Abstract
Time course, in vivo imaging of brain cells is crucial to fully understand the progression of secondary cellular damage and recovery in murine models of injury. We have combined high-resolution gradient index lens technology with a model of diffuse axonal injury in rodents to enable repeated visualization of fine features of individual cells in three-dimensional space over several weeks. For example, we recorded changes in morphology in the same axons in the external capsule numerous times over 30 to 60 days, before and after induced traumatic brain injury. We observed the expansion of secondary injury and limited recovery of individual axons in this subcortical white matter tract over time. In another application, changes in microglial activation state were visualized in the penumbra region of mice before and after ischemia induced by middle carotid artery occlusion. The ability to collect a series of high-resolution images of cellular features of the same cells pre- and post-injury enables a unique opportunity to study the progression of damage, spontaneous healing, and effects of therapeutics in mouse models of neurodegenerative disease and brain injury.
Highlights
Multiphoton microscopy together with promoter-directed expression of fluorescent proteins for cell-type specific labeling has propelled in vivo neuroscience research into previously inaccessible regions of the animal brain
We present the design and use of an implanted small-diameter, high-resolution gradient index lens system for longitudinal studies that is optimized to image changes in fine cellular features associated with secondary injury in murine models of traumatic brain injury (TBI) and ischemic stroke
We describe how to integrate this system with murine models of brain injury, including midline fluid percussion injury, a model of diffuse brain injury[16], and middle carotid artery occlusion, a model of ischemic stroke[17]
Summary
Multiphoton microscopy together with promoter-directed expression of fluorescent proteins for cell-type specific labeling has propelled in vivo neuroscience research into previously inaccessible regions of the animal brain. We present the design and use of an implanted small-diameter, high-resolution gradient index (hrGRIN) lens system for longitudinal studies that is optimized to image changes in fine cellular features associated with secondary injury in murine models of TBI and ischemic stroke This chronically-implanted lens system, for use with multiphoton microscopy, has the temporal resolution to study dynamic events ranging from seconds to months. Using this system, we were able to observe mobile microglia in the penumbra region a day after ischemic stroke and the progression of secondary axonal damage over several weeks after TBI. Given the ability to monitor the same cells over multiple time points, this system will be useful for studying the effects of potential therapeutics on arresting neurodegeneration and promoting recovery
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