article · 09/04/2013
Fast functional imaging of multiple brain regions in intact zebrafish larvae using Selective Plane Illumination Microscopy
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Résumé
The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2\% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30\% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a similar or equal to 20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed.
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Panier, T., Romano, S.-A., Olive, R., Pietri, T., Sumbre, G., Candelier, R., & Debregeas, G. (2013). Fast functional imaging of multiple brain regions in intact zebrafish larvae using Selective Plane Illumination Microscopy. Frontiers in Neural Circuits, 7(65). https://doi.org/10.3389/fncir.2013.00065@article{Panier2013_5,
author = {Panier, Thomas and Romano, Sebastian A. and Olive, Raphael and Pietri, Thomas and Sumbre, German and Candelier, Raphael and Debregeas, Georges},
year = {2013},
month = {4},
title = {Fast functional imaging of multiple brain regions in intact zebrafish larvae using Selective Plane Illumination Microscopy},
journal = {Frontiers in Neural Circuits},
publisher = {FRONTIERS RESEARCH FOUNDATION},
volume = {7},
number = {65},
address = {PO BOX 110, LAUSANNE, 1015, SWITZERLAND},
abstract = {The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2\textbackslash{}\% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30\textbackslash{}\% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a similar or equal to 20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed.},
url = {http://www.dx.doi.org/10.3389/fncir.2013.00065},
doi = {10.3389/fncir.2013.00065},
issn = {1662-5110},
}TY - JOUR
AU - Panier, Thomas
AU - Romano, Sebastian A.
AU - Olive, Raphael
AU - Pietri, Thomas
AU - Sumbre, German
AU - Candelier, Raphael
AU - Debregeas, Georges
PY - 2013
DA - 2013/04/09
TI - Fast functional imaging of multiple brain regions in intact zebrafish larvae using Selective Plane Illumination Microscopy
JO - Frontiers in Neural Circuits
VL - 7
IS - 65
PB - FRONTIERS RESEARCH FOUNDATION
SN - 1662-5110
AB - The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2\% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30\% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a similar or equal to 20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed.
DO - 10.3389/fncir.2013.00065
UR - http://www.dx.doi.org/10.3389/fncir.2013.00065
ER -