Long-term high-resolution in vivo imaging of cerebral cortical structures following ischemic stroke
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Biophysics Reports
P R OTO C O L
Long-term high-resolution in vivo imaging of cerebral cortical structures following ischemic stroke Lei Wang1, Lirui Zhu1, Junru Liu1, Shengxiang Zhang1& 1
Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, School of Life Sciences, Lanzhou University, Lanzhou 730000, China
Received: 3 June 2019 / Accepted: 14 January 2020
Abstract
The development of multi-photon microscopic technique has made it possible to image submicron structures deep in biological tissues. This technique is widely used for imaging of cortical structures in developing and adult animals, and there have been detail descriptions of in vivo imaging of synaptic structures in normal animals through a thinned-skull or open-skull cranial window. However, several challenges should be considered carefully for high-resolution imaging of cortical structures under pathological conditions. Here we describe a protocol for in vivo imaging of dendritic structures following ischemic stroke through thinned skull. This protocol can also be applied for acute or chronic imaging of neuronal structural plasticity, glial activation, cerebral microcirculation, or synaptic functions in other pathological conditions.
Keywords Two-photon microscopy, In vivo imaging, Ischemic stroke, Spine
INTRODUCTION The invention of two-photon fluorescence microscopy enables direct visualization of submicron fluorescent structures deep in biological tissues with two-photon excitation (Denk et al. 1990). Excitation of fluorescent molecules based on simultaneous absorption of two photons of relatively longer wavelength has improved the imaging depth greatly. The application of twophoton microscopy to study synaptic plasticity in living animals has been facilitated with the emergence of transgenic mice expressing fluorescent proteins (Feng et al. 2000; Grutzendler et al. 2002; Trachtenberg et al. 2002). In the past decades, live imaging technique based on multi-photon excitation has been found widely applied in time-lapse observation of structural and functional changes of the nervous system during development and in a variety of disease models such as stroke, Alzheimer’s disease, Parkinson’s disease and spinal cord injury (Davalos et al. 2008; Eichhoff and & Correspondence: [email protected] (S. Zhang)
Ó The Author(s) 2020
Garaschuk 2011; Guo et al. 2015; Li and Murphy 2008; Reichenbach et al. 2018; Zhang et al. 2005; Zuo et al. 2005). There have been detailed descriptions of the procedures for in vivo imaging of cortical structures in normal animals through thinned-skull or open-skull window (Grutzendler et al. 2011; Holtmaat et al. 2009; Yang et al. 2010; Zuo et al. 2013). However, these methods mainly focus on intravital microscopy of cortical structures in healthy animals, and pathological changes that may appear in disease models such as extravasation (leakage of blood plasma), inflammation, deformations of vascular structures are not emphasized. These pathological changes may affect refractive index of cortical
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