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Microscopy Method Observes Spinal Cells in Freely Behaving Mice

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A microscope about the size of a penny has been used to observe everyday activity of cells within the spinal cords of mice, revealing that astrocytes — cells in the nervous system that do not conduct electrical signals and were traditionally viewed as merely supportive — unexpectedly react to intense sensation.

Researchers from the Salk Institute developed fluorescence imaging approaches based on two- and miniaturized one-photon microscopy. The miniaturized microscope and related imaging methods offer insight into nervous system function and could lead to novel pain treatments for spinal cord injuries, chronic itch and neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS).

Salk Institute scientists have observed the involvement of cells called astrocytes in spinal sensory processing.
Salk Institute scientists have observed the involvement of cells called astrocytes in spinal sensory processing. Here, astrocytes (genetically labeled in green) in a spinal cord (costained with glial fibrillary acidic protein, red, to visualize its outline) react to the activity of sensation with their own chemical signals. Courtesy of the Salk Institute.

In the study, published in Nature Communications (doi: 10.1038/ncomms11450) professor Axel Nimmerjahn and his team improved upon miniaturized microscopes they first described in 2008. The new version features numerous hardware and software improvements and enabled them to visualize changes in cellular activity in awake, freely behaving mice.

Most of the Salk team's previous work focused on deploying microscopes to observe the brains of living animals. The spinal cord, by contrast, presented a bigger challenge; unlike the brain, multiple, independently moving vertebrae surround the spinal cord. The spinal cord is also closer to pulsating organs such as the heart and lungs, which can hinder stable views of the cells within. By developing new microscopy and procedural and computational approaches, the team was able to capture the action of living cells in real time and during vigorous movements.

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The researchers reported that distinct stimuli such as light touch or pressure activated different subsets of spinal sensory neurons. Certain features, like the intensity or duration of a given stimulus, were reflected in the activity of the neurons.

Salk researchers (from left, Kohei Sekiguchi and Axel Nimmerjahn) have revealed what they say is the world’s first imaging data on spinal cellular activity during behavior, enabled by miniaturized microscopes.
Salk researchers (from left, Kohei Sekiguchi and Axel Nimmerjahn) have revealed what they say is the world’s first imaging data on spinal cellular activity during behavior, enabled by miniaturized microscopes. Courtesy of the Salk Institute.

To the team's surprise, astrocytes, traditionally thought to be passive support cells, also responded to stimuli, albeit differently than the neurons. Though the astrocytes cannot send electrical signals like neurons can, they generated their own chemical signals in a coordinated way during intense stimuli.

Astrocytes are increasingly appreciated as important players in how the nervous system develops and operates and could serve as promising new drug targets, Nimmerjahn said. The microscopy protocols will enable study of normal sensory processing, but also disease contexts and the effect of treatment on cells.

The team is now working to simultaneously record touch or pain-related activity in the brain and spinal cord using additional iterations of the miniaturized microscopes, which allow them to monitor and manipulate multiple cell types at even higher resolutions.

The video below describes how the imaging technology offers a new view into the spinal cord. Courtesy of the  Salk Institute.


Published: May 2016
Glossary
fluorescence
Fluorescence is a type of luminescence, which is the emission of light by a substance that has absorbed light or other electromagnetic radiation. Specifically, fluorescence involves the absorption of light at one wavelength and the subsequent re-emission of light at a longer wavelength. The emitted light occurs almost instantaneously and ceases when the excitation light source is removed. Key characteristics of fluorescence include: Excitation and emission wavelengths: Fluorescent materials...
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