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Go, Mary Ann ; Mueller, Max ; Castañares, Michael Lawrence ; Egger, Veronica ; Daria, Vincent R.

A compact holographic projector module for high-resolution 3D multi-site two-photon photostimulation

Go, Mary Ann, Mueller, Max, Castañares, Michael Lawrence, Egger, Veronica und Daria, Vincent R. (2019) A compact holographic projector module for high-resolution 3D multi-site two-photon photostimulation. PLOS ONE 14 (1), e0210564.

Veröffentlichungsdatum dieses Volltextes: 07 Feb 2019 10:20
Artikel
DOI zum Zitieren dieses Dokuments: 10.5283/epub.38312


Zusammenfassung

Patterned two-photon (2P) photolysis via holographic illumination is a powerful method to investigate neuronal function because of its capability to emulate multiple synaptic inputs in three dimensions (3D) simultaneously. However, like any optical system, holographic projectors have a finite space-bandwidth product that restricts the spatial range of patterned illumination or field-of-view (FOV) ...

Patterned two-photon (2P) photolysis via holographic illumination is a powerful method to investigate neuronal function because of its capability to emulate multiple synaptic inputs in three dimensions (3D) simultaneously. However, like any optical system, holographic projectors have a finite space-bandwidth product that restricts the spatial range of patterned illumination or field-of-view (FOV) for a desired resolution. Such trade-off between holographic FOV and resolution restricts the coverage within a limited domain of the neuron's dendritic tree to perform highly resolved patterned 2P photolysis on individual spines. Here, we integrate a holographic projector into a commercial 2P galvanometer-based 2D scanning microscope with an uncaging unit and extend the accessible holographic FOV by using the galvanometer scanning mirrors to reposition the holographic FOV arbitrarily across the imaging FOV. The projector system utilizes the microscope's built-in imaging functions. Stimulation positions can be selected from within an acquired 3D image stack (the volume-of-interest, VOI) and the holographic projector then generates 3D illumination patterns with multiple uncaging foci. The imaging FOV of our system is 800x800 mu m(2) within which a holographic VOI of 70x70x70 mu m(3) can be chosen at arbitrary positions and also moved during experiments without moving the sample. We describe the design and alignment protocol as well as the custom software plugin that controls the 3D positioning of stimulation sites. We demonstrate the neurobiological application of the system by simultaneously uncaging glutamate at multiple spines within dendritic domains and consequently observing summation of postsynaptic potentials at the soma, eventually resulting in action potentials. At the same time, it is possible to perform two-photon Ca2+ imaging in 2D in the dendrite and thus to monitor synaptic Ca2+ entry in selected spines and also local regenerative events such as dendritic action potentials.



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Details

DokumentenartArtikel
Titel eines Journals oder einer ZeitschriftPLOS ONE
Verlag:PLOS
Ort der Veröffentlichung:SAN FRANCISCO
Band:14
Nummer des Zeitschriftenheftes oder des Kapitels:1
Seitenbereich:e0210564
Datum28 Januar 2019
InstitutionenBiologie und Vorklinische Medizin > Institut für Zoologie > Neurophysiologie (Prof. Dr. Veronica Egger)
Biologie und Vorklinische Medizin > Institut für Zoologie > Neurophysiologie (Prof. Dr. Veronica Egger)
Identifikationsnummer
WertTyp
10.1371/journal.pone.0210564DOI
Stichwörter / KeywordsDENDRITIC SPINES; OPTICAL TWEEZERS; MICROSCOPY; EXCITATION; NETWORK; INTEGRATION; CALCIUM;
Dewey-Dezimal-Klassifikation500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie
StatusVeröffentlicht
BegutachtetJa, diese Version wurde begutachtet
An der Universität Regensburg entstandenJa
URN der UB Regensburgurn:nbn:de:bvb:355-epub-383127
Dokumenten-ID38312

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