Abstract
Astrocytes play important roles in normal brain function and neurological diseases. In vivo two-photon excitation laser scanning microscopy has the potential to reveal rapid, dynamic structural changes in cells in a variety of physiological and pathological conditions. The type of in vivo imaging method has been shown to affect the plasticity of dendritic spines of neurons, but the optimal in vivo imaging methods of astrocytes have not been established. We compared open-skull and thinned-skull imaging methods for two-photon laser microscopy of live astrocytes in neocortex of GFAP-GFP transgenic mice. The thinned-skull method provided stable image intensity and morphological features of astrocytes in vivo over at least one week, with no evidence of astrogliosis. In contrast, the open-skull method resulted in significant changes in image intensity and induced astrogliosis. The thinned-skull method is the preferred approach for in vivo imaging of astrocytes under most conditions involving gross astrocyte modulation or causing astrogliosis.
Highlights
Astrocytes, a subtype of glial cell in the central nervous system, are responsible for maintenance of homeostasis in the brain by regulating local ion concentrations, pH, energetic state, and metabolism [1]
Morphological features of astrocytes were assessed with respect to total astrocyte size and soma size, based on area calculations from the projected Z-stacks (S1 Fig)
With the thinned-skull technique, the image intensity did not change significantly at 24 hr, 3d, or 1 wk after surgery compared with baseline, it decreased slightly at 3 d and 1 wk compared with 24 hr (Fig 1E)
Summary
Astrocytes, a subtype of glial cell in the central nervous system, are responsible for maintenance of homeostasis in the brain by regulating local ion concentrations, pH, energetic state, and metabolism [1]. Astrodegeneration, gliosis, and other structural changes in astrocytes are often identified in pathological specimens from animal models and patients with epilepsy and other neurological disorders. Novel imaging methods for studying astrocytes provide important insights into both the physiological and pathological roles of astrocytes in normal brain function and neurological diseases. In contrast to the fixed, static view provided by conventional pathological studies, in vivo two-photon excitation laser scanning microscopy (2PLSM) is an indispensable method to investigate dynamic changes in cellular and subcellular structure and function in live tissue [4,5]. Several recent studies utilizing in vivo time-lapse 2PLSM in the mouse brain demonstrate that astrocytes may undergo rapid, dynamic structural changes under a variety of PLOS ONE | DOI:10.1371/journal.pone.0170005. Several recent studies utilizing in vivo time-lapse 2PLSM in the mouse brain demonstrate that astrocytes may undergo rapid, dynamic structural changes under a variety of PLOS ONE | DOI:10.1371/journal.pone.0170005 January 20, 2017
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