Table1.XLS

<p>Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.</p

From a resin block to serial 2d image montages.

<p><b>A</b> Serial EM is performed on a block of tissue embedded in hardened plastic resin. <b>B</b> Sections are imaged with multiple overlapping image tiles. <b>C</b> The imprecision in the positioning of the camera and the numerous non-linear deformations demand of an automatic multi-section image registration procedure that computes the best possible transformation for each tile without introducing gross deformations. <b>D</b> TrakEM2 operates only on original images, which are treated as read-only. A <i>preprocessor script</i> specified invidually for every image alters the image after loading from disk and before the rest of TrakEM2 has access to it, enabling changes of scale, of look-up table, data type, and any pixel-level operation. A <i>Patch</i> object encapsulates the image file path and a set of properties such as the alpha mask, the coordinate transforms (linear and non-linear image transformations) and the desired image display range and composite mode, among others. The precomputed mipmaps store most of the <i>Patch</i> information in compressed 8-bit files ready for display. The image for the field of view is constructed from composing multiple <i>Patch</i> instances according to their location and composite rules (overlay, subtract, add, multiply, difference and Colorize YCbCr), and is then filtered, if desired, for dynamic interactive image enhancement. <b>E</b> The TrakEM2 <i>Display</i> presents the field of view showing a single section and the images, segmentations and annotations present in that section. The <i>Display</i> provides access to tools for manipulating and analyzing all imported images and reconstructed elements.</p

Neural circuit reconstruction with skeletonized neural arbors and connectors to relate them at synaptic sites.

<p><b>A</b> Snapshot illustrating the use of connectors to relate neural arbors. The connector in green (notice the ‘o’ node with a yellow circle around; it has three targets–it’s a polyadic insect synapse), each of which is represented within the section by a node with an arrow head that falls within the circle of each target. To the left, notice the use of text annotations to describe the synapse. <b>B</b> Search with regular expressions locates any objects of interest, in this case a “membrane specializations” tag in a neuronal arbor. <b>C</b> The tabular view for a neural arbor lists all nodes, branch nodes, end nodes or a subset whose tags match a regular expression. All columns are sortable, and clicking on each row positions the display on the node. The last column titled “Reviews” indicates which cables of the neuron have already been reviewed (in green) to correct for missing branches or synapses or other issues. <b>D</b> A review stack is precomputed for fast visualization of the cable of interest, each section centered on the node. The visual flow through the stack helps in catching reconstruction errors. <b>E</b> “Area trees” are skeleton arbors whose nodes have 2d areas associated. <b>F</b> 3d rendering of two “area trees”, a section of which are depicted in E. <b>G</b> 3d rendering of the nucleus (represented by a “ball”) and the arbor (represented by a “treeline”) of a neuron in the insect brain. <b>H–J</b> Cartons of the skeletons used for reconstruction. The root node is labeled with an “S”, the branch nodes with “Y” and the end nodes with “e”. In H, a “connector” relates the nodes of two arbors, with specific confidence value for the relationship. These confidence values exist on the edges that relate the arbor’s nodes as well (not shown). <b>I</b> Rerooting changes the perspective, but not the topology, of the tree. By convention we position the root node at the soma. <b>J</b> Two common and trivial operations on trees are split and merge.</p

Image1.PDF

<p>Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.</p

Image2.PDF

<p>Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.</p

Video1.mp4

<p>Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.</p