NeurphologyJ: An automatic neuronal morphology quantification method and its application in pharmacological discovery

Abstract

BackgroundAutomatic quantification of neuronal morphology from images of fluorescence microscopy plays an increasingly important role in high-content screenings. However, there exist very few freeware tools and methods which provide automatic neuronal morphology quantification for pharmacological discovery.ResultsThis study proposes an effective quantification method, called NeurphologyJ, capable of automatically quantifying neuronal morphologies such as soma number and size, neurite length, and neurite branching complexity (which is highly related to the numbers of attachment points and ending points). NeurphologyJ is implemented as a plugin to ImageJ, an open-source Java-based image processing and analysis platform. The high performance of NeurphologyJ arises mainly from an elegant image enhancement method. Consequently, some morphology operations of image processing can be efficiently applied. We evaluated NeurphologyJ by comparing it with both the computer-aided manual tracing method NeuronJ and an existing ImageJ-based plugin method NeuriteTracer. Our results reveal that NeurphologyJ is comparable to NeuronJ, that the coefficient correlation between the estimated neurite lengths is as high as 0.992. NeurphologyJ can accurately measure neurite length, soma number, neurite attachment points, and neurite ending points from a single image. Furthermore, the quantification result of nocodazole perturbation is consistent with its known inhibitory effect on neurite outgrowth. We were also able to calculate the IC50 of nocodazole using NeurphologyJ. This reveals that NeurphologyJ is effective enough to be utilized in applications of pharmacological discoveries.ConclusionsThis study proposes an automatic and fast neuronal quantification method NeurphologyJ. The ImageJ plugin with supports of batch processing is easily customized for dealing with high-content screening applications. The source codes of NeurphologyJ (interactive and high-throughput versions) and the images used for testing are freely available (see Availability).

The Unfolded Protein Response and Cholesterol Biosynthesis Link Luman/CREB3 to Regenerative Axon Growth in Sensory Neurons

“…Total axon length/neuron (identified by ␤III-tubulin immunofluorescence) was calculated for all neurons in each of ϳ50 random fields per experimental treatment group per replicate (N ϭ 3) using NeurphologyJ (Ho et al, 2011) 48 h after plating for each experimental condition examined.…”

Image Informatics Strategies for Deciphering Neuronal Network Connectivity

“…All global segmentation methods rely on binarization (i.e. thresholding) and skeletonization of a pre-processed image ( Figure 2B) 62 . The complexity of the pre-processing steps (apart from those mentioned in 2.1) is what truly discriminates different methods, and this is usually based on the image quality and density of the cell culture.…”

Ccm3, a gene associated with cerebral cavernous malformations, is required for neuronal migration

“…Emx1/Ccm3 cKO, Cdk5 and p35 mutants (this study) (Ohshima et al, 1996;Chae et al, 1997;Gilmore et al, 1998;Kwon and Tsai, 1998;Ohshima et al, 2007;Hoerder-Suabedissen et al, 2009) have nearly identical cortical lamination defects and strikingly similar Ctgf, Er81 and Cux2 profiles (Fig. 1D,H,L) (Ohshima et al, 2007;Hoerder-Suabedissen et al, 2009) raising the possibility of CCM3 interaction with the Cdk5/p35 pathway, which regulates actin and microtubule dynamics during neuronal migration, as well as dendritic development of pyramidal neurons (Xie et al, 2003;Kawauchi et al, 2006;Ohshima et al, 2007), processes also disrupted in the Ccm3 cKO mutants.…”

“…1D,H,L) (Ohshima et al, 2007;Hoerder-Suabedissen et al, 2009) raising the possibility of CCM3 interaction with the Cdk5/p35 pathway, which regulates actin and microtubule dynamics during neuronal migration, as well as dendritic development of pyramidal neurons (Xie et al, 2003;Kawauchi et al, 2006;Ohshima et al, 2007), processes also disrupted in the Ccm3 cKO mutants. Cdk5 modulates actin reorganization via RhoA suppression and regulation of cofilin activity (Kawauchi et al, 2006), and controls microtubule organization via FAK phosphorylation (Xie et al, 2003).…”