AxonEM Dataset: 3D Axon Instance Segmentation of Brain Cortical Regions

Electron microscopy (EM) enables the reconstruction of neural circuits at the level of individual synapses, which has been transformative for scientific discoveries. However, due to the complex morphology, an accurate reconstruction of cortical axons has become a major challenge. Worse still, there is no publicly available large-scale EM dataset from the cortex that provides dense ground truth segmentation for axons, making it difficult to develop and evaluate large-scale axon reconstruction methods. To address this, we introduce the AxonEM dataset, which consists of two 30x30x30 um^3 EM image volumes from the human and mouse cortex, respectively. We thoroughly proofread over 18,000 axon instances to provide dense 3D axon instance segmentation, enabling large-scale evaluation of axon reconstruction methods. In addition, we densely annotate nine ground truth subvolumes for training, per each data volume. With this, we reproduce two published state-of-the-art methods and provide their evaluation results as a baseline. We publicly release our code and data at https://connectomics-bazaar.github.io/proj/AxonEM/index.html to foster the development of advanced methods.

RhoanaNet Pipeline: Dense Automatic Neural Annotation

Reconstructing a synaptic wiring diagram, or connectome, from electron microscopy (EM) images of brain tissue currently requires many hours of manual annotation or proofreading (Kasthuri and Lichtman, 2010; Lichtman and Sanes, 2008; Seung, 2009). The desire to reconstruct ever larger and more complex networks has pushed the collection of ever larger EM datasets. A cubic millimeter of raw imaging data would take up 1 PB of storage and present an annotation project that would be impractical without relying heavily on automatic segmentation methods. The RhoanaNet image processing pipeline was developed to automatically segment large volumes of EM data and ease the burden of manual proofreading and annotation. Based on (Kaynig et al., 2015), we updated every stage of the software pipeline to provide better throughput performance and higher quality segmentation results. We used state of the art deep learning techniques to generate improved membrane probability maps, and Gala (Nunez-Iglesias et al., 2014) was used to agglomerate 2D segments into 3D objects. We applied the RhoanaNet pipeline to four densely annotated EM datasets, two from mouse cortex, one from cerebellum and one from mouse lateral geniculate nucleus (LGN). All training and test data is made available for benchmark comparisons. The best segmentation results obtained gave $V^\text{Info}_\text{F-score}$ scores of 0.9054 and 09182 for the cortex datasets, 0.9438 for LGN, and 0.9150 for Cerebellum. The RhoanaNet pipeline is open source software. All source code, training data, test data, and annotations for all four benchmark datasets are available at www.rhoana.org.

Quantifying mesoscale neuroanatomy using X-ray microtomography

Methods for resolving the 3D microstructure of the brain typically start by thinly slicing and staining the brain, and then imaging each individual section with visible light photons or electrons. In contrast, X-rays can be used to image thick samples, providing a rapid approach for producing large 3D brain maps without sectioning. Here we demonstrate the use of synchrotron X-ray microtomography ($\mu$CT) for producing mesoscale $(1~\mu m^3)$ resolution brain maps from millimeter-scale volumes of mouse brain. We introduce a pipeline for $\mu$CT-based brain mapping that combines methods for sample preparation, imaging, automated segmentation of image volumes into cells and blood vessels, and statistical analysis of the resulting brain structures. Our results demonstrate that X-ray tomography promises rapid quantification of large brain volumes, complementing other brain mapping and connectomics efforts.

Automatic Annotation of Axoplasmic Reticula in Pursuit of Connectomes using High-Resolution Neural EM Data

Accurately estimating the wiring diagram of a brain, known as a connectome, at an ultrastructure level is an open research problem. Specifically, precisely tracking neural processes is difficult, especially across many image slices. Here, we propose a novel method to automatically identify and annotate small subcellular structures present in axons, known as axoplasmic reticula, through a 3D volume of high-resolution neural electron microscopy data. Our method produces high precision annotations, which can help improve automatic segmentation by using our results as seeds for segmentation, and as cues to aid segment merging.

Automatic Annotation of Axoplasmic Reticula in Pursuit of Connectomes

In this paper, we present a new pipeline which automatically identifies and annotates axoplasmic reticula, which are small subcellular structures present only in axons. We run our algorithm on the Kasthuri11 dataset, which was color corrected using gradient-domain techniques to adjust contrast. We use a bilateral filter to smooth out the noise in this data while preserving edges, which highlights axoplasmic reticula. These axoplasmic reticula are then annotated using a morphological region growing algorithm. Additionally, we perform Laplacian sharpening on the bilaterally filtered data to enhance edges, and repeat the morphological region growing algorithm to annotate more axoplasmic reticula. We track our annotations through the slices to improve precision, and to create long objects to aid in segment merging. This method annotates axoplasmic reticula with high precision. Our algorithm can easily be adapted to annotate axoplasmic reticula in different sets of brain data by changing a few thresholds. The contribution of this work is the introduction of a straightforward and robust pipeline which annotates axoplasmic reticula with high precision, contributing towards advancements in automatic feature annotations in neural EM data.

Large-Scale Automatic Reconstruction of Neuronal Processes from Electron Microscopy Images

Automated sample preparation and electron microscopy enables acquisition of very large image data sets. These technical advances are of special importance to the field of neuroanatomy, as 3D reconstructions of neuronal processes at the nm scale can provide new insight into the fine grained structure of the brain. Segmentation of large-scale electron microscopy data is the main bottleneck in the analysis of these data sets. In this paper we present a pipeline that provides state-of-the art reconstruction performance while scaling to data sets in the GB-TB range. First, we train a random forest classifier on interactive sparse user annotations. The classifier output is combined with an anisotropic smoothing prior in a Conditional Random Field framework to generate multiple segmentation hypotheses per image. These segmentations are then combined into geometrically consistent 3D objects by segmentation fusion. We provide qualitative and quantitative evaluation of the automatic segmentation and demonstrate large-scale 3D reconstructions of neuronal processes from a $\mathbf{27,000}$ $\mathbf{\mu m^3}$ volume of brain tissue over a cube of $\mathbf{30 \; \mu m}$ in each dimension corresponding to 1000 consecutive image sections. We also introduce Mojo, a proofreading tool including semi-automated correction of merge errors based on sparse user scribbles.