FRET based ratiometric Ca(2+) imaging to investigate immune-mediated neuronal and axonal damage processes in experimental autoimmune encephalomyelitis

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Item Type:	Article
Title:	FRET based ratiometric Ca(2+) imaging to investigate immune-mediated neuronal and axonal damage processes in experimental autoimmune encephalomyelitis
Creators Name:	Siffrin, V., Birkenstock, J., Luchtmann, D.W., Gollan, R., Baumgart, J., Niesner, R.A., Griesbeck, O. and Zipp, F.
Abstract:	BACKGROUND: Irreversible axonal and neuronal damage are the correlate of disability in patients suffering from multiple sclerosis (MS). A sustained increase of cytoplasmic free [Ca2+] is a common upstream event of many neuronal and axonal damage processes and could represent an early and potentially reversible step. NEW METHOD: We propose a method to specifically analyze the neurodegenerative aspects of experimental autoimmune encephalomyelitis by Forster Resonance Energy Transfer (FRET) imaging of neuronal and axonal Ca2+ dynamics by two-photon laser scanning microscopy (TPLSM). RESULTS: Using the genetically encoded Ca2+ sensor TN-XXL expressed in neurons and their corresponding axons, we confirm the increase of cytoplasmic free [Ca2+] in axons and neurons of autoimmune inflammatory lesions compared to those in non-inflamed brains. We show that these relative [Ca2+] increases were associated with immune-neuronal interactions. COMPARISON WITH EXISTING METHODS: In contrast to Ca2+-sensitive dyes the use of a genetically encoded Ca2+ sensor allows reliable intraaxonal free [Ca2+] measurements in living anesthetized mice in health and disease. This method detects early axonal damage processes in contrast to e.g. cell/axon morphology analysis, that rather detects late signs of neurodegeneration. CONCLUSIONS: Thus, we describe a method to analyze and monitor early neuronal damage processes in the brain in vivo.
Keywords:	EAE/MS, Two-Photon Laser Scanning Microscopy, Intravital Microscopy, FRET, Ca2+ Imaging Neurodegeneration, Animals, Mice
Source:	Journal of Neuroscience Methods
ISSN:	0165-0270
Publisher:	Elsevier
Volume:	249
Page Range:	8-15
Date:	15 July 2015
Official Publication:	https://doi.org/10.1016/j.jneumeth.2015.04.005
PubMed:	View item in PubMed

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