title: Hybrid multiphoton volumetric functional imaging of large-scale bioengineered neuronal networks.
abstract: Planar neural networks and interfaces serve as versatile in vitro models of central nervous system physiology, but adaptations of related methods to three dimensions (3D) have met with limited success. Here, we demonstrate for the first time volumetric functional imaging in a bioengineered neural tissue growing in a transparent hydrogel with cortical cellular and synaptic densities, by introducing complementary new developments in nonlinear microscopy and neural tissue engineering. Our system uses a novel hybrid multiphoton microscope design combining a 3D scanning-line temporal-focusing subsystem and a conventional laser-scanning multiphoton microscope to provide functional and structural volumetric imaging capabilities: dense microscopic 3D sampling at tens of volumes per second of structures with mm-scale dimensions containing a network of over 1,000 developing cells with complex spontaneous activity patterns. These developments open new opportunities for large-scale neuronal interfacing and for applications of 3D engineered networks ranging from basic neuroscience to the screening of neuroactive substances.
authors.affiliation: Department of Biomedical Engineering, Technion, Haifa 32000, Israel.
authors.forename: Hod
authors.initials: H
authors.lastname: Dana
authors.affiliation: Department of Biomedical Engineering, Technion, Haifa 32000, Israel.|The Interdepartmental Program in Biotechnology, Technion, Haifa 32000, Israel.
authors.forename: Anat
authors.initials: A
authors.lastname: Marom
authors.affiliation: Department of Biomedical Engineering, Technion, Haifa 32000, Israel.
authors.forename: Shir
authors.initials: S
authors.lastname: Paluch
authors.affiliation: Faculty of Medicine and Network Biology Research Laboratories. Technion, Haifa 32000, Israel.
authors.forename: Roman
authors.initials: R
authors.lastname: Dvorkin
authors.affiliation: Department of Biomedical Engineering, Technion, Haifa 32000, Israel.
authors.forename: Inbar
authors.initials: I
authors.lastname: Brosh
authors.affiliation: Department of Biomedical Engineering, Technion, Haifa 32000, Israel.
authors.forename: Shy
authors.initials: S
authors.lastname: Shoham
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002540:Cerebral Cortex; D059907:Functional Neuroimaging / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D066298:In Vitro Techniques; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D008959:Models, Neurological; D009415:Nerve Net; D009474:Neurons / Q000502:physiology* / Q000648:ultrastructure; D051381:Rats; D023822:Tissue Engineering