BrainAligner: 3D registration atlases of Drosophila brains - PubMed (original) (raw)

BrainAligner: 3D registration atlases of Drosophila brains

Hanchuan Peng et al. Nat Methods. 2011 Jun.

Abstract

Analyzing Drosophila melanogaster neural expression patterns in thousands of three-dimensional image stacks of individual brains requires registering them into a canonical framework based on a fiducial reference of neuropil morphology. Given a target brain labeled with predefined landmarks, the BrainAligner program automatically finds the corresponding landmarks in a subject brain and maps it to the coordinate system of the target brain via a deformable warp. Using a neuropil marker (the antibody nc82) as a reference of the brain morphology and a target brain that is itself a statistical average of data for 295 brains, we achieved a registration accuracy of 2 μm on average, permitting assessment of stereotypy, potential connectivity and functional mapping of the adult fruit fly brain. We used BrainAligner to generate an image pattern atlas of 2954 registered brains containing 470 different expression patterns that cover all the major compartments of the fly brain.

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Figures

Figure 1

Figure 1

BrainAligner registers images of neurons from different brains onto a common coordinate system. (a–b) Maximum intensity projections of confocal images of a64-GAL4 and a74-GAL4 brains. Neurons are visualized by membrane-targeted GFP and brain morphology is visualized by staining with the antibody nc82. (c) Aligned and overlaid neuronal patterns of (a) and (b). (d) Alignment of many GAL4 expression patterns. Patterns of interest can be selected and displayed in the common coordinate system. R1 and R2, regions of interest. (e) V3D-AtlasViewer software for viewing the 3D pattern atlas. (f–h) Zoomed-in single-section views of R1 and R2 in (d). Scale bars in all sub-figures: 50 µm.

Figure 2

Figure 2

Schematic illustration of the BrainAligner algorithm. (a) BrainAligner performs a global alignment (G) followed by nonlinear local alignments (L) using landmarks. Scale bars: 50 µm. (b) The Reliable Landmark Matching (RLM) algorithm for detecting corresponding feature points in subject and target images. Dots of the same color indicate the matching landmarks; PT, a target brain landmark position; PS, a subject brain landmark; PMI, PINT, PCC, the best matching positions based on mutual information (MI), voxel intensity (INT), and correlation coefficient (CC) of local image patches. In the tetrahedron-pruning step, the landmarks in a subject image that clearly violate the relative position relationships of the target are discarded.

Figure 3

Figure 3

Stereotypy of neuronal morphology and reproducibility of GAL4 expression patterns. (a) Two aligned and overlaid examples (magenta and green) of the a278-GAL4 expression pattern, from different brains. Scale bar: 20 µm. (b) 3D reconstruction of the major neurite tracts in (a). Magenta and green, surface representations of the reconstructed tracts. Gray, GAL4 pattern. Scale bar: 20 µm. (c) 3D reconstructed neurite tracts (gray) from 20 aligned a278-GAL4 images, along with their mean tract model (red). (d) Average deviation of the mean tract model from each reconstructed tract.

Figure 4

Figure 4

Expression pattern overlap by computational and biological methods. (a) Maximum intensity projection of a278-GAL4; UAS-mCD8-GFP. Scale bar: 100 µm. (b) Maximum intensity projection of LexAP036; lexop-CD2-GFP. Scale bar: 100 µm. (c) Aligned image of GAL4 and LexA expression patterns in (a) and (b), with a zoomed-in view to the right. Scale bar: 50 µm. (d) Co-expression of the GAL4 and LexA patterns, with a zoomed-in view to the right. Scale bar: 50 µm. Arrows indicate the 11 locations where colocalization of the two patterns was measured; the yellow arrow indicates a region of substantial overlap. (e–f) Cross-sectional views of single slices of the aligned (e) and co-expressed (f) samples at a position corresponding to the yellow arrow in (c) and (d). Scale bars: 25 µm.

Figure 5

Figure 5

Comparison of computational alignment of separate brains with co-expression within the same brain. For all images, grey shows N-cadherin (N-Cad) labeling, which serves as the reference signal for alignment to the nc82-labeled target. Magenta, FasII antibody staining; green, GAL4 expression pattern (anti-GFP stain). (a) Wild-type w1118 adult brain. (b–d) Expression patterns of the indicated lines shown as maximum intensity projections of 20X confocal image stacks. (e, g, i) Cross-sectional views of computational alignments of FasII expression from (a) with GAL4 patterns from (b–d). (f, h, j) Matched cross-sectional views of brains expressing the GAL4 lines and labeled with both anti-GFP and anti-FasII to show biological co-localization. OK107 and 201Y expression patterns overlap with FasII (yellow arrows), but C232 expresses in adjacent but non-overlapping brain regions (red arrow). Scale bars, 100 µm.

Figure 6

Figure 6

A 3D atlas of neurite tracts reconstructed from aligned GAL4 patterns. (a) 269 stereotyped neurite tracts and their distribution in the brain. The width of each tract equals the respective spatial deviation. The tracts are color-coded randomly for better visualization. Scale bar: 100 µm. (b) Distribution of the spatial deviation of the neurite tracts.

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