Direkt zum Inhalt

Owner only: item control page
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 and Daria, Vincent R. (2019) A compact holographic projector module for high-resolution 3D multi-site two-photon photostimulation. PLOS ONE 14 (1), e0210564.

Date of publication of this fulltext: 07 Feb 2019 10:20
Article
DOI to cite this document: 10.5283/epub.38312


Abstract

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.



Involved Institutions


Details

Item typeArticle
Journal or Publication TitlePLOS ONE
Publisher:PLOS
Open Access Type:Gold (with APC)
Place of Publication:SAN FRANCISCO
Volume:14
Number of Issue or Book Chapter:1
Page Range:e0210564
Date28 January 2019
InstitutionsBiology, Preclinical Medicine > Institut für Zoologie > Neurophysiologie (Prof. Dr. Veronica Egger)
Biology, Preclinical Medicine > Institut für Zoologie > Neurophysiologie (Prof. Dr. Veronica Egger)
Identification Number
ValueType
10.1371/journal.pone.0210564DOI
KeywordsDENDRITIC SPINES; OPTICAL TWEEZERS; MICROSCOPY; EXCITATION; NETWORK; INTEGRATION; CALCIUM;
Dewey Decimal Classification500 Science > 570 Life sciences
StatusPublished
RefereedYes, this version has been refereed
Created at the University of RegensburgYes
URN of the UB Regensburgurn:nbn:de:bvb:355-epub-383127
Item ID38312

Export bibliographical data

Owner only: item control page

nach oben