Cortical microvascular blood flow velocity mapping by combining dynamic light scattering optical coherence tomography and two-photon microscopy.

Significance: The accurate large-scale mapping of cerebral microvascular blood flow velocity is crucial for a better understanding of cerebral blood flow (CBF) regulation. Although optical imaging techniques enable both high-resolution microvascular angiography and fast absolute CBF velocity measurements in the mouse cortex, they usually require different imaging techniques with independent system configurations to maximize their performances. Consequently, it is still a challenge to accurately combine functional and morphological measurements to co-register CBF speed distribution from hundreds of microvessels with high-resolution microvascular angiograms. Aim: We propose a data acquisition and processing framework to co-register a large set of microvascular blood flow velocity measurements from dynamic light scattering optical coherence tomography (DLS-OCT) with the corresponding microvascular angiogram obtained using two-photon microscopy (2PM). Approach: We used DLS-OCT to first rapidly acquire a large set of microvascular velocities through a sealed cranial window in mice and then to acquire high-resolution microvascular angiograms using 2PM. The acquired data were processed in three steps: (i) 2PM angiogram coregistration with the DLS-OCT angiogram, (ii) 2PM angiogram segmentation and graphing, and (iii) mapping of the CBF velocities to the graph representation of the 2PM angiogram. Results: We implemented the developed framework on the three datasets acquired from the mice cortices to facilitate the coregistration of the large sets of DLS-OCT flow velocity measurements with 2PM angiograms. We retrieved the distributions of red blood cell velocities in arterioles, venules, and capillaries as a function of the branching order from precapillary arterioles and postcapillary venules from more than 1000 microvascular segments. Conclusions: The proposed framework may serve as a useful tool for quantitative analysis of large microvascular datasets obtained by OCT and 2PM in studies involving normal brain functioning, progression of various diseases, and numerical modeling of the oxygen advection and diffusion in the realistic microvascular networks.

Aerobic exercise reverses aging-induced depth-dependent decline in cerebral microcirculation.

Aging is a major risk factor for cognitive impairment. Aerobic exercise benefits brain function and may promote cognitive health in older adults. However, underlying biological mechanisms across cerebral gray and white matter are poorly understood. Selective vulnerability of the white matter to small vessel disease and a link between white matter health and cognitive function suggests a potential role for responses in deep cerebral microcirculation. Here, we tested whether aerobic exercise modulates cerebral microcirculatory changes induced by aging. To this end, we carried out a comprehensive quantitative examination of changes in cerebral microvascular physiology in cortical gray and subcortical white matter in mice (3-6 vs. 19-21 months old), and asked whether and how exercise may rescue age-induced deficits. In the sedentary group, aging caused a more severe decline in cerebral microvascular perfusion and oxygenation in deep (infragranular) cortical layers and subcortical white matter compared with superficial (supragranular) cortical layers. Five months of voluntary aerobic exercise partly renormalized microvascular perfusion and oxygenation in aged mice in a depth-dependent manner, and brought these spatial distributions closer to those of young adult sedentary mice. These microcirculatory effects were accompanied by an improvement in cognitive function. Our work demonstrates the selective vulnerability of the deep cortex and subcortical white matter to aging-induced decline in microcirculation, as well as the responsiveness of these regions to aerobic exercise.

Aerobic exercise reverses aging-induced depth-dependent decline in cerebral microcirculation.

Aging is a major risk factor for cognitive impairment. Aerobic exercise benefits brain function and may promote cognitive health in older adults. However, underlying biological mechanisms across cerebral gray and white matter are poorly understood. Selective vulnerability of the white matter to small vessel disease and a link between white matter health and cognitive function suggests a potential role for responses in deep cerebral microcirculation. Here, we tested whether aerobic exercise modulates cerebral microcirculatory changes induced by aging. To this end, we carried out a comprehensive quantitative examination of changes in cerebral microvascular physiology in cortical gray and subcortical white matter in mice (3-6 vs. 19-21 months old), and asked whether and how exercise may rescue age-induced deficits. In the sedentary group, aging caused a more severe decline in cerebral microvascular perfusion and oxygenation in deep (infragranular) cortical layers and subcortical white matter compared with superficial (supragranular) cortical layers. Five months of voluntary aerobic exercise partly renormalized microvascular perfusion and oxygenation in aged mice in a depth-dependent manner, and brought these spatial distributions closer to those of young adult sedentary mice. These microcirculatory effects were accompanied by an improvement in cognitive function. Our work demonstrates the selective vulnerability of the deep cortex and subcortical white matter to aging-induced decline in microcirculation, as well as the responsiveness of these regions to aerobic exercise.

Hyperspectral wide-field time domain single-pixel diffuse optical tomography platform.

We present the design and comprehensive instrumental characterization of a time domain diffuse optical tomography (TD-DOT) platform based on wide-field illumination and wide-field hyperspectral time-resolved single-pixel detection for functional and molecular imaging in turbid media. The proposed platform combines two digital micro-mirror devices (DMDs) to generate structured light and a spectrally resolved multi-anode photomultiplier tube (PMT) detector in time domain for hyperspectral data acquisition over 16 wavelength channels based on the time-correlated single-photon counting (TCSPC) technique. The design of the proposed platform is described in detail and its characteristics in spatial, temporal and spectral dimensions are calibrated and presented. The performance of the system is further validated through a phantom study where two absorbers in glass tubes with spectral contrast are mapped in a turbid medium of ~20 mm thickness. The method presented here offers the potential of accelerating the imaging process and improving reconstruction results in TD-DOT and thus facilitates its wide spread use in preclinical and clinical in vivo imaging scenarios.

Compressive hyperspectral time-resolved wide-field fluorescence lifetime imaging.

Spectrally resolved fluorescence lifetime imaging1-3 and spatial multiplexing1,4,5 have offered information content and collection-efficiency boosts in microscopy, but efficient implementations for macroscopic applications are still lacking. An imaging platform based on time-resolved structured light and hyperspectral single-pixel detection has been developed to perform quantitative macroscopic fluorescence lifetime imaging (MFLI) over a large field of view (FOV) and multiple spectral bands simultaneously. The system makes use of three digital micromirror device (DMD)-based spatial light modulators (SLMs) to generate spatial optical bases and reconstruct N by N images over 16 spectral channels with a time-resolved capability (~40 ps temporal resolution) using fewer than N2 optical measurements. We demonstrate the potential of this new imaging platform by quantitatively imaging near-infrared (NIR) Förster resonance energy transfer (FRET) both in vitro and in vivo. The technique is well suited for quantitative hyperspectral lifetime imaging with a high sensitivity and paves the way for many important biomedical applications.

Wide-field fluorescence molecular tomography with compressive sensing based preconditioning.

Wide-field optical tomography based on structured light illumination and detection strategies enables efficient tomographic imaging of large tissues at very fast acquisition speeds. However, the optical inverse problem based on such instrumental approach is still ill-conditioned. Herein, we investigate the benefit of employing compressive sensing-based preconditioning to wide-field structured illumination and detection approaches. We assess the performances of Fluorescence Molecular Tomography (FMT) when using such preconditioning methods both in silico and with experimental data. Additionally, we demonstrate that such methodology could be used to select the subset of patterns that provides optimal reconstruction performances. Lastly, we compare preconditioning data collected using a normal base that offers good experimental SNR against that directly acquired with optimal designed base. An experimental phantom study is provided to validate the proposed technique.

Hyperspectral time-resolved wide-field fluorescence molecular tomography based on structured light and single-pixel detection.

We present a time-resolved fluorescence diffuse optical tomography platform that is based on wide-field structured illumination, single-pixel detection, and hyperspectral acquisition. Two spatial light modulators (digital micro-mirror devices) are employed to generate independently wide-field illumination and detection patterns, coupled with a 16-channel spectrophotometer detection module to capture hyperspectral time-resolved tomographic data sets. The main system characteristics are reported, and we demonstrate the feasibility of acquiring dense 4D tomographic data sets (space, time, spectra) for time domain 3D quantitative multiplexed fluorophore concentration mapping in turbid media.

Spatial light modulator based active wide-field illumination for ex vivo and in vivo quantitative NIR FRET imaging.

Fluorescence lifetime imaging is playing an increasing role in drug development by providing a sensitive method to monitor drug delivery and receptor-ligand interactions. However, the wide dynamic range of fluorescence intensity emitted by ex vivo and in vivo samples presents challenges in retrieving information over the whole subject accurately and quantitatively. To overcome this challenge, we developed an active wide-field illumination (AWFI) strategy based on a spatial light modulator that acquires optimal fluorescence signals by enhancing the dynamic range, signal to noise ratio, and estimation of lifetime-based parameters. We demonstrate the ability of AWFI to estimate Förster resonance energy transfer (FRET) donor fraction from dissected organs with high accuracy (standard deviation <6%) over the whole field of view, in contrast with the homogenous wide-field illumination. We further report its successful application to quantitative FRET imaging in a live mouse. AWFI allows improved detection of weak signals and enhanced quantitative accuracy in ex vivo and in vivo molecular fluorescence quantitative imaging. The technique allows for robust quantitative estimation of the bio-distribution of molecular probes and lifetime-based parameters over an extended imaging field exhibiting a large range of fluorescence intensities and at a high acquisition speed (less than 1 min).