Through-skull brain imaging in vivo at visible wavelengths via dimensionality reduction adaptive-optical microscopy

Abstract

Compensation of sample-induced optical aberrations is crucial for visualizing microscopic structures deep within biological tissues. However, strong multiple scattering poses a fundamental limitation for identifying and correcting the tissue-induced aberrations. Here, we introduce a label-free deep-tissue imaging technique termed dimensionality reduction adaptive-optical microscopy (DReAM) to selectively attenuate multiple scattering. We established a theoretical framework in which dimensionality reduction of a time-gated reflection matrix can attenuate uncorrelated multiple scattering while retaining a single-scattering signal with a strong wave correlation, irrespective of sample-induced aberrations. We performed mouse brain imaging in vivo through the intact skull with the probe beam at visible wavelengths. Despite the strong scattering and aberrations, DReAM offered a 17-fold enhancement of single scattering–to–multiple scattering ratio and provided high-contrast images of neural fibers in the brain cortex with the diffraction-limited spatial resolution of 412 nanometers and a 33-fold enhanced Strehl ratio.

Phase contrast reflectance confocal brain imaging at 1650 nm

“…The signal collected provided vascular images only at superficial depths (under 200 μm), hence restricting the capabilities of the imaging setup to perform through skull longitudinal observation. It is clear that the deterioration of the resolution arises from the passage through the skull, which could be avoided with different techniques, such as skull optical clearing 18 or adaptive imaging via frontwave manipulation 19 . However, optical clearing requires access to the skull surface by fixing a QWP onto the surface of the skull, which prevents the manipulation required.…”

“…It is clear that the deterioration of the resolution arises from the passage through the skull, which could be avoided with different techniques, such as skull optical clearing 18 or adaptive imaging via frontwave manipulation. 19 However, optical clearing requires access to the skull surface by fixing a QWP onto the surface of the skull, which prevents the manipulation required. Implementation of an adaptive optic component to the system may be more suitable for future experiments.…”

3D morphometric analysis of mouse skulls using microcomputed tomography and computer vision

“…The ability to image brain structure and physiology through the skull has important applications in neuroscience, particularly in studies where immune disruption caused by implantation of cranial windows 13,14 is undesirable. Methods for imaging the brain through the skull include using NIR light 15 , visible light 16 , optical coherence tomography 17 , or three-photon imaging 18 . Other approaches include thinned skull imaging and polished skull techniques 19 .…”