arbors at 2 μm resolution, 6 Hz for sequentially sampling 3 volumes encompassing the dendritic arbor and soma, and 200-250 Hz for scanning individual planes through the dendritic arbor. This system allows investigations of sensory-evoked information input-output relationships of neurons within the intact and awake brain.

DOI10.3389/fncir.2020.00033
Alternate JournalFront Neural Circuits
PubMed ID32612514
PubMed Central IDPMC7308460
Grant ListFDN-148468 / / CIHR / Canada