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Improved tools to study astrocytes

Abstract

Astrocytes are a type of glial cell that tile the CNS. They interact with multiple cell types, including neurons, glial cells and blood vessels, and are involved or implicated in brain disorders. Progress has been made in understanding astrocytes, but the field lacks detailed information concerning how they perform their multifarious functions, and how and when they influence the operations of the neural circuits with which they interact. One recognized bottleneck to progress has been the paucity of reliable tools with which to explore astrocytes within the adult vertebrate CNS in vivo. However, improved tools for molecular, genetic, morphological and physiological assessments have been developed recently or have been adapted from their original purposes to study neurons and are now being used to systematically document and interrogate astrocyte biology in vivo. These tools, their uses and limitations, and the insights that they afford are summarized in this Review.

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Fig. 1: Transcriptomic profiling of astrocytes from cell-specific RNA sequencing in adult mice.
Fig. 2: Methods to study astrocyte morphology.
Fig. 3: Cartoon of the tools used to manipulate astrocyte intracellular Ca2+ signalling.
Fig. 4: Causative roles of astrocyte intracellular Ca2+ signalling revealed by new tools.

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Acknowledgements

The authors are supported by the US National Institutes of Health (NS111583, DA047444, NS060677 and MH104069), a Paul G. Allen Distinguished Investigator Award and the CHDI Foundation (to B.S.K.). B.S.K. was also partly supported by the Ressler Family Foundation. X.Y. was supported partly by an American Heart Association Postdoctoral Fellowship (16POST27260256). J.N. was partly supported by a Japan Society for the Promotion of Science (JSPS) Overseas Research Fellowship (H28-729) and the Uehara Memorial Foundation Overseas Postdoctoral Research Fellowship (201730082). The authors regret that many papers could not be cited (especially early studies), because of space limits and the requirement to focus primarily on the past 5 years. The authors thank members of the Khakh laboratory for useful discussions, and the anonymous reviewers for their comments.

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All authors contributed equally to all aspects of the manuscript.

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Correspondence to Baljit S. Khakh.

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Related links

Adult Astrocyte RNA-seq Explorer: http://astrocyternaseq.org/

Allen Brain Atlas, RNA-seq data: https://celltypes.brain-map.org/rnaseq

Astrocyte Ageing Transcription: http://igc1.salk.edu:3838/astrocyte_aging_transcriptome/

Brain RNA-seq: http://www.brainrnaseq.org/

DropViz: http://dropviz.org/

Mouse Brain Atlas: http://mousebrain.org

SCope: http://scope.aertslab.org

Glossary

Gliotransmission

The release of transmitters from astrocytes owing to intracellular Ca2+ elevations, through vesicular exocytosis or by another regulated mechanism.

Immunopanning

A method of cell purification that uses cell-type-specific antibodies immobilized to a solid surface (such as a cell culture plate) to immunoprecipitate specific cell populations.

Bacterial artificial chromosome (BAC) transgenic

Describing a transgenic organism generated by random but stable integration of large segments of DNA (up to 300 kb) from BAC vectors into the genome.

Tamoxifen

A selective oestrogen receptor modulator that specifically activates tamoxifen-inducible Cre recombinase (for example, Cre/ERT2) and results in the shuttling of the Cre recombinase into the nucleus, where the recombination occurs.

Adeno-associated virus (AAV) vector

A vector based on an AAV, composed of exogenous DNA (up to 5 kb) flanked by two 145-nucleotide-long inverted terminal repeats.

Tetracycline

An antibiotic used to either repress (Tet-Off) or activate (Tet-On) gene expression in a Tet system. Its derivative, doxycycline, is also widely used.

Microglial reactivity

A microglial activation response to tissue damage or pathological insults, to mediate inflammatory responses.

Iontophoresis

An approach to deliver charged molecules (such as fluorescent dye) into cells by applying electric current.

Ezrin

A protein that functions as a linker between the plasma membrane and the actin cytoskeleton.

Lck-GFP

A green fluorescent protein (GFP) tagged with a membrane-targeting sequence of Lck (from Src tyrosine kinase).

Spaghetti monster fluorescent proteins

Modified fluorescent proteins with multiple epitope tags that can be targeted using highly specific antibodies.

Förster resonance energy transfer

(FRET). Energy transfer between two appropriate light-sensitive molecules, such as fluorophores, via dipole–dipole coupling. The transfer typically reports distances on the tens of nanometres scale or lower and so can be used to assess the proximity between cognate fluorophores that undergo Förster resonance energy transfer.

Microdomains

Highly localized, small (several micrometres wide) territories of a cell.

Photoisomerable

Able to undergo reversible structural conversion between isomers in response to photoexcitation.

Remote memory

The enduring memory of events that happened or were learnt in the distant past.

Retinal ganglion cells

Output neurons in the ganglion cell layer of the retina, receiving visual input from photoreceptors and interneurons.

Tet system

An inducible gene expression system that reversibly activates (Tet-On) or represses (Tet-Off) transcription in the presence of a tetracycline transactivator (tTA) protein, a DNA sequence called the tetracycline response element (TRE) and treatment with tetracycline or its derivatives.

Acidification

The act of something such as a cellular compartment becoming more acidic (that is, becoming protonated).

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Yu, X., Nagai, J. & Khakh, B.S. Improved tools to study astrocytes. Nat Rev Neurosci 21, 121–138 (2020). https://doi.org/10.1038/s41583-020-0264-8

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