Abstract
BackgroundThe Golgi–Cox stain is an established method for characterising neuron cell morphology. The method highlights neurite processes of stained cells allowing the complexity of dendritic branching to be measured. New methodsConventional rapid Golgi and Golgi–Cox methods all require fresh impregnation in unfixed brain blocks. Here, we describe a modified method that gives high quality staining on brain tissue blocks perfusion-fixed with 4% paraformaldehyde (PFA) and post-fixed by immersion for 24h. ResultsTissue perfused with 4% PFA and post fixed for 24h remained viable for the modified Golgi–Cox silver impregnation staining of mouse striatum from perfused wild type and zQ175. It was not found necessary to impregnate tissue blocks with Golgi solutions prior to sectioning, as post-sectioned tissues yielded equally good impregnation. Impregnation for 14 days resulted in optimal visualisation of striatal neuron and dendritic morphology. Although no modifications applied to the rapid Golgi method were reliable, the modified Golgi–Cox method yielded consistently reliable high-quality staining. Comparison with existing methodsThe current method used fixed tissues to reduce damage and preserve cell morphology. The revised method was found to be fast, reliable and cost effective without the need for expensive staining kits and could be performed in any neuroscience lab with limited specialist equipment. ConclusionsThe present study introduces a robust reproducible and inexpensive staining method for identifying neuronal morphological changes in the post fixed mouse brain, and is suitable for assessing changes in cell morphology in models of neurodegeneration and in response to experimental treatment.
Highlights
Camillo Golgi discovered a basic method for visualising neurons in the nervous system in 1873, which was originally named the black reaction
We describe a modified method that gives high quality staining on brain tissue blocks perfusion-fixed with 4% paraformaldehyde (PFA) and post-fixed by immersion for 24 h
Tissue perfused with 4% paraformaldehyde in 0.1 M PBS (4% PFA) and post fixed for 24 h remained viable for the modified Golgi–Cox silver impregnation staining of mouse striatum from perfused wild type and zQ175
Summary
Camillo Golgi discovered a basic method for visualising neurons in the nervous system in 1873, which was originally named the black reaction (la reazione nera). The Golgi technique stains only 3–5% of cells in the brain, at random and by unidentified mechanism (Spacek, 1989). Golgi revealed that his procedure of nervous tissue hardening took place after incubation in potassium dichromate, followed by impregnation with silver nitrate (Golgi, 1873). Bayram-Weston et al / Journal of Neuroscience Methods 265 (2016) 81–88 allowing visualisation of neurons in their entirety including cell soma, axons, dendrites, and spines This allows researchers to examine, characterise and quantify axonal and dendritic morphology and spines throughout the nervous system (Risher et al, 2014). Results: Tissue perfused with 4% PFA and post fixed for 24 h remained viable for the modified Golgi–Cox silver impregnation staining of mouse striatum from perfused wild type and zQ175. Conclusions: The present study introduces a robust reproducible and inexpensive staining method for identifying neuronal morphological changes in the post fixed mouse brain, and is suitable for assessing changes in cell morphology in models of neurodegeneration and in response to experimental treatment
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