Polarization-sensitive optical coherence tomography and scleral collagen fiber orientation in osteogenesis imperfecta.

Osteogenesis imperfecta (OI), a rare genetic connective tissue disorder, primarily arises from pathogenic variants affecting the production or structure of collagen type I. In addition to skeletal fragility, individuals with OI may face an increased risk of developing ophthalmic diseases. This association is believed to stem from the widespread presence of collagen type I throughout various parts of the eye. However, the precise consequences of abnormal collagen type I on different ocular tissues remain unknown. Of particular significance is the sclera, where collagen type I is abundant and crucial for maintaining the structural integrity of the eye. Recent research on healthy individuals has uncovered a unique organizational pattern of collagen fibers within the sclera, characterized by fiber arrangement in both circular and radial layers around the optic nerve head. While the precise function of this organizational pattern remains unclear, it is hypothesized to play a role in providing mechanical support to the optic nerve. The objective of this study is to investigate the impact of abnormal collagen type I on the sclera by assessing the fiber organization near the optic nerve head in individuals with OI and comparing them to healthy individuals. Collagen fiber orientation of the sclera was measured using polarization-sensitive optical coherence tomography (PS-OCT), an extension of the conventional OCT that is sensitive to materials that exhibit birefringence (axial changes in light refraction). Birefringence was quantified and used as imaging contrast to extract collagen fiber orientation as well as the thickness of the radially oriented scleral layer. Three individuals with OI, exhibiting different degrees of disease severity, were assessed and analyzed, along with seventeen healthy individuals. Mean values obtained from individuals with OI were descriptively compared to those of the healthy participant group. PS-OCT revealed a similar orientation pattern of scleral collagen fibers around the optic nerve head between OI individuals and healthy individuals. However, two OI participants exhibited reduced mean birefringence of the radially oriented scleral layer compared to the healthy participant group (OI participant 1 oculus dexter et sinister (ODS): 0.34°/µm, OI participant 2: ODS 0.26°/µm, OI participant 3: OD: 0.29°/µm, OS: 0.28°/µm, healthy participants: ODS 0.38 ± 0.05°/µm). The radially oriented scleral layer was thinner in all OI participants although within ±2 standard deviations of the mean observed in healthy individuals (OI participant 1 OD: 101 µm, OS 104 µm, OI participant 2: OD 97 µm, OS 98 µm, OI participant 3: OD: 94 µm, OS 120 µm, healthy participants: OD 122.8 ± 13.6 µm, OS 120.8 ± 15.1 µm). These findings imply abnormalities in collagen organization or composition, underscoring the necessity for additional research to comprehend the ocular phenotype in OI.

Assessing Lung Fibrosis with ML-Assisted Minimally Invasive OCT Imaging.

This paper presents a combined optical coherence tomography (OCT) imaging/machine learning (ML) technique for real-time analysis of lung tissue morphology to determine the presence and level of invasiveness of idiopathic lung fibrosis (ILF). This is an important clinical problem as misdiagnosis is common, resulting in patient exposure to costly and invasive procedures and substantial use of healthcare resources. Therefore, biopsy is needed to confirm or rule out radiological findings. Videoscopic-assisted thoracoscopic wedge biopsy (VATS) under general anesthesia is typically necessary to obtain enough tissue to make an accurate diagnosis. This kind of biopsy involves the placement of several tubes through the chest wall, one of which is used to cut off a piece of lung to send for evaluation. The removed tissue is examined histopathologically by microscopy to confirm the presence and the pattern of fibrosis. However, VATS pulmonary biopsy can have multiple side effects, including inflammation, tissue morbidity, and severe bleeding, which further degrade the quality of life for the patient. Furthermore, the results are not immediately available, requiring tissue processing and analysis. Here, we report an initial attempt of using ML-assisted polarization sensitive OCT (PS-OCT) imaging for lung fibrosis assessment. This approach has been preliminarily tested on a rat model of lung fibrosis. Our preliminary results show that ML-assisted PS-OCT imaging can detect the presence of ILF with an average of 77% accuracy and 89% specificity.

Quantification of smooth muscle in human airways by polarization-sensitive optical coherence tomography requires correction for perichondrium.

Quantifying airway smooth muscle (ASM) in patients with asthma raises the possibility of improved and personalized disease management. Endobronchial polarization-sensitive optical coherence tomography (PS-OCT) is a promising quantitative imaging approach that is in the early stages of clinical translation. To date, only animal tissues have been used to assess the accuracy of PS-OCT to quantify absolute (rather than relative) ASM in cross sections with directly matched histological cross sections as validation. We report the use of whole fresh human and pig airways to perform a detailed side-by-side qualitative and quantitative validation of PS-OCT against gold-standard histology. We matched and quantified 120 sections from five human and seven pig (small and large) airways and linked PS-OCT signatures of ASM to the tissue structural appearance in histology. Notably, we found that human cartilage perichondrium can share with ASM the properties of birefringence and circumferential alignment of fibers, making it a significant confounder for ASM detection. Measurements not corrected for perichondrium overestimated ASM content several-fold (P < 0.001, paired t test). After careful exclusion of perichondrium, we found a strong positive correlation (r = 0.96, P < 0.00001) of ASM area measured by PS-OCT and histology, supporting the method's application in human subjects. Matching human histology further indicated that PS-OCT allows conclusions on the intralayer composition and in turn potential contractile capacity of ASM bands. Together these results form a reliable basis for future clinical studies.NEW & NOTEWORTHY Polarization-sensitive optical coherence tomography (PS-OCT) may facilitate in vivo measurement of airway smooth muscle (ASM). We present a quantitative validation correlating absolute ASM area from PS-OCT to directly matched histological cross sections using human tissue. A major confounder for ASM quantification was observed and resolved: fibrous perichondrium surrounding hyaline cartilage in human airways presents a PS-OCT signature similar to ASM for birefringence and optic axis orientation. Findings impact the development of automated methods for ASM segmentation.

Enhancing epidural needle guidance using a polarization-sensitive optical coherence tomography probe with convolutional neural networks.

Epidural anesthesia helps manage pain during different surgeries. Nonetheless, the precise placement of the epidural needle remains a challenge. In this study, we developed a probe based on polarization-sensitive optical coherence tomography (PS-OCT) to enhance the epidural anesthesia needle placement. The probe was tested on six porcine spinal samples. The multimodal imaging guidance used the OCT intensity mode and three distinct PS-OCT modes: (1) phase retardation, (2) optic axis, and (3) degree of polarization uniformity (DOPU). Each mode enabled the classification of different epidural tissues through distinct imaging characteristics. To further streamline the tissue recognition procedure, convolutional neural network (CNN) were used to autonomously identify the tissue types within the probe's field of view. ResNet50 models were developed for all four imaging modes. DOPU imaging was found to provide the highest cross-testing accuracy of 91.53%. These results showed the improved precision by PS-OCT in guiding epidural anesthesia needle placement.

Information-Rich Multi-Functional OCT for Adult Zebrafish Intra- and Extracranial Imaging.

The zebrafish serves as a valuable animal model for both intra- and extracranial research, particularly in relation to the brain and skull. To effectively investigate the development and regeneration of adult zebrafish, a versatile in vivo imaging technique capable of showing both intra- and extracranial conditions is essential. In this paper, we utilized a high-resolution multi-functional optical coherence tomography (OCT) to obtain rich intra- and extracranial imaging outcomes of adult zebrafish, encompassing pigmentation distribution, tissue-specific information, cranial vascular imaging, and the monitoring of traumatic brain injury (TBI). Notably, it is the first that the channels through the zebrafish cranial suture, which may have a crucial function in maintaining the patency of the cranial sutures, have been observed. Rich imaging results demonstrated that a high-resolution multi-functional OCT system can provide a wealth of novel and interpretable biological information for intra- and extracranial studies of adult zebrafish.

Polarization coherency matrix tomography.

In this paper, a polarization-sensitive optical coherence tomography (PS-OCT) based polarization coherency matrix tomography (PCMT) combining polarization coherency matrix with Mueller matrix is proposed for the determination of complete polarization properties of tissue. PCMT measures the Jones matrix of biological sample based on similar transformation, in which four elements have initial random phase from different polarization states based on traditional PS-OCT. The results indicate that PCMT can eliminate the phase difference of incident lights with different polarization states. In addition, the polarization coherency matrix using three polarization states has complete information of the sample Jones matrix. Finally, the 16 elements of the sample Mueller matrix are applied for deriving fully polarized optical properties of the sample based on the elliptical diattenuator and the elliptical retarder. Thus, the method based on the PCM and Mueller matrix has the advantage over the traditional PS-OCT.

Development of polarization-sensitive optical coherence tomography imaging platform and metrics to quantify electrostimulation-induced peripheral nerve injury in vivo in a small animal model.

Significance: Neuromodulation devices are rapidly evolving for the treatment of neurological diseases and conditions. Injury from implantation or long-term use without obvious functional losses is often only detectable through terminal histology. New technologies are needed that assess the peripheral nervous system (PNS) under normal and diseased or injured conditions. Aim: We aim to demonstrate an imaging and stimulation platform that can elucidate the biological mechanisms and impacts of neurostimulation in the PNS and apply it to the sciatic nerve to extract imaging metrics indicating electrical overstimulation. Approach: A sciatic nerve injury model in a 15-rat cohort was observed using a newly developed imaging and stimulation platform that can detect electrical overstimulation effects with polarization-sensitive optical coherence tomography. The sciatic nerve was electrically stimulated using a custom-developed nerve holder with embedded electrodes for 1 h, followed by a 1-h recovery period, delivered at above-threshold Shannon model k -values in experimental groups: sham control (SC, n = 5 , 0.0 mA / 0 Hz ), stimulation level 1 (SL1, n = 5 , 3.4 mA / 50 Hz , and k = 2.57 ), and stimulation level 2 (SL2, n = 5 , 6.8 mA / 100 Hz , and k = 3.17 ). Results: The stimulation and imaging system successfully captured study data across the cohort. When compared to a SC after a 1-week recovery, the fascicle closest to the stimulation lead showed an average change of + 4 % / - 309 % (SL1/SL2) in phase retardation and - 79 % / - 148 % in optical attenuation relative to SC. Analysis of immunohistochemistry (IHC) shows a + 1 % / - 36 % difference in myelin pixel counts and - 13 % / + 29 % difference in axon pixel counts, and an overall increase in cell nuclei pixel count of + 20 % / + 35 % . These metrics were consistent with IHC and hematoxylin/eosin tissue section analysis. Conclusions: The poststimulation changes observed in our study are manifestations of nerve injury and repair, specifically degeneration and angiogenesis. Optical imaging metrics quantify these processes and may help evaluate the safety and efficacy of neuromodulation devices.

Improved in vivo optical coherence tomography imaging of animal peripheral nerves using a prism nerve holder.

Significance: Modern optical volumetric imaging modalities, such as optical coherence tomography (OCT), provide enormous information about the structure, function, and physiology of living tissue. Although optical imaging achieves lateral resolution on the order of the wavelength of light used, and OCT achieves axial resolution on a similar micron scale, tissue optical properties, particularly high scattering and absorption, limit light penetration to only a few millimeters. In addition, in vivo imaging modalities are susceptible to significant motion artifacts due to cardiac and respiratory function. These effects limit access to artifact-free optical measurements during peripheral neurosurgery to only a portion of the exposed nerve without further modification to the procedure. Aim: We aim to improve in vivo OCT imaging during peripheral neurosurgery in small and large animals by increasing the amount of visualized nerve volume as well as suppressing motion of the imaged area. Approach: We designed a nerve holder with embedded mirror prisms for peripheral nerve volumetric imaging as well as a specific beam steering strategy to acquire prism and direct view volumes in one session with minimal motion artifacts. Results: The axially imaged volumes from mirror prisms increased the OCT signal intensity by > 22 dB over a 1.25-mm imaging depth in tissue-mimicking phantoms. We then demonstrated the new imaging capabilities in visualizing peripheral nerves from direct and side views in living rats and minipigs using a polarization-sensitive OCT system. Prism views have shown nerve fascicles and vasculature from the bottom half of the imaged nerve which was not visible in direct view. Conclusions: We demonstrated improved OCT imaging during neurosurgery in small and large animals by combining the use of a prism nerve holder with a specifically designed beam scanning protocol. Our strategy can be applied to existing OCT imaging systems with minimal hardware modification, increasing the nerve tissue volume visualized. Enhanced imaging depth techniques may lead to a greater adoption of structural and functional optical biomarkers in preclinical and clinical medicine.

Zebrafish brain and skull imaging based on polarization-sensitive optical coherence tomography.

Zebrafish brain imaging is very important for the study of brain disease and regeneration. We scanned the adult zebrafish brain before and after skull removal and monitored the recovery process of a head wound by polarization-sensitive optical coherence tomography (PS-OCT) in this paper. We analyzed the structure and polarization characteristics of the brain and skull in PS-OCT images, and found their internal microstructure can be clearly identified with the polarization information. Further, we estimated the pigment distribution of the skull area and found that the density of pigment in skull is a critical factor of affecting zebrafish brain in vivo polarization imaging. Our results demonstrated that more features of brain can be displayed by introducing the polarization information, and proved high-resolution PS-OCT will play a great potential role in studying the zebrafish brain and skull.

Requirements and limitations of imaging airway smooth muscle throughout the lung in vivo.

Clinical visualization and quantification of the amount and distribution of airway smooth muscle (ASM) in the lungs of individuals with asthma has major implications for our understanding of airway wall remodeling as well as treatments targeted at the ASM. This paper theoretically investigates the feasibility of quantifying airway wall thickness (focusing on the ASM) throughout the lung in vivo by means of bronchoscopic polarization-sensitive optical coherence tomography (PS-OCT). Using extensive human biobank data from subjects with and without asthma in conjunction with a mathematical model of airway compliance, we define constraints that airways of various sizes pose to any endoscopic imaging technique and how this is impacted by physiologically relevant processes such as constriction, inflation and deflation. We identify critical PS-OCT system parameters and pinpoint parts of the airway tree that are conducive to successful quantification of ASM. We further quantify the impact of breathing and ASM contraction on the measurement error and recommend strategies for standardization and normalization.

Degree of polarization uniformity for dental calculus visualization.

During the treatment for periodontitis, the removal of dental calculus is essential. Previously, we have proposed the DAM algorithm for intuitive identification of the site of lesion, enabling the non-contact assessment during the operation. Nonetheless, the delineation of dental calculus was still imperfect. To this end, here we utilized the power of polarization-sensitive optical coherence tomography and evaluated the contrast called degree of polarization uniformity for dental calculus visualization. The result showed that the selected index demonstrated excellent contrast of dental calculus from other normal dental hard tissues. The proposed contrast is promising for accurate dental calculus delineation.

The Optical Coherence Tomography and Raman Spectroscopy for Sensing of the Bone Demineralization Process.

The presented research was intended to seek new optical methods to investigate the demineralization process of bones. Optical examination of the bone condition could facilitate clinical trials and improve the safety of patients. The authors used a set of complementary methods: polarization-sensitive optical coherence tomography (PS-OCT) and Raman spectroscopy. Chicken bone samples were used in this research. To stimulate in laboratory conditions the process of demineralization and gradual removal of the hydroxyapatite, the test samples of bones were placed into 10% acetic acid. Measurements were carried out in two series. The first one took two weeks with data acquired every day. In the second series, the measurements were made during one day at an hourly interval (after 1, 2, 3, 5, 7, 10, and 24 h). The relation between the content of hydroxyapatite and images recorded using OCT was analyzed and discussed. Moreover, the polarization properties of the bones, including retardation angles of the bones, were evaluated. Raman measurement confirmed the disappearance of the hydroxyapatite and the speed of this process. This work presents the results of the preliminary study on the possibility of measuring changes in bone mineralization by means of the proposed methods and confirms their potential for practical use in the future.

Quantification of volumetric morphometry and optical property in the cortex of human cerebellum at micrometer resolution.

The surface of the human cerebellar cortex is much more tightly folded than the cerebral cortex. Volumetric analysis of cerebellar morphometry in magnetic resonance imaging studies suffers from insufficient resolution, and therefore has had limited impact on disease assessment. Automatic serial polarization-sensitive optical coherence tomography (as-PSOCT) is an emerging technique that offers the advantages of microscopic resolution and volumetric reconstruction of large-scale samples. In this study, we reconstructed multiple cubic centimeters of ex vivo human cerebellum tissue using as-PSOCT. The morphometric and optical properties of the cerebellar cortex across five subjects were quantified. While the molecular and granular layers exhibited similar mean thickness in the five subjects, the thickness varied greatly in the granular layer within subjects. Layer-specific optical property remained homogenous within individual subjects but showed higher cross-subject variability than layer thickness. High-resolution volumetric morphometry and optical property maps of human cerebellar cortex revealed by as-PSOCT have great potential to advance our understanding of cerebellar function and diseases.

Imaging of a retinal pigment epithelium aperture using polarization-sensitive optical coherence tomography.

PURPOSE: To evaluate cases with a retinal pigment epithelium (RPE) aperture using polarization-sensitive optical coherence tomography (PS-OCT). STUDY DESIGN: Retrospective consecutive case series. METHODS: A retrospective study that included three eyes (three patients) with RPE aperture and age-related macular degeneration (AMD) evaluated at the Macular Clinic in Tokyo University Hospital. A three-dimensional dataset of depolarization information was obtained with a clinical prototype of PS-OCT. RESULTS: All patients were categorized as intermediate AMD. RPE apertures were identified with PS-OCT as discontinuities of depolarization in the RPE layer of the pigment epithelial detachment (PED). A nonuniform decrease of depolarization in the RPE layer was also observed around the aperture. Two findings were observed above the aperture, intraretinal focal areas with high reflectivity and increased depolarization and subretinal bands with moderate reflectivity and low depolarization. Retinal sensitivity according to fundus microperimetry measured at 25 points was significantly associated with the degree of depolarization at the corresponding area (r-square = 0.60, p = 0.0001). CONCLUSION: The RPE aperture was characterized as a round discontinuity of depolarization. The findings with PS-OCT suggest atrophic changes in the overlying RPE of the PED. The degree of depolarization was associated with retinal sensitivity. The current results indicate that RPE apertures developed within the spectrum of atrophic AMD.

Compressive sensing for polarization sensitive optical coherence tomography.

In this report, we report on the implementation of compressive sensing (CS) and sparse sampling in polarization sensitive optical coherence tomography (PS-OCT) to reduce the number of B-scans (frames consisting of an array of A-scans, where each represents a single depth profile of reflections) required for effective volumetric (3D dataset composed of an array of B-scans) PS-OCT measurements (i.e. OCT intensity, and phase retardation) reconstruction. Sparse sampling of PS-OCT is achieved through randomization of step sizes along the slow-axis of PS-OCT imaging, covering the same spatial ranges as those with equal slow-axis step sizes, but with a reduced number of B-scans. Tested on missing B-scan rates of 25%, 50% and 75%, we found CS could reconstruct reasonably good (as evidenced by a correlation coefficient >0.6) PS-OCT measurements with a maximum reduced B-scan rate of 50%, thereby accelerating and doubling the rate of volumetric PS-OCT measurements.

In Vivo 3D Determination of Peripapillary Scleral and Retinal Layer Architecture Using Polarization-Sensitive Optical Coherence Tomography.

Purpose: The purpose of this paper was to determine the architecture of the collagen fibers of the peripapillary sclera, the retinal nerve fiber layer (RNFL), and Henle's fiber layer in vivo in 3D using polarization-sensitive optical coherence tomography (PS-OCT). Methods: Seven healthy volunteers were imaged with our in-house built PS-OCT system. PS-OCT imaging included intensity, local phase retardation, relative optic axis, and optic axis uniformity (OAxU). Differential Mueller matrix calculus was used for the first time in ocular tissues to visualize local orientations that varied with depth, incorporating a correction method for the fiber orientation in preceding layers. Results: Scleral collagen fiber orientation images clearly showed an inner layer with an orientation parallel to the RNFL orientation, and a deeper layer where the collagen was circularly oriented. RNFL orientation images visualized the nerve fibers leaving the optic nerve head (ONH) in a radial pattern. The phase retardation and orientation of Henle's fiber layer were visualized locally for the first time. Conclusions: PS-OCT successfully showed the orientation of the retinal nerve fibers, sclera, and Henle's fiber layer, and is to the extent of our knowledge the only technique able to do so in 3D in vivo. Translational Relevance: In vivo 3D imaging of scleral collagen architecture and the retinal neural fibrous structures can improve our understanding of retinal biomechanics and structural alterations in different disease stages of myopia and glaucoma.

Detection of Oral Dysplastic and Early Cancerous Lesions by Polarization-Sensitive Optical Coherence Tomography.

Detection of oral dysplastic and early-stage cancerous lesions is difficult with the current tools. Half of oral cancers are diagnosed in a late stage. Detection of early stromal change to predict malignant transformation is a new direction in the diagnosis of early-stage oral cancer. The application of new optical tools to image stroma in vivo is under investigation, and polarization-sensitive optical coherence tomography (PS-OCT) is potentially one of those tools. This is a preliminary study to sequentially image oral stromal changes from normal, hyperplasia, and dysplasia to early-stage cancer by PS-OCT in vivo. We used 4-Nitroquinoline-1-oxide drinking water to induce dysplasia and early-stage oral cancer in 19 K14-EGFP-miR-211-GFP transgenic mice. A total of 8 normal, 12 hyperplastic, 11 dysplastic, and 4 early-stage cancerous lesions were enrolled. A new analytic process of PS-OCT imaging was proposed, called an en-face birefringence map. From the birefringence map, the sensitivity, specificity, positive predictive value, and negative predictive values to detect dysplasia and early-stage cancer were 100.00%, 95.00%, 93.75%, and 100.00%, respectively, and the kappa value of these images between two investigators was 0.942. The mean size of malignant lesions detected in this study is 1.66 ± 0.93 mm. This pilot animal study validates the use of PS-OCT to detect small and early-stage oral malignancy with high accuracy and consistency.

Jones matrix-based speckle-decorrelation angiography using polarization-sensitive optical coherence tomography.

We show that polarization-sensitive optical coherence tomography angiography (PS-OCTA) based on full Jones matrix assessment of speckle decorrelation offers improved contrast and depth of vessel imaging over conventional OCTA. We determine how best to combine the individual Jones matrix elements and compare the resulting image quality to that of a conventional OCT scanner by co-locating and imaging the same skin locations with closely matched scanning setups. Vessel projection images from finger and forearm skin demonstrate the benefits of Jones matrix-based PS-OCTA. Our study provides a promising starting point and a useful reference for future pre-clinical and clinical applications of Jones matrix-based PS-OCTA.

Intracardiac radiofrequency ablation in living swine guided by polarization-sensitive optical coherence tomography.

SIGNIFICANCE: Pulmonary vein isolation with catheter-based radiofrequency ablation (RFA) is carried out frequently to treat atrial fibrillation. However, RFA lesion creation is only guided by indirect information (e.g., temperature, impedance, and contact force), which may result in poor lesion quality (e.g., nontransmural) and can lead to reoccurrence or complications. AIM: The feasibility of guiding intracardiac RFA with an integrated polarization-sensitive optical coherence tomography (PSOCT)-RFA catheter in the right atria (RA) of living swine is demonstrated. APPROACH: In total, 12 sparse lesions were created in the RA of three living swine using an integrated PSOCT-RFA catheter with standard ablation protocol. PSOCT images were displayed in real time to guide catheter-tissue apposition. After experiments, post-processed PSOCT images were analyzed to assess lesion quality and were compared with triphenyltetrazolium chloride (TTC) lesion quality analysis. RESULTS: Five successful lesions identified with PSOCT images were all confirmed by TTC analysis. In two ablations, PSOCT imaging detected gas bubble formation, indicating overtreatment. Unsuccessful lesions observed with PSOCT imaging were confirmed by TTC analysis. CONCLUSIONS: The results demonstrate that the PSOCT-RFA catheter provides real-time feedback to guide catheter-tissue apposition, monitor lesion quality, and possibly help avoid complications due to overtreatment, which may enable more effective and safer RFA treatment.

Contrast-enhanced serial optical coherence scanner with deep learning network reveals vasculature and white matter organization of mouse brain.

Optical coherence tomography provides volumetric reconstruction of brain structure with micrometer resolution. Gray matter and white matter can be highlighted using conventional and polarization-based contrasts; however, vasculature in ex-vivo fixed brain has not been investigated at large scale due to lack of intrinsic contrast. We present contrast enhancement to visualize the vasculature by perfusing titanium dioxide particles transcardially into the mouse vascular system. The brain, after dissection and fixation, is imaged by a serial optical coherence scanner. Accumulation of particles in blood vessels generates distinguishable optical signals. Among these, the cross-polarization images reveal the vasculature organization remarkably well. The conventional and polarization-based contrasts are still available for probing the gray matter and white matter structures. The segmentation and reconstruction of the vasculature are presented by using a deep learning algorithm. Axonal fiber pathways in the mouse brain are delineated by utilizing the retardance and optic axis orientation contrasts. This is a low-cost method that can be further developed to study neurovascular diseases and brain injury in animal models.