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Subventricular zone cell migration: lessons from quantitative two-photon microscopy.

James R, Kim Y, Hockberger PE, Szele FG - Front Neurosci (2011)

Bottom Line: This involved analyzing 3D stacks of images over time and uncovered several novel aspects of SVZ migration: chains remain stable, cells can be immotile for extensive periods, morphology does not necessarily correlate with motility, neuroblasts exhibit local exploratory motility, dorsoventral migration occurs throughout the striatal SVZ, and neuroblasts turn at distinctive angles.In this review we also discuss some technical considerations when setting up a two-photon microscope imaging system.Throughout the review we identify several unsolved questions about SVZ neuroblast migration that might be addressed with current or emerging techniques.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford Oxford, UK.

ABSTRACT
Neuroblasts born in the adult subventricular zone (SVZ) migrate long distances in the rostral migratory stream (RMS) to the olfactory bulbs where they integrate into circuitry as functional interneurons. As very little was known about the dynamic parameters of SVZ neuroblast migration, we used two-photon time-lapse microscopy to analyze migration in acute slices. This involved analyzing 3D stacks of images over time and uncovered several novel aspects of SVZ migration: chains remain stable, cells can be immotile for extensive periods, morphology does not necessarily correlate with motility, neuroblasts exhibit local exploratory motility, dorsoventral migration occurs throughout the striatal SVZ, and neuroblasts turn at distinctive angles. We investigated these novel findings in the SVZ and RMS from the population to the single cell level. In this review we also discuss some technical considerations when setting up a two-photon microscope imaging system. Throughout the review we identify several unsolved questions about SVZ neuroblast migration that might be addressed with current or emerging techniques.

No MeSH data available.


Flow chart of image analysis. (A) Raw z-stacks are captured with Fluoview software over time. Three actual stacks collected at 3 min scan intervals. We typically collect 50 Z optical sections whereas only five are illustrated for simplicity. (B) Images are imported into Volocity and rendered into 3D. (C) Images filtered in Volocity. (D) Quicktime movies are generated. (E) Cell positions are tracked in Volocity and exported to Graphis for illustration.
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Figure 2: Flow chart of image analysis. (A) Raw z-stacks are captured with Fluoview software over time. Three actual stacks collected at 3 min scan intervals. We typically collect 50 Z optical sections whereas only five are illustrated for simplicity. (B) Images are imported into Volocity and rendered into 3D. (C) Images filtered in Volocity. (D) Quicktime movies are generated. (E) Cell positions are tracked in Volocity and exported to Graphis for illustration.

Mentions: We have used Olympus Confocal software (Fluoview) to capture images (Figure 2A). Fluoview is rather easy to use and allows control over the number of optical slices, the distance between slices, the digital zoom and initial Kallman filtering. Once the time-lapse is completed files are imported into Volocity (Improvision) software and are rendered into 3D images (Figure 2B). Although Volocity can run into $10–30K depending on the suite of modules one chooses, we have found it ideal for subsequent filtering (Figure 2C) and especially for tracking and quantifying cells in 3D. Although the program requires a high-performance computer with large memory capacity, the computer should still only amount to 10–20% of cost of the image analysis system. We have found it extremely useful to routinely convert movies from Volocity into Quicktime files for archiving, rapid examination of parameters and for presentations (Figure 2D). To image the tracks taken by migrating cells we frequently export data to Graphis (Kylebank Software) which is convenient for visualizing the pathways from multiple angles (Figure 2E).


Subventricular zone cell migration: lessons from quantitative two-photon microscopy.

James R, Kim Y, Hockberger PE, Szele FG - Front Neurosci (2011)

Flow chart of image analysis. (A) Raw z-stacks are captured with Fluoview software over time. Three actual stacks collected at 3 min scan intervals. We typically collect 50 Z optical sections whereas only five are illustrated for simplicity. (B) Images are imported into Volocity and rendered into 3D. (C) Images filtered in Volocity. (D) Quicktime movies are generated. (E) Cell positions are tracked in Volocity and exported to Graphis for illustration.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3064983&req=5

Figure 2: Flow chart of image analysis. (A) Raw z-stacks are captured with Fluoview software over time. Three actual stacks collected at 3 min scan intervals. We typically collect 50 Z optical sections whereas only five are illustrated for simplicity. (B) Images are imported into Volocity and rendered into 3D. (C) Images filtered in Volocity. (D) Quicktime movies are generated. (E) Cell positions are tracked in Volocity and exported to Graphis for illustration.
Mentions: We have used Olympus Confocal software (Fluoview) to capture images (Figure 2A). Fluoview is rather easy to use and allows control over the number of optical slices, the distance between slices, the digital zoom and initial Kallman filtering. Once the time-lapse is completed files are imported into Volocity (Improvision) software and are rendered into 3D images (Figure 2B). Although Volocity can run into $10–30K depending on the suite of modules one chooses, we have found it ideal for subsequent filtering (Figure 2C) and especially for tracking and quantifying cells in 3D. Although the program requires a high-performance computer with large memory capacity, the computer should still only amount to 10–20% of cost of the image analysis system. We have found it extremely useful to routinely convert movies from Volocity into Quicktime files for archiving, rapid examination of parameters and for presentations (Figure 2D). To image the tracks taken by migrating cells we frequently export data to Graphis (Kylebank Software) which is convenient for visualizing the pathways from multiple angles (Figure 2E).

Bottom Line: This involved analyzing 3D stacks of images over time and uncovered several novel aspects of SVZ migration: chains remain stable, cells can be immotile for extensive periods, morphology does not necessarily correlate with motility, neuroblasts exhibit local exploratory motility, dorsoventral migration occurs throughout the striatal SVZ, and neuroblasts turn at distinctive angles.In this review we also discuss some technical considerations when setting up a two-photon microscope imaging system.Throughout the review we identify several unsolved questions about SVZ neuroblast migration that might be addressed with current or emerging techniques.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford Oxford, UK.

ABSTRACT
Neuroblasts born in the adult subventricular zone (SVZ) migrate long distances in the rostral migratory stream (RMS) to the olfactory bulbs where they integrate into circuitry as functional interneurons. As very little was known about the dynamic parameters of SVZ neuroblast migration, we used two-photon time-lapse microscopy to analyze migration in acute slices. This involved analyzing 3D stacks of images over time and uncovered several novel aspects of SVZ migration: chains remain stable, cells can be immotile for extensive periods, morphology does not necessarily correlate with motility, neuroblasts exhibit local exploratory motility, dorsoventral migration occurs throughout the striatal SVZ, and neuroblasts turn at distinctive angles. We investigated these novel findings in the SVZ and RMS from the population to the single cell level. In this review we also discuss some technical considerations when setting up a two-photon microscope imaging system. Throughout the review we identify several unsolved questions about SVZ neuroblast migration that might be addressed with current or emerging techniques.

No MeSH data available.