BigNeuron: a resource to benchmark and predict performance of algorithms for automated tracing of neurons in light microscopy datasets.

BigNeuron is an open community bench-testing platform with the goal of setting open standards for accurate and fast automatic neuron tracing. We gathered a diverse set of image volumes across several species that is representative of the data obtained in many neuroscience laboratories interested in neuron tracing. Here, we report generated gold standard manual annotations for a subset of the available imaging datasets and quantified tracing quality for 35 automatic tracing algorithms. The goal of generating such a hand-curated diverse dataset is to advance the development of tracing algorithms and enable generalizable benchmarking. Together with image quality features, we pooled the data in an interactive web application that enables users and developers to perform principal component analysis, t-distributed stochastic neighbor embedding, correlation and clustering, visualization of imaging and tracing data, and benchmarking of automatic tracing algorithms in user-defined data subsets. The image quality metrics explain most of the variance in the data, followed by neuromorphological features related to neuron size. We observed that diverse algorithms can provide complementary information to obtain accurate results and developed a method to iteratively combine methods and generate consensus reconstructions. The consensus trees obtained provide estimates of the neuron structure ground truth that typically outperform single algorithms in noisy datasets. However, specific algorithms may outperform the consensus tree strategy in specific imaging conditions. Finally, to aid users in predicting the most accurate automatic tracing results without manual annotations for comparison, we used support vector machine regression to predict reconstruction quality given an image volume and a set of automatic tracings.

An objective comparison of cell-tracking algorithms.

We present a combined report on the results of three editions of the Cell Tracking Challenge, an ongoing initiative aimed at promoting the development and objective evaluation of cell segmentation and tracking algorithms. With 21 participating algorithms and a data repository consisting of 13 data sets from various microscopy modalities, the challenge displays today's state-of-the-art methodology in the field. We analyzed the challenge results using performance measures for segmentation and tracking that rank all participating methods. We also analyzed the performance of all of the algorithms in terms of biological measures and practical usability. Although some methods scored high in all technical aspects, none obtained fully correct solutions. We found that methods that either take prior information into account using learning strategies or analyze cells in a global spatiotemporal video context performed better than other methods under the segmentation and tracking scenarios included in the challenge.

Automated neuron tracing using probability hypothesis density filtering.

Motivation: The functionality of neurons and their role in neuronal networks is tightly connected to the cell morphology. A fundamental problem in many neurobiological studies aiming to unravel this connection is the digital reconstruction of neuronal cell morphology from microscopic image data. Many methods have been developed for this, but they are far from perfect, and better methods are needed. Results: Here we present a new method for tracing neuron centerlines needed for full reconstruction. The method uses a fundamentally different approach than previous methods by considering neuron tracing as a Bayesian multi-object tracking problem. The problem is solved using probability hypothesis density filtering. Results of experiments on 2D and 3D fluorescence microscopy image datasets of real neurons indicate the proposed method performs comparably or even better than the state of the art. Availability and Implementation: Software implementing the proposed neuron tracing method was written in the Java programming language as a plugin for the ImageJ platform. Source code is freely available for non-commercial use at https://bitbucket.org/miroslavradojevic/phd . Contact: meijering@imagescience.org. Supplementary information: Supplementary data are available at Bioinformatics online.

Deep-Learning-Based Automated Neuron Reconstruction From 3D Microscopy Images Using Synthetic Training Images.

Digital reconstruction of neuronal structures from 3D microscopy images is critical for the quantitative investigation of brain circuits and functions. It is a challenging task that would greatly benefit from automatic neuron reconstruction methods. In this paper, we propose a novel method called SPE-DNR that combines spherical-patches extraction (SPE) and deep-learning for neuron reconstruction (DNR). Based on 2D Convolutional Neural Networks (CNNs) and the intensity distribution features extracted by SPE, it determines the tracing directions and classifies voxels into foreground or background. This way, starting from a set of seed points, it automatically traces the neurite centerlines and determines when to stop tracing. To avoid errors caused by imperfect manual reconstructions, we develop an image synthesizing scheme to generate synthetic training images with exact reconstructions. This scheme simulates 3D microscopy imaging conditions as well as structural defects, such as gaps and abrupt radii changes, to improve the visual realism of the synthetic images. To demonstrate the applicability and generalizability of SPE-DNR, we test it on 67 real 3D neuron microscopy images from three datasets. The experimental results show that the proposed SPE-DNR method is robust and competitive compared with other state-of-the-art neuron reconstruction methods.

Spherical-Patches Extraction for Deep-Learning-Based Critical Points Detection in 3D Neuron Microscopy Images.

Digital reconstruction of neuronal structures is very important to neuroscience research. Many existing reconstruction algorithms require a set of good seed points. 3D neuron critical points, including terminations, branch points and cross-over points, are good candidates for such seed points. However, a method that can simultaneously detect all types of critical points has barely been explored. In this work, we present a method to simultaneously detect all 3 types of 3D critical points in neuron microscopy images, based on a spherical-patches extraction (SPE) method and a 2D multi-stream convolutional neural network (CNN). SPE uses a set of concentric spherical surfaces centered at a given critical point candidate to extract intensity distribution features around the point. Then, a group of 2D spherical patches is generated by projecting the surfaces into 2D rectangular image patches according to the orders of the azimuth and the polar angles. Finally, a 2D multi-stream CNN, in which each stream receives one spherical patch as input, is designed to learn the intensity distribution features from those spherical patches and classify the given critical point candidate into one of four classes: termination, branch point, cross-over point or non-critical point. Experimental results confirm that the proposed method outperforms other state-of-the-art critical points detection methods. The critical points based neuron reconstruction results demonstrate the potential of the detected neuron critical points to be good seed points for neuron reconstruction. Additionally, we have established a public dataset dedicated for neuron critical points detection, which has been released along with this article.

Automated Neuron Reconstruction from 3D Fluorescence Microscopy Images Using Sequential Monte Carlo Estimation.

Microscopic images of neuronal cells provide essential structural information about the key constituents of the brain and form the basis of many neuroscientific studies. Computational analyses of the morphological properties of the captured neurons require first converting the structural information into digital tree-like reconstructions. Many dedicated computational methods and corresponding software tools have been and are continuously being developed with the aim to automate this step while achieving human-comparable reconstruction accuracy. This pursuit is hampered by the immense diversity and intricacy of neuronal morphologies as well as the often low quality and ambiguity of the images. Here we present a novel method we developed in an effort to improve the robustness of digital reconstruction against these complicating factors. The method is based on probabilistic filtering by sequential Monte Carlo estimation and uses prediction and update models designed specifically for tracing neuronal branches in microscopic image stacks. Moreover, it uses multiple probabilistic traces to arrive at a more robust, ensemble reconstruction. The proposed method was evaluated on fluorescence microscopy image stacks of single neurons and dense neuronal networks with expert manual annotations serving as the gold standard, as well as on synthetic images with known ground truth. The results indicate that our method performs well under varying experimental conditions and compares favorably to state-of-the-art alternative methods.

Automated Neuron Detection in High-Content Fluorescence Microscopy Images Using Machine Learning.

The study of neuronal morphology in relation to function, and the development of effective medicines to positively impact this relationship in patients suffering from neurodegenerative diseases, increasingly involves image-based high-content screening and analysis. The first critical step toward fully automated high-content image analyses in such studies is to detect all neuronal cells and distinguish them from possible non-neuronal cells or artifacts in the images. Here we investigate the performance of well-established machine learning techniques for this purpose. These include support vector machines, random forests, k-nearest neighbors, and generalized linear model classifiers, operating on an extensive set of image features extracted using the compound hierarchy of algorithms representing morphology, and the scale-invariant feature transform. We present experiments on a dataset of rat hippocampal neurons from our own studies to find the most suitable classifier(s) and subset(s) of features in the common practical setting where there is very limited annotated data for training. The results indicate that a random forests classifier using the right feature subset ranks best for the considered task, although its performance is not statistically significantly better than some support vector machine based classification models.

Fuzzy-Logic Based Detection and Characterization of Junctions and Terminations in Fluorescence Microscopy Images of Neurons.

Digital reconstruction of neuronal cell morphology is an important step toward understanding the functionality of neuronal networks. Neurons are tree-like structures whose description depends critically on the junctions and terminations, collectively called critical points, making the correct localization and identification of these points a crucial task in the reconstruction process. Here we present a fully automatic method for the integrated detection and characterization of both types of critical points in fluorescence microscopy images of neurons. In view of the majority of our current studies, which are based on cultured neurons, we describe and evaluate the method for application to two-dimensional (2D) images. The method relies on directional filtering and angular profile analysis to extract essential features about the main streamlines at any location in an image, and employs fuzzy logic with carefully designed rules to reason about the feature values in order to make well-informed decisions about the presence of a critical point and its type. Experiments on simulated as well as real images of neurons demonstrate the detection performance of our method. A comparison with the output of two existing neuron reconstruction methods reveals that our method achieves substantially higher detection rates and could provide beneficial information to the reconstruction process.