Review of mesoscopic optical tomography for depth-resolved imaging of hemodynamic changes and neural activities

Abstract

Understanding the functional wiring of neural circuits and their patterns of activation following sensory stimulations is a fundamental task in the field of neuroscience. Furthermore, charting the activity patterns is undoubtedly important to elucidate how neural networks operate in the living brain. However, optical imaging must overcome the effects of light scattering in the tissue, which limit the light penetration depth and affect both the imaging quantitation and sensitivity. Laminar optical tomography (LOT) is a three-dimensional (3-D) in-vivo optical imaging technique that can be used for functional imaging. LOT can achieve both a resolution of 100 to [Formula: see text] and a penetration depth of 2 to 3 mm based either on absorption or fluorescence contrast, as well as large field-of-view and high acquisition speed. These advantages make LOT suitable for 3-D depth-resolved functional imaging of the neural functions in the brain and spinal cords. We review the basic principles and instrumentations of representative LOT systems, followed by recent applications of LOT on 3-D imaging of neural activities in the rat forepaw stimulation model and mouse whisker-barrel system.

Keywords: angled fluorescence laminar optical tomography; functional brain mapping; imaging three-dimensional neural activity; laminar optical tomography; mesoscopic fluorescence molecular tomography; voltage-sensitive dye.

Fig. 1

(a) Schematic of LOT source (S) and detector (D1 to D7) setup. (b) LOT system for depth-resolved hemodynamic imaging of rat cortex.

Fig. 2

(a) Schematic of the aFLOT system. LD, laser diode; O, objective lens; P, polarizer; S, shutter; I, iris; CL, cylindrical lens; F, filter; DM, dichroic mirror. (b) Time-resolved image acquisition protocol.

Fig. 3

LOT of the cortical hemodynamic response to forepaw stimulus in rat. (a) CCD image of rat cortical surface through thinned skull. The region imaged using LOT is indicated by the white dotted lines. m, medial; c: caudal. (b) Depth-resolved LOT images of oxy-, deoxy- and total hemoglobin concentration changes in the cortex 0.6 s after cessation of a 4 s forepaw stimulus at cortical depths of 0, 200, and $600\mu \mathrm{m}$. (c) Depth-resolved cross-section of the ${\mathrm{HbO}}_2$ response at the position indicated with (i) in panel (b), representing a large draining vein. The corresponding ${\mathrm{HbO}}_2$, HbR, and HbT depth-resolved time-courses around $x=1800\mu \mathrm{m}$ (dotted white line) are shown to the right. (d) Depth-resolved cross-section of the ${\mathrm{HbO}}_2$ response at the position indicated with (ii) in panel b. The corresponding ${\mathrm{HbO}}_2$, HbR, and HbT depth resolved time-courses around $x=750\mu \mathrm{m}$ are shown to the right. Numbers on each temporal trace represent their depth of origin in microns. “a,” “v,” and “c” denote regions identified as arteriole, vein, and capillary. (e) Isosurface rendering of hemodynamic response resolved into arterial, capillary, and venous compartments based on their distinctive temporal behaviors (40% isosurface).

Fig. 4

(a) Time course of LOT signals, induced by left hind paw stimulation, collected over 15 s at the $0.9\times \text{muscle threshold}$ (detector 1 with a source–detector separation of $575\mu \mathrm{m}$). (b) Photo of the exposed cortex (left) and maximum IOS acquired simultaneously on the somatosensory cortex (right). (c) 3-D map of neural activation in the spinal cord induced by left hind paw stimulation at the $0.9\times \text{muscle threshold}$. Ipsilateral activation around $z=0.4\mathrm{mm}$ is consistent with interneuron activation. (d) Reconstruction viewed across the segmented volume along the line in (c).

Fig. 5

3-D PSFs of the aFLOT system at $302\mu \mathrm{m}$ (a), $664\mu \mathrm{m}$ (b), and (c) $785\mu \mathrm{m}$. Insets show the isosurface of PSFs with ${\mu }_{\mathrm{s}}^{\prime }=0.5/\mathrm{mm}$. Legends report FWHM in $\mu \mathrm{m}$ in $x$, $y$, and $z$ directions. (d) FWHM versus depths. Filled and open circles and multiplication symbols represent, respectively, the FWHM in $x$, $y$, and $z$ directions of a single PSF at the corresponding depth. (e,f) Reconstructed 3-D aFLOT fluorescence images of $100\text{-}\mu \mathrm{m}$ glass capillary tube superimposed with OCT data.

Fig. 6

3-D changes in fluorescence [$\mathrm{\Delta }F/F$ (%), ordinate] in response to the C2 whisker stimulation reconstructed by the aFLOT system. Time period after stimulation is indicated at the bottom right corner of each image.