Methods of dendritic spine detection: From Golgi to high-resolution optical imaging

Abstract

Dendritic spines, the bulbous protrusions that form the postsynaptic half of excitatory synapses, are one of the most prominent features of neurons and have been imaged and studied for over a century. In that time, changes in the number and morphology of dendritic spines have been correlated to the developmental process as well as the pathophysiology of a number of neurodegenerative diseases. Due to the sheer scale of synaptic connectivity in the brain, work to date has merely scratched the surface in the study of normal spine function and pathology. This review will highlight traditional approaches to the imaging of dendritic spines and newer approaches made possible by advances in microscopy, protein engineering, and image analysis. The review will also describe recent work that is leading researchers toward the possibility of a systematic and comprehensive study of spine anatomy throughout the brain.

A close look at brain dynamics: Cells and vessels seen by in vivo two-photon microscopy

“…While generally considered to range between 800 mm and 1 mm, maximum depth may also be lower depending on the object to visualize. The very small structures such as dendritic spines or thin microglial ramifications, that are sized close to the microscope resolution limit, require high resolution to be properly quantified (Mancuso et al, 2013). Work dealing with fine measures of dendritic spines or microglia morphology is usually done within a range of 200-250 mm beneath the outer cerebral surface, where proper resolution can be achieved (Chaigneau et al, 2011;Herz et al, 2010).…”

Merging advanced technologies with classical methods to uncover dendritic spine dynamics: A hot spot of synaptic plasticity

“…Both neurons and spines can be visualized with fluorescent microscopes using this labeling method. Recently, scientists have used plasmid-encoded soluble enhanced GFP (eGFP), membrane-tagged eGFP, and mRFP-ruby tagged Life act, a small actin-binding peptide, to label neurons (Mancuso et al, 2013). Similarly, neuron-specific promoters (e.g.…”

“…1). An analysis of the ingredients used in the Golgi stain suggests that the potassium dichromate impregnates the neurons and glia and hardens them, whereas the silver nitrate turns the neurons black Mallick, 2010, 2012;Mancuso et al, 2013). It is not known why some neurons are stained and others are not; in fact the staining mechanism remains enigmatic today, well over hundred years after its introduction.…”

“…Using the appropriate software it can also be used to reconstruct image stacks to show high-resolution 3-D structure, thus providing information about hidden spines in the z-direction (Castano et al, 1995). The development of new fluorescent dyes, immuno-labeling techniques, protein engineering tools, and genetically engineered animals allowed for major advances in the study of the functional aspects of dendritic spines using CLSM (Mancuso et al, 2013;Sala and Segal, 2014). More recently the introduction of calcium-sensitive fluorescent dyes (e.g.…”

Bioimage Informatics for Big Data

“…In addition, the spatial relationship of neuron(s) and synapses should be characterized. Achieving these complex computational analyses is often technically challenging (Micheva et al 2010;Kim et al 2012;Mancuso et al 2013;Collman et al 2015).…”