Structural and functional brain imaging using extended-focus optical coherence tomography and microscopy

Neuroimaging techniques aim at revealing the anatomy and functional organisation of cerebral structures. Over the past decades, functional magnetic resonance imaging (fMRI) has revolutionized our understanding of human cerebral physiology through its ability to probe neural activity throughout the entire brain in a non-invasive fashion. Nevertheless, despite recent technological improvements, the spatial resolution of fMRI remains limited to a few hundreds of microns, restricting its use to macroscopic studies. Microscopic imaging solutions have been proposed to circumvent this limitation and have enabled revealing the existence of various cerebral structures, such as neuronal and vascular networks and their contribution to information processing and blood flow regulation within the brain. Optical imaging has proven, through its increased resolution and available contrast mechanisms, to be a valuable complement to fMRI for cellular-scale imaging. In this context, we demonstrate here the capabilities of an extension of optical coherence tomography, termed extended-focus optical coherence tomography (xf-OCT), in imaging cerebral structure and function at high resolution and very high acquisitions rates. Optical coherence tomography is an interferometric imaging technique using a low-coherence illumination source to provide fast, three-dimensional imaging of the back-scattering of tissue and cells. By multiplexing the interferometric ranging over several spectral channels, Fourier-domain OCT performs depth-resolved imaging at very high acquisition rates and high sensitivity. Increasing the lateral resolution of optical systems typically reduces the available depth-of-field and thus hampers this depth multiplexing advantage of OCT. Extended-focus systems aim at alleviating this trade-off between imaging depth and lateral resolution through the use of diffraction-less beams such as Bessel beams, providing high resolution imaging over large depths. The xf-OCT system therefore combines fast acquisition rates and high resolution, both characteristics required to image and study the structure and function of microscopic constituents of cerebral tissue. In this work, we performed functional brain imaging using the ability of xf-OCT to obtain quantita- tive measurements of blood flow in the brain. Changes in blood velocity evoked by neuronal activation were monitored and maps of hemodynamic activity were generated by adapting tools routinely used in fMRI to xf-OCT imaging. Additionally, three novel xf-OCT instruments are presented, wherein the advantages of different spectral ranges are exploited to reach specific imaging parameters. The increased contrast and resolution afforded by an illumination in the visible spectral range was used in two extended-focus optical coherence microscopy (xf-OCM) implementations for subcellular imaging of ex-vivo brain slices and cellular imaging of neurons, capillaries and myelinated axons in the superficial cortex in-vivo. Subsequently, an xf-OCT system is presented, operating in the infrared spectral range, wherein the reduced scattering enabled imaging the smallest capillaries deep in the murine cortex in-vivo

3D Time-lapse Imaging and Quantification of Mitochondrial Dynamics

We present a 3D time-lapse imaging method for monitoring mitochondrial dynamics in living HeLa cells based on photothermal optical coherence microscopy and using novel surface functionalization of gold nanoparticles. The biocompatible protein-based biopolymer coating contains multiple functional groups which impart better cellular uptake and mitochondria targeting efficiency. The high stability of the gold nanoparticles allows continuous imaging over an extended time up to 3000 seconds without significant cell damage. By combining temporal autocorrelation analysis with a classical diffusion model, we quantify mitochondrial dynamics and cast these results into 3D maps showing the heterogeneity of diffusion parameters across the whole cell volume

Statistical parametric mapping of stimuli evoked changes in total blood flow velocity in the mouse cortex obtained with extended-focus optical coherence microscopy

Functional magnetic resonance (fMRI) imaging is the current gold-standard in neuroimaging. fMRI exploits local changes in blood oxygenation to map neuronal activity over the entire brain. However, its spatial resolution is currently limited to a few hundreds of microns. Here we use extended-focus optical coherence microscopy (xfOCM) to quantitatively measure changes in blood flow velocity during functional hyperaemia at high spatio-temporal resolution in the somatosensory cortex of mice. As optical coherence microscopy acquires hundreds of depth slices simultaneously, blood flow velocity measurements can be performed over several vessels in parallel. We present the proof-of-principle of an optimised statistical parametric mapping framework to analyse quantitative blood flow timetraces acquired with xfOCM using the general linear model. We demonstrate the feasibility of generating maps of cortical hemodynamic reactivity at the capillary level with optical coherence microscopy. To validate our method, we exploited 3 stimulation paradigms, covering different temporal dynamics and stimulated limbs, and demonstrated its repeatability over 2 trials, separated by a week

Statistical parametric mapping of stimuli evoked changes in total blood flow velocity in the mouse cortex obtained with extended-focus optical coherence microscopy

Functional magnetic resonance (fMRI) imaging is the current gold-standard in neuroimaging. fMRI exploits local changes in blood oxygenation to map neuronal activity over the entire brain. However, its spatial resolution is currently limited to a few hundreds of microns. Here we use extended-focus optical coherence microscopy (xfOCM) to quantitatively measure changes in blood flow velocity during functional hyperaemia at high spatio-temporal resolution in the somatosensory cortex of mice. As optical coherence microscopy acquires hundreds of depth slices simultaneously, blood flow velocity measurements can be performed over several vessels in parallel. We present the proof-of-principle of an optimised statistical parametric mapping framework to analyse quantitative blood flow timetraces acquired with xfOCM using the general linear model. We demonstrate the feasibility of generating maps of cortical hemodynamic reactivity at the capillary level with optical coherence microscopy. To validate our method, we exploited 3 stimulation paradigms, covering different temporal dynamics and stimulated limbs, and demonstrated its repeatability over 2 trials, separated by a week

Optical projection tomography for rapid whole mouse brain imaging

In recent years, three-dimensional mesoscopic imaging has gained significant importance in life sciences for fundamental studies at the whole-organ level. In this manuscript, we present an optical projection tomography (OPT) method designed for imaging of the intact mouse brain. The system features an isotropic resolution of ~50 μm and an acquisition time of four to eight minutes, using a 3-day optimized clearing protocol. Imaging of the brain autofluorescence in 3D reveals details of the neuroanatomy, while the use of fluorescent labels displays the vascular network and amyloid deposition in 5xFAD mice, an important model of Alzheimer’s disease (AD). Finally, the OPT images are compared with histological slices

Genetic and Functional Assessment of the Role of the rs13431652-A and rs573225-A Alleles in the G6PC2 Promoter That Are Strongly Associated With Elevated Fasting Glucose Levels

OBJECTIVE Genome-wide association studies have identified a single nucleotide polymorphism (SNP), rs560887, located in a G6PC2 intron that is highly correlated with variations in fasting plasma glucose (FPG). G6PC2 encodes an islet-specific glucose-6-phosphatase catalytic subunit. This study examines the contribution of two G6PC2 promoter SNPs, rs13431652 and rs573225, to the association signal. RESEARCH DESIGN AND METHODS We genotyped 9,532 normal FPG participants (FPG <6.1 mmol/l) for three G6PC2 SNPs, rs13431652 (distal promoter), rs573225 (proximal promoter), rs560887 (3rd intron). We used regression analyses adjusted for age, sex, and BMI to assess the association with FPG and haplotype analyses to assess comparative SNP contributions. Fusion gene and gel retardation analyses characterized the effect of rs13431652 and rs573225 on G6PC2 promoter activity and transcription factor binding. RESULTS Genetic analyses provide evidence for a strong contribution of the promoter SNPs to FPG variability at the G6PC2 locus (rs13431652: β = 0.075, P = 3.6 × 10−35; rs573225 β = 0.073 P = 3.6 × 10−34), in addition to rs560887 (β = 0.071, P = 1.2 × 10−31). The rs13431652-A and rs573225-A alleles promote increased NF-Y and Foxa2 binding, respectively. The rs13431652-A allele is associated with increased FPG and elevated promoter activity, consistent with the function of G6PC2 in pancreatic islets. In contrast, the rs573225-A allele is associated with elevated FPG but reduced promoter activity. CONCLUSIONS Genetic and in situ functional data support a potential role for rs13431652, but not rs573225, as a causative SNP linking G6PC2 to variations in FPG, though a causative role for rs573225 in vivo cannot be ruled out