Abstract
In brain networks, neurons are constantly involved in a dynamic interaction with the other cell populations and, particularly, with astrocytes, the most abundant glial cells in the brain. Astrocytes respond to neurotransmitters with Ca2+ elevations which represent a key event in the modulation of local brain circuits played by these glial cells. Due to technical limitations, the study of Ca2+ signal dynamics in astrocytes has focused for decades almost exclusively on somatic and perisomatic regions. Accordingly, Ca2+ signal in astrocytic fine protrusions, which are in close contact with the synapse, has been poorly investigated. Over the last years, the diffusion of novel tools such as the viral vector gene delivery of genetically encoded Ca2+ indicators (GECI), the optogenetics, and multiphoton laser scanning microscopy has boosted significantly our capability to study astrocytic Ca2+ signals in the different subcellular compartments. Here we report a protocol that combines these techniques to study astrocyte Ca2+ signaling in response to somatostatin (SST)-expressing interneurons, one of the main classes of GABAergic inhibitory interneurons.
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Change history
08 June 2019
Figures 1 and 2 were inadvertently switched during the production and this has been corrected so the figures appear in the proper order.
References
Araque A et al (2014) Gliotransmitters travel in time and space. Neuron 81:728–739
Bazargani N, Attwell D (2016) Astrocyte calcium signaling: the third wave. Nat Neurosci 19:182–189
Pekny M, Pekna M, Messing A, Steinhäuser C, Lee JM, Parpura V, Hol EM, Sofroniew MV, Verkhratsky A (2016) Astrocytes: a central element in neurological diseases. Acta Neuropathol 131(3):323–345
Verkhratsky A, Zorec R, Parpura V (2017) Stratification of astrocytes in healthy and diseased brain. Brain Pathol 27(5):629–644
Tong X, Shigetomi E, Looger LL, Khakh BS (2013) Genetically encoded calcium indicators and astrocyte calcium microdomains. Neuroscientist 19(3):274–291
Rusakov DA (2015) Disentangling calcium-driven astrocyte physiology. Nat Rev Neurosci 16(4):226–233
Losi G, Mariotti L, Sessolo M, Carmignoto G (2017) New tools to study astrocyte Ca2+ signal dynamics in brain networks in vivo. Front Cell Neurosci 11:134
Deisseroth K (2015) Optogenetics: 10 years of microbial opsins in neuroscience. Nat Neurosci 18(9):1213–1225
Rost BR, Schneider-Warme F, Schmitz D, Hegemann P (2017) Optogenetic tools for subcellular applications in neuroscience. Neuron 96(3):572–603
Mariotti L, Losi G, Lia A, Melone M, Chiavegato A, Gómez-Gonzalo M, Sessolo M, Bovetti S, Forli A, Zonta M, Requie LM, Marcon I, Pugliese A, Viollet C, Bettler B, Fellin T, Conti F, Carmignoto G (2018) Interneuron-specific signaling evokes distinctive somatostatin-mediated responses in adult cortical astrocytes. Nat Commun 9(1):82
McCown TJ (2011) Adeno-associated virus (AAV) vectors in the CNS. Curr Gene Ther 11(3):181
Watakabe A, Ohtsuka M, Kinoshita M, Takaji M, Isa K, Mizukami H, Ozawa K, Isa T, Yamamori T (2015) Comparative analyses of adeno-associated viral vector serotypes 1, 2, 5, 8 and 9 in marmoset, mouse and macaque cerebral cortex. Neurosci Res 93:144–157
Acknowledgments
We are grateful to Serena Bovetti and Tommaso Fellin for excellent initial support on viral vector injection and optogenetic stimulation and to Vanessa Henriques for mouse preparation of slices shown in Fig. 3b, d.
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Losi, G., Lia, A.M., Gomez-Gonzalo, M., Zonta, M., Carmignoto, G. (2019). Optogenetic Interneuron Stimulation and Calcium Imaging in Astrocytes. In: Raffaello, A., Vecellio Reane, D. (eds) Calcium Signalling. Methods in Molecular Biology, vol 1925. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9018-4_16
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DOI: https://doi.org/10.1007/978-1-4939-9018-4_16
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