SynapCountJ: A Validated Tool for Analyzing Synaptic Densities in Neurons

Abstract

The quantification of synapses is instrumental to measure the evolution of synaptic densities of neurons under the effect of some physiological conditions, neuronal diseases or even drug treatments. However, the manual quantification of synapses is a tedious, error-prone, time-consuming and subjective task; therefore, tools that might automate this process are desirable. In this paper, we present SynapCountJ, an ImageJ plugin, that can measure synaptic density of individual neurons obtained by immunofluorescence techniques, and also can be applied for batch processing of neurons that have been obtained in the same experiment or using the same setting. The procedure to quantify synapses implemented in SynapCountJ is based on the colocalization of three images of the same neuron (the neuron marked with two antibody markers and the structure of the neuron) and is inspired by methods coming from Computational Algebraic Topology. SynapCountJ provides a procedure to semi-automatically quantify the number of synapses of neuron cultures; as a result, the time required for such an analysis is greatly reduced.

Neuronal Autophagy Regulates Presynaptic Neurotransmission by Controlling the Axonal Endoplasmic Reticulum

“…Average intensities of fluorescent puncta (synapses) were measured by centering 9 × 9 pixel (~1 × 1 μm) regions on maxima determined by ImageJ processing function. SynapCountJ, an ImageJ plugin, was utilized to determine synapses via colocalization of Homer and vGLUT in traced neurites, as described previously ( Mata et al., 2016 ). For quantifying ER antibody stainings in neurites MAP2 and Tau signal were used as template for a mask, restricting the quantified area to the shape of the dendrites or axons.…”

Taurine Promotes Neurite Outgrowth and Synapse Development of Both Vertebrate and Invertebrate Central Neurons

“…It, therefore, remains unknown whether taurine affects the level of clustering of synaptic proteins at the single neuron-, neurite-, or network-levels. We took advantage of well-established methods such as SynQuant (Wang et al, 2020 ) and SynapCountJ (Mata et al, 2016 ) to analyze different synaptic parameters in entire images (networks) and primary neurites. We found that taurine not only regulated synaptic protein expression in networks but also their intensity in synaptic puncta along neurites.…”

“…Synaptophysin and PSD95 fluorescent intensity were measured in ImageJ as described for β-tubulin above. The ImageJ plugin SynapCountJ was used to measure the number of synapses (colocalization of synaptophysin and PSD95) per area of traced neurite as previously described (Mata et al, 2016 ). Synaptic puncta parameters were measured with the ImageJ plugin SynQuant (Wang et al, 2020 ).…”

Astrocytic and Neuronal Apolipoprotein E Isoforms Differentially Affect Neuronal Excitability

“…VGAT- and vGlut1-positive synaptic density was quantified per neuron using ImageJ ( Figures 5E,F ). The tracings of neurites were selected based on phalloidin staining using ImageJ plugin NeuroJ ( Meijering et al, 2004 ) and synaptic density per 100 μm neurite length was determined based on VGAT-phalloidin and vGlut1-phalloidin combinations using the SynapCount v2 plugin ( Mata et al, 2017 ). A total of five neurons for each embryo were analyzed.…”