Abstract
A central effort of today’s neuroscience is to study the brain’s ’wiring diagram’. The nervous system is believed to be a network of neurons interacting with each other through synaptic connection between axons and dendrites, therefore the neuronal connectivity map not only depicts the underlying anatomy, but also has important behavioral implications. Different approaches have been utilized to decipher neuronal circuits, including electron microscopy (EM) and light microscopy (LM). However, these approaches typically demand extensive sectioning and reconstruction for a brain sample. Recently, tissue clearing methods have enabled the investigation of a fully assembled biological system with greatly improved light penetration. Yet, most of these implementations, still require either genetic or exogenous contrast labeling for light microscopy. Here we demonstrate a high-speed approach, termed as Clearing Assisted Scattering Tomography (CAST), where intact brains can be imaged at optical resolution without labeling by leveraging tissue clearing and the scattering contrast of optical frequency domain imaging (OFDI).
Highlights
Another approach for deep penetration optical imaging without sectioning, tissue clearing, originated decades ago[12] has recently received renewed interest[13]
By adjusting the degree of lipid removal, we found that it was possible for Clearing Assisted Scattering Tomography (CAST) to image through very thick tissue while maintaining sufficient contrast
To introduce optical frequency domain imaging (OFDI) imaging on cleared samples, we first constructed an OFDI imaging system (Method). This platform was designed to have a nearly isotropic optical resolution volume of 10 × 10 × 9 μm[3] (X ×Y ×Z) in air to demonstrate that CAST can work efficiently at the resolution gap between conventional light microscopy (LM) (1 μm) and MRI (100 μm)
Summary
Another approach for deep penetration optical imaging without sectioning, tissue clearing, originated decades ago[12] has recently received renewed interest[13]. While deeper than confocal (tens of micron)[29] and two-photon microscopy (hundreds of micron)[30], all existing OFDI/OCT imaging are still limited to 1~2 millimeters by tissue scattering. This prevents all previous OFDI/OCT studies[31,32,33] from imaging deep into an intact brain. As we will show in this work, by adapting the OFDI apparatus and controlling photon scattering through the clearing procedure, CAST enables high resolution, cross-sectional and volumetric imaging through thick intact tissue without any contrast agent. The significant merits of this new method CAST, lie in that it provides a unique way to image deep tissue at optical resolution in a label-free manner
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