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Abstract Speckle contrast optical spectroscopy (SCOS) is an emerging camera-based technique that can measure human cerebral blood flow (CBF) with high signal-to-noise ratio (SNR). At low photon flux levels typically encountered in human CBF measurements, camera noise and nonidealities could significantly impact SCOS measurement SNR and accuracy. Thus, a guide for characterizing, selecting, and optimizing a camera for SCOS measurements is crucial for the development of next-generation optical devices for monitoring human CBF and brain function. Here, we provide such a guide and illustrate it by evaluating three commercially available complementary metal–oxide–semiconductor cameras, considering a variety of factors including linearity, read noise, and quantization distortion. We show that some cameras that are well-suited for general intensity imaging could be challenged in accurately quantifying spatial contrast for SCOS. We then determine the optimal operating parameters for the preferred camera among the three and demonstrate measurement of human CBF with this selected low-cost camera. This work establishes a guideline for characterizing and selecting cameras as well as for determining optimal parameters for SCOS systems.

Abstract Laser speckle contrast imaging (LSCI) is a rapidly developing technology broadly applied for the full-field characterization of tissue perfusion. Over the recent years, significant advancements have been made in interpreting LSCI measurements and improving the technique’s accuracy. On the other hand, the method’s precision has yet to be studied in detail, despite being as important as accuracy for many biomedical applications. Here we combine simulation, theory and animal experiments to systematically evaluate and re-analyze the role of key factors defining LSCI precision—speckle-to-pixel size ratio, polarisation, exposure time and camera-related noise. We show that contrary to the established assumptions, smaller speckle size and shorter exposure time can improve the precision, while the camera choice is less critical and does not affect the signal-to-noise ratio significantly.

Abstract Cerebral blood flow (CBF) is crucial for brain health. Speckle contrast optical spectroscopy (SCOS) is a technique that has been recently developed to measure CBF, but the use of SCOS to measure human brain function at large source-detector separations with comparable or greater sensitivity to cerebral rather than extracerebral blood flow has not been demonstrated. We describe a fiber-based SCOS system capable of measuring human brain activation induced CBF changes at 33 mm source detector separations using CMOS detectors. The system implements a pulsing strategy to improve the photon flux and uses a data processing pipeline to improve measurement accuracy. We show that SCOS outperforms the current leading optical modality for measuring CBF, i.e. diffuse correlation spectroscopy (DCS), achieving more than 10x SNR improvement at a similar financial cost. Fiber-based SCOS provides an alternative approach to functional neuroimaging for cognitive neuroscience and health science applications.

Abstract In age-related neurodegenerative diseases, pathology often develops slowly across the lifespan. As one example, in diseases such as Alzheimer’s, vascular decline is believed to onset decades ahead of symptomology. However, challenges inherent in current microscopic methods make longitudinal tracking of such vascular decline difficult. Here, we describe a suite of methods for measuring brain vascular dynamics and anatomy in mice for over seven months in the same field of view. This approach is enabled by advances in optical coherence tomography (OCT) and image processing algorithms including deep learning. These integrated methods enabled us to simultaneously monitor distinct vascular properties spanning morphology, topology, and function of the microvasculature across all scales: large pial vessels, penetrating cortical vessels, and capillaries. We have demonstrated this technical capability in wild-type and 3xTg male mice. The capability will allow comprehensive and longitudinal study of a broad range of progressive vascular diseases, and normal aging, in key model systems.

Abstract The study of aging and neurodegenerative processes in the human brain requires a comprehensive understanding of cytoarchitectonic, myeloarchitectonic, and vascular structures. Recent computational advances have enabled volumetric reconstruction of the human brain using thousands of stained slices, however, tissue distortions and loss resulting from standard histological processing have hindered deformation‐free reconstruction. Here, the authors describe an integrated serial sectioning polarization‐sensitive optical coherence tomography (PSOCT) and two photon microscopy (2PM) system to provide label‐free multi‐contrast imaging of intact brain structures, including scattering, birefringence, and autofluorescence of human brain tissue. The authors demonstrate high‐throughput reconstruction of 4 × 4 × 2cm3 sample blocks and simple registration between PSOCT and 2PM images that enable comprehensive analysis of myelin content, vascular structure, and cellular information. The high‐resolution 2PM images provide microscopic validation and enrichment of the cellular information provided by the PSOCT optical properties on the same sample, revealing the densely packed fibers, capillaries, and lipofuscin‐filled cell bodies in the cortex and white matter. It is shown that the imaging system enables quantitative characterization of various pathological features in aging process, including myelin degradation, lipofuscin accumulation, and microvascular changes, which opens up numerous opportunities in the study of neurodegenerative diseases in the future.

Background Normal brain function depends on the ability of the vasculature to increase blood flow to regions with high metabolic demands. Impaired neurovascular coupling, such as the local hyperemic response to neuronal activity, may contribute to poor neurological outcome after stroke despite successful recanalization, that is, futile recanalization. Methods and Results Mice implanted with chronic cranial windows were trained for awake head‐fixation before experiments. One‐hour occlusion of the anterior middle cerebral artery branch was induced using single‐vessel photothrombosis. Cerebral perfusion and neurovascular coupling were assessed by optical coherence tomography and laser speckle contrast imaging. Capillaries and pericytes were studied in perfusion‐fixed tissue by labeling lectin and platelet‐derived growth factor receptor β. Arterial occlusion induced multiple spreading depolarizations over 1 hour associated with substantially reduced blood flow in the peri‐ischemic cortex. Approximately half of the capillaries in the peri‐ischemic area were no longer perfused at the 3‐ and 24‐hour follow‐up (45% [95% CI, 33%–58%] and 53% [95% CI, 39%–66%] reduction, respectively; P

Abstract Optical coherence tomography (OCT) is an emerging 3D imaging technique that allows quantification of intrinsic optical properties such as scattering coefficient and back-scattering coefficient, and has proved useful in distinguishing delicate microstructures in the human brain. The origins of scattering in brain tissues are contributed by the myelin content, neuron size and density primarily; however, no quantitative relationships between them have been reported, which hampers the use of OCT in fundamental studies of architectonic areas in the human brain and the pathological evaluations of diseases. Here, we built a generalized linear model based on Mie scattering theory that quantitatively links tissue scattering to myelin content and neuron density in the human brain. We report a strong linear relationship between scattering coefficient and the myelin content that is retained across different regions of the brain. Neuronal cell body turns out to be a secondary contribution to the overall scattering. The optical property of OCT provides a label-free solution for quantifying volumetric myelin content and neuron cells in the human brain.

Functional neuroimaging, which measures hemodynamic responses to brain activity, has great potential for monitoring recovery in stroke patients and guiding rehabilitation during recovery. However, hemodynamic responses after stroke are almost always altered relative to responses in healthy subjects and it is still unclear if these alterations reflect the underlying brain physiology or if the alterations are purely due to vascular injury. In other words, we do not know the effect of stroke on neurovascular coupling and are therefore limited in our ability to use functional neuroimaging to accurately interpret stroke pathophysiology. To address this challenge, we simultaneously captured neural activity, through fluorescence calcium imaging, and hemodynamics, through intrinsic optical signal imaging, during longitudinal stroke recovery. Our data suggest that neurovascular coupling was preserved in the chronic phase of recovery (2 weeks and 4 weeks post-stoke) and resembled pre-stroke neurovascular coupling. This indicates that functional neuroimaging faithfully represents the underlying neural activity in chronic stroke. Further, neurovascular coupling in the sub-acute phase of stroke recovery was predictive of long-term behavioral outcomes. Stroke also resulted in increases in global brain oscillations, which showed distinct patterns between neural activity and hemodynamics. Increased neural excitability in the contralesional hemisphere was associated with increased contralesional intrahemispheric connectivity. Additionally, sub-acute increases in hemodynamic oscillations were associated with improved sensorimotor outcomes. Collectively, these results support the use of hemodynamic measures of brain activity post-stroke for predicting functional and behavioral outcomes.

Optical coherence tomography (OCT) images of ex vivo human brain tissue are corrupted by multiplicative speckle noise that degrades the contrast to noise ratio (CNR) of microstructural compartments. This work proposes a novel algorithm to reduce noise corruption in OCT images that minimizes the penalized negative log likelihood of gamma distributed speckle noise. The proposed method is formulated as a majorize-minimize problem that reduces to solving an iterative regularized least squares optimization. We demonstrate the usefulness of the proposed method by removing speckle in simulated data, phantom data and real OCT images of human brain tissue. We compare the proposed method with state of the art filtering and non-local means based denoising methods. We demonstrate that our approach removes speckle accurately, improves CNR between different tissue types and better preserves small features and edges in human brain tissue.

Previous studies suggest that producing and comprehending semantically related words relies on inhibitory control over competitive lexical selection which results in the recruitment of the left inferior frontal gyrus (IFG). Few studies, however, have examined the involvement of other regions of the frontal cortex, such as the dorsolateral prefrontal cortex (DLPFC), despite its role in cognitive control related to lexical processing. The primary objective of this study was to elucidate the role of the DLPFC in the production and comprehension of semantically and phonologically related words in blocked cyclic naming and picture–word matching paradigms. Twenty-one adults participated in neuroimaging with functional near-infrared spectroscopy to measure changes in oxygenated and deoxygenated hemoglobin concentrations across the bilateral frontal cortex during blocked cyclic picture naming and blocked cyclic picture–word-matching tasks. After preprocessing, oxygenated and deoxygenated hemoglobin concentrations were obtained for each task (production, comprehension), condition (semantic, phonological) and region (DLPFC, IFG). The results of pairwise t-tests adjusted for multiple comparisons showed significant increases in oxygenated hemoglobin concentration over baseline in the bilateral DLPFC during picture naming for phonologically related words. For picture–word matching, we found significant increases in oxygenated hemoglobin concentration over baseline in the right DLPFC for semantically related words and in the right IFG for phonologically related words. We discuss the results in light of the inhibitory attentional control over competitive lexical access theory in contrast to alternative potential explanations for the findings.

Brain reorganization patterns associated with language recovery after stroke have long been debated. Studying mechanisms of spontaneous and treatment-induced language recovery in post-stroke aphasia requires a network-based approach given the potential for recruitment of perilesional left hemisphere language regions, homologous right hemisphere language regions, and/or spared bilateral domain-general regions. Recent hardware, software, and methodological advances in functional near-infrared spectroscopy (fNIRS) make it well-suited to examine this question. fNIRS is cost-effective with minimal contraindications, making it a robust option to monitor treatment-related brain activation changes over time. Establishing clear activation patterns in neurotypical adults during language and domain-general cognitive processes via fNIRS is an important first step. Some fNIRS studies have investigated key language processes in healthy adults, yet findings are challenging to interpret in the context of methodological limitations. This pilot study used fNIRS to capture brain activation during language and domain-general processing in neurotypicals and individuals with aphasia. These findings will serve as a reference when interpreting treatment-related changes in brain activation patterns in post-stroke aphasia in the future. Twenty-four young healthy controls, seventeen older healthy controls, and six individuals with left hemisphere stroke-induced aphasia completed two language tasks (i.e., semantic feature, picture naming) and one domain-general cognitive task (i.e., arithmetic) twice during fNIRS. The probe covered bilateral frontal, parietal, and temporal lobes and included short-separation detectors for scalp signal nuisance regression. Younger and older healthy controls activated core language regions during semantic feature processing (e.g., left inferior frontal gyrus pars opercularis) and lexical retrieval (e.g., left inferior frontal gyrus pars triangularis) and domain-general regions (e.g., bilateral middle frontal gyri) during hard versus easy arithmetic as expected. Consistent with theories of post-stroke language recovery, individuals with aphasia activated areas outside the traditional networks: left superior frontal gyrus and left supramarginal gyrus during semantic feature judgment; left superior frontal gyrus and right precentral gyrus during picture naming; and left inferior frontal gyrus pars opercularis during arithmetic processing. The preliminary findings in the stroke group highlight the utility of using fNIRS to study language and domain-general processing in aphasia.

Ischemic stroke leads to a reduction or complete loss of blood supply causing injury to brain tissue, which ultimately leads to behavioral impairment. Optical techniques are widely used to study the structural and functional changes that result as a consequence of ischemic stroke both in the acute and chronic phases of stroke recovery. It is currently a challenge to accurately estimate the spatial extent of the infarct without the use of histological parameters however, and in order to follow recovery mechanisms longitudinally at the mesoscopic scale it is essential to know the spatial extent of the stroke core. In this paper we first establish optical coherence tomography (OCT) as a reliable indicator of the stroke core by analyzing signal attenuation and spatially correlating it with the infarct, determined by staining with triphenyl-tetrazolium chloride (TTC). We then introduce spatial frequency domain imaging (SFDI) as a mesoscopic optical technique that can be used to accurately measure the infarct spatial extent by exploiting changes in optical scattering that occur as a consequence of ischemic stroke. Additionally, we follow the progression of ischemia through the acute and sub-acute phases of stroke recovery using both OCT and SFDI and show a consistently high spatial overlap in estimating infarct location. The use of SFDI in assessing infarct location will allow longitudinal studies targeted at following functional recovery mechanisms on a mesoscopic level without having to sacrifice the mouse acutely.

Chronic cranial windows allow for longitudinal brain imaging experiments in awake, behaving mice. Different imaging technologies have their unique advantages and combining multiple imaging modalities offers measurements of a wide spectrum of neuronal, glial, vascular, and metabolic parameters needed for comprehensive investigation of physiological and pathophysiological mechanisms. Here, we detail a suite of surgical techniques for installation of different cranial windows targeted for specific imaging technologies and their combination. Following these techniques and practices will yield higher experimental success and reproducibility of results.

Objective and Impact Statement. Segmentation of blood vessels from two-photon microscopy (2PM) angiograms of brains has important applications in hemodynamic analysis and disease diagnosis. Here, we develop a generalizable deep learning technique for accurate 2PM vascular segmentation of sizable regions in mouse brains acquired from multiple 2PM setups. The technique is computationally efficient, thus ideal for large-scale neurovascular analysis. Introduction. Vascular segmentation from 2PM angiograms is an important first step in hemodynamic modeling of brain vasculature. Existing segmentation methods based on deep learning either lack the ability to generalize to data from different imaging systems or are computationally infeasible for large-scale angiograms. In this work, we overcome both these limitations by a method that is generalizable to various imaging systems and is able to segment large-scale angiograms. Methods. We employ a computationally efficient deep learning framework with a loss function that incorporates a balanced binary-cross-entropy loss and total variation regularization on the network’s output. Its effectiveness is demonstrated on experimentally acquired in vivo angiograms from mouse brains of dimensions up to 808×808×702 μm. Results. To demonstrate the superior generalizability of our framework, we train on data from only one 2PM microscope and demonstrate high-quality segmentation on data from a different microscope without any network tuning. Overall, our method demonstrates 10× faster computation in terms of voxels-segmented-per-second and 3× larger depth compared to the state-of-the-art. Conclusion. Our work provides a generalizable and computationally efficient anatomical modeling framework for brain vasculature, which consists of deep learning-based vascular segmentation followed by graphing. It paves the way for future modeling and analysis of hemodynamic response at much greater scales that were inaccessible before.

Abstract A high‐speed, contrast‐free, quantitative ultrasound velocimetry (vUS) for blood flow velocity imaging throughout the rodent brain is developed based on the normalized first‐order temporal autocorrelation function of the ultrasound field signal. vUS is able to quantify blood flow velocity in both transverse and axial directions, and is validated with numerical simulation, phantom experiments, and in vivo measurements. The functional imaging ability of vUS is demonstrated by monitoring the blood flow velocity changes during whisker stimulation in awake mice. Compared to existing Power‐Doppler‐ and Color‐Doppler‐based functional ultrasound imaging techniques, vUS shows quantitative accuracy in estimating both axial and transverse flow speeds and resistance to acoustic attenuation and high‐frequency noise.

In the first study comparing high angular resolution diffusion MRI (dMRI) in the human brain to axonal orientation measurements from polarization-sensitive optical coherence tomography (PSOCT), we compare the accuracy of orientation estimates from various dMRI sampling schemes and reconstruction methods. We find that, if the reconstruction approach is chosen carefully, single-shell dMRI data can yield the same accuracy as multi-shell data, and only moderately lower accuracy than a full Cartesian-grid sampling scheme. Our results suggest that current dMRI reconstruction approaches do not benefit substantially from ultra-high b-values or from very large numbers of diffusion-encoding directions. We also show that accuracy remains stable across dMRI voxel sizes of 1 ​mm or smaller but degrades at 2 ​mm, particularly in areas of complex white-matter architecture. We also show that, as the spatial resolution is reduced, axonal configurations in a dMRI voxel can no longer be modeled as a small set of distinct axon populations, violating an assumption that is sometimes made by dMRI reconstruction techniques. Our findings have implications for in vivo studies and illustrate the value of PSOCT as a source of ground-truth measurements of white-matter organization that does not suffer from the distortions typical of histological techniques.

For the robust estimation of evoked brain activity from functional Near-Infrared Spectroscopy (fNIRS) signals, it is crucial to reduce nuisance signals from systemic physiology and motion. The current best practice incorporates short-separation (SS) fNIRS measurements as regressors in a General Linear Model (GLM). However, several challenging signal characteristics such as non-instantaneous and non-constant coupling are not yet addressed by this approach and additional auxiliary signals are not optimally exploited. We have recently introduced a new methodological framework for the unsupervised multivariate analysis of fNIRS signals using Blind Source Separation (BSS) methods. Building onto the framework, in this manuscript we show how to incorporate the advantages of regularized temporally embedded Canonical Correlation Analysis (tCCA) into the supervised GLM. This approach allows flexible integration of any number of auxiliary modalities and signals. We provide guidance for the selection of optimal parameters and auxiliary signals for the proposed GLM extension. Its performance in the recovery of evoked HRFs is then evaluated using both simulated ground truth data and real experimental data and compared with the GLM with short-separation regression. Our results show that the GLM with tCCA significantly improves upon the current best practice, yielding significantly better results across all applied metrics: Correlation (HbO max. +45%), Root Mean Squared Error (HbO max. −55%), F-Score (HbO up to 3.25-fold) and p-value as well as power spectral density of the noise floor. The proposed method can be incorporated into the GLM in an easily applicable way that flexibly combines any available auxiliary signals into optimal nuisance regressors. This work has potential significance both for conventional neuroscientific fNIRS experiments as well as for emerging applications of fNIRS in everyday environments, medicine and BCI, where high Contrast to Noise Ratio is of importance for single trial analysis.

Within a decade, single trial analysis of functional Near Infrared Spectroscopy (fNIRS) signals has gained significant momentum, and fNIRS joined the set of modalities frequently used for active and passive Brain Computer Interfaces (BCI). A great variety of methods for feature extraction and classification have been explored using state-of-the-art Machine Learning methods. In contrast, signal preprocessing and cleaning pipelines for fNIRS often follow simple recipes and so far rarely incorporate the available state-of-the-art in adjacent fields. In neuroscience, where fMRI and fNIRS are established neuroimaging tools, evoked hemodynamic brain activity is typically estimated across multiple trials using a General Linear Model (GLM). With the help of the GLM, subject, channel, and task specific evoked hemodynamic responses are estimated, and the evoked brain activity is more robustly separated from systemic physiological interference using independent measures of nuisance regressors, such as short-separation fNIRS measurements. When correctly applied in single trial analysis, e.g., in BCI, this approach can significantly enhance contrast to noise ratio of the brain signal, improve feature separability and ultimately lead to better classification accuracy. In this manuscript, we provide a brief introduction into the GLM and show how to incorporate it into a typical BCI preprocessing pipeline and cross-validation. Using a resting state fNIRS data set augmented with synthetic hemodynamic responses that provide ground truth brain activity, we compare the quality of commonly used fNIRS features for BCI that are extracted from (1) conventionally preprocessed signals, and (2) signals preprocessed with the GLM and physiological nuisance regressors. We show that the GLM-based approach can provide better single trial estimates of brain activity as well as a new feature type, i.e., the weight of the individual and channel-specific hemodynamic response function (HRF) regressor. The improved estimates yield features with higher separability, that significantly enhance accuracy in a binary classification task when compared to conventional preprocessing—on average +7.4% across subjects and feature types. We propose to adapt this well-established approach from neuroscience to the domain of single-trial analysis and preprocessing wherever the classification of evoked brain activity is of concern, for instance in BCI.

Functional near infrared spectroscopy (fNIRS) is a non-invasive optical imaging method that provides continuous measure of cortical brain functions. One application has been its use in the evaluation of pain. Previous studies have delineated a deoxygenation process associated with pain in the medial anterior prefrontal region, more specifically, the medial Brodmann Area 10 (BA 10). Such response to painful stimuli has been consistently observed in awake, sedated and anesthetized patients. In this study, we administered oral morphine (15 mg) or placebo to 14 healthy male volunteers with no history of pain or opioid abuse in a crossover double blind design, and performed fNIRS scans prior to and after the administration to assess the effect of morphine on the medial BA 10 pain signal. Morphine is the gold standard for inhibiting nociceptive processing, most well described for brain effects on sensory and emotional regions including the insula, the somatosensory cortex (the primary somatosensory cortex, S1, and the secondary somatosensory cortex, S2), and the anterior cingulate cortex (ACC). Our results showed an attenuation effect of morphine on the fNIRS-measured pain signal in the medial BA 10, as well as in the contralateral S1 (although observed in a smaller number of subjects). Notably, the extent of signal attenuation corresponded with the temporal profile of the reported plasma concentration for the drug. No clear attenuation by morphine on the medial BA 10 response to innocuous stimuli was observed. These results provide further evidence for the role of medial BA 10 in the processing of pain, and also suggest that fNIRS may be used as an objective measure of drug-brain profiles independent of subjective reports.

As near-infrared spectroscopy (NIRS) broadens its application area to different age and disease groups, motion artifacts in the NIRS signal due to subject movement is becoming an important challenge. Motion artifacts generally produce signal fluctuations that are larger than physiological NIRS signals, thus it is crucial to correct for them before obtaining an estimate of stimulus evoked hemodynamic responses. There are various methods for correction such as principle component analysis (PCA), wavelet-based filtering and spline interpolation. Here, we introduce a new approach to motion artifact correction, targeted principle component analysis (tPCA), which incorporates a PCA filter only on the segments of data identified as motion artifacts. It is expected that this will overcome the issues of filtering desired signals that plagues standard PCA filtering of entire data sets. We compared the new approach with the most effective motion artifact correction algorithms on a set of data acquired simultaneously with a collodion-fixed probe (low motion artifact content) and a standard Velcro probe (high motion artifact content). Our results show that tPCA gives statistically better results in recovering hemodynamic response function (HRF) as compared to wavelet-based filtering and spline interpolation for the Velcro probe. It results in a significant reduction in mean-squared error (MSE) and significant enhancement in Pearson's correlation coefficient to the true HRF. The collodion-fixed fiber probe with no motion correction performed better than the Velcro probe corrected for motion artifacts in terms of MSE and Pearson's correlation coefficient. Thus, if the experimental study permits, the use of a collodion-fixed fiber probe may be desirable. If the use of a collodion-fixed probe is not feasible, then we suggest the use of tPCA in the processing of motion artifact contaminated data.