Accurate sensor placement is vital for non-invasive brain imaging, particularly for functional near-infrared spectroscopy (fNIRS) and diffuse optical tomography (DOT), which lack standardized layouts such as those in electroencephalography (EEG). Custom, manually prepared probe layouts on textile caps are often imprecise and labor intensive. We introduce a method for creating personalized, 3D-printed headgear, enabling the accurate translation of 3D brain coordinates to 2D printable panels for custom fNIRS and EEG sensor layouts while reducing costs and manual labor. Our approach uses atlas-based or subject-specific head models and a spring-relaxation algorithm for flattening 3D coordinates onto 2D panels, using 10-5 EEG coordinates for reference. This process ensures geometrical fidelity, crucial for accurate probe placement. Probe geometries and holder types are customizable and printed directly on the cap, making the approach agnostic to instrument manufacturers and probe types. Our ninjaCap method offers 2.7±1.8 mm probe placement accuracy. Over the last five years, we have developed and validated this approach with over 50 cap models and 500 participants. A cloud-based ninjaCap generation pipeline along with detailed instructions is now available at openfnirs.org. The ninjaCap marks a significant advancement in creating individualized neuroimaging caps, reducing costs and labor while improving probe placement accuracy, thereby reducing variability in research.
SignificanceWidefield microscopy of the entire dorsal part of mouse cerebral cortex enables large-scale (“mesoscopic”) imaging of different aspects of neuronal activity with spectrally compatible fluorescent indicators as well as hemodynamics via oxy- and deoxyhemoglobin absorption. Versatile and cost-effective imaging systems are needed for large-scale, color-multiplexed imaging of multiple fluorescent and intrinsic contrasts.AimWe aim to develop a system for mesoscopic imaging of two fluorescent and two reflectance channels.ApproachExcitation of red and green fluorescence is achieved through epi-illumination. Hemoglobin absorption imaging is achieved using 525- and 625-nm light-emitting diodes positioned around the objective lens. An aluminum hemisphere placed between objective and cranial window provides diffuse illumination of the brain. Signals are recorded sequentially by a single sCMOS detector.ResultsWe demonstrate the performance of our imaging system by recording large-scale spontaneous and stimulus-evoked neuronal, cholinergic, and hemodynamic activity in awake, head-fixed mice with a curved “crystal skull” window expressing the red calcium indicator jRGECO1a and the green acetylcholine sensor GRABACh3.0. Shielding of illumination light through the aluminum hemisphere enables concurrent recording of pupil diameter changes.ConclusionsOur widefield microscope design with a single camera can be used to acquire multiple aspects of brain physiology and is compatible with behavioral readouts of pupil diameter.
Prof. Na Ji (UC Berkeley) discusses her pioneering work and motivation in adaptive optics, microscopy, and beyond, in an interview with former trainee Anderson Chen (Prisma Therapeutics, Inc.).
We present DeepVIDv2, a resolution-improved self-supervised voltage imaging denoising approach that achieves higher spatial resolution while preserving fast neuronal dynamics. While existing methods enhance signal-to-noise ratio (SNR), they compromise spatial resolution and result in blurry outputs. By disentangling spatial and temporal performance into two parameters, DeepVIDv2 overcomes the tradeoff faced by its predecessor. This advancement enables more effective analysis of high-speed, large-population voltage imaging data.
SignificanceBrief disruptions in capillary flow, commonly referred to as capillary “stalling,” have gained interest recently for their potential role in disrupting cerebral blood flow and oxygen delivery. Approaches to studying this phenomenon have been hindered by limited volumetric imaging rates and cumbersome manual analysis. The ability to precisely and efficiently quantify the dynamics of these events will be key in understanding their potential role in stroke and neurodegenerative diseases, such as Alzheimer’s disease.AimOur study aimed to demonstrate that the fast volumetric imaging rates offered by Bessel beam two-photon microscopy combined with improved data analysis throughput allows for faster and more precise measurement of capillary stall dynamics.ResultsWe found that while our analysis approach was unable to achieve full automation, we were able to cut analysis time in half while also finding stalling events that were missed in traditional blind manual analysis. The resulting data showed that our Bessel beam system was captured more stalling events compared to optical coherence tomography, particularly shorter stalling events. We then compare differences in stall dynamics between a young and old group of mice as well as a demonstrate changes in stalling before and after photothrombotic model of stroke. Finally, we also demonstrate the ability to monitor arteriole dynamics alongside stall dynamics.ConclusionsBessel beam two-photon microscopy combined with high throughput analysis is a powerful tool for studying capillary stalling due to its ability to monitor hundreds of capillaries simultaneously at high frame rates.
High-speed low-light two-photon voltage imaging is an emerging tool to simultaneously monitor neuronal activity from a large number of neurons. However, shot noise dominates pixel-wise measurements and the neuronal signals are difficult to be identified in the single-frame raw measurement. We developed a self-supervised deep learning framework for voltage imaging denoising, DeepVID, without the need for any high-SNR measurements. DeepVID infers the underlying fluorescence signal based on independent temporal and spatial statistics of the measurement that is attributable to shot noise. DeepVID achieved a 15-fold improvement in SNR when comparing denoised and raw image data.
Neurophotonics was launched in 2014 coinciding with the launch of the BRAIN Initiative focused on development of technologies for advancement of neuroscience. For the last seven years, Neurophotonics’ agenda has been well aligned with this focus on neurotechnologies featuring new optical methods and tools applicable to brain studies. While the BRAIN Initiative 2.0 is pivoting towards applications of these novel tools in the quest to understand the brain, in this article we review an extensive and diverse toolkit of novel methods to explore brain function that have emerged from the BRAIN Initiative and related large-scale efforts for measurement and manipulation of brain structure and function. Here, we focus on neurophotonic tools mostly applicable to animal studies. A companion article, scheduled to appear later this year, will cover diffuse optical imaging methods applicable to noninvasive human studies. For each domain, we outline the current state-of-the-art of the respective technologies, identify the areas where innovation is needed and provide an outlook for the future directions.
Serial Sectioning Optical Coherence Tomography(serial sectioning OCT) has been widely used to investigate the structural and pathological features of brain samples. OCT is an optical imaging technique that provides both the 3D structure of the tissue as well as the optical properties including the scattering coefficient (μs) and back-scattering coefficient (μb). Serial sectioning OCT allows the reconstruction of distortion-free volumetric images at high contrast and high resolution, which has proven to be useful for the detection of cancerous tissue boundaries, visualizing 3D vascular structures and measuring neuron density. The tissue optical properties extracted from the OCT depth profile has been shown to be related to myelin content and neuron density. However, no quantitative correlation of the tissue optical properties with myelin content and neuron density has been reported. Establishing a quantitative relationship will potentially benefit the segmentation of anatomical layers and the characterization of demyelination and neuron loss, which are related to neurological diseases such as Alzheimer’s and Chronic Traumatic Encephalopathy(CTE). Here, we demonstrate using block-face imaging with optical coherence tomography (OCT) to quantitatively measure myelin content and neuron density in the human brain. By correlating the OCT measurements of tissue optical properties with the ground truth of myelin content and neuron density provided by histology, we found that the scattering coefficient possesses a linear relationship with the myelin content across different regions of the human brain, while the neuron density only slightly modulate the overall tissue scattering properties
We combined a home-built two photon microscope with a Bessel focus capable of volumetric imaging with a custom scan pattern that allowed us to generate image pairs with a time delay of ~12 ms. This approach allows us to track red blood cell (RBC) flow along individual capillaries and therefore calculate RBC speed. We demonstrate the ability to measure flow speeds in capillaries across a 475x475x150 µm volume which will enable us to monitor network wide changes in capillary flow.
The signal of optical coherence tomography (OCT) decays exponentially in depth due to tissue scattering, resulting in indistinct tissue features in three-dimension. Moreover, due to limited light penetration depth, extensive volumetric investigation is usually constrained for large-scale biological samples. By integrating serial sectioning technology with block-face imaging, we establish a volumetric OCT acquisition and reconstruction pipeline that incorporates depth-resolved attenuation coefficient estimation, volumetric stitching and filtering, and feature enhancement visualization. We demonstrate this pipeline on ex vivo human brain volumes of several cubic centimeters with 5 um isotropic resolution.
Scanning microscopy’s resolution of highly localized dynamics in awake animals is limited by bulk motion. We present a modular optical flow sensor based on spectral domain OCT which measures the specimen’s 3D displacement in real-time. A rose-function scan pattern rapidly acquires B-lines used to estimate displacements via an adaptive cross-correlation approach. Axial displacements are estimated from phase fluctuations apparent in the cross-correlation of the complex SD-OCT signal. The technique’s 3-dimensional readout rate and sensitivity to physiologically-relevant motion frequencies and magnitudes is evaluated.
Significance: The optical properties of biological samples provide information about the structural characteristics of the tissue and any changes arising from pathological conditions. Optical coherence tomography (OCT) has proven to be capable of extracting tissue’s optical properties using a model that combines the exponential decay due to tissue scattering and the axial point spread function that arises from the confocal nature of the detection system, particularly for higher numerical aperture (NA) measurements. A weakness in estimating the optical properties is the inter-parameter cross-talk between tissue scattering and the confocal parameters defined by the Rayleigh range and the focus depth.
Aim: In this study, we develop a systematic method to improve the characterization of optical properties with high-NA OCT.
Approach: We developed a method that spatially parameterizes the confocal parameters in a previously established model for estimating the optical properties from the depth profiles of high-NA OCT.
Results: The proposed parametrization model was first evaluated on a set of intralipid phantoms and then validated using a low-NA objective in which cross-talk from the confocal parameters is negligible. We then utilize our spatially parameterized model to characterize optical property changes introduced by a tissue index matching process using a simple immersion agent, 2,2’-thiodiethonal.
Conclusions: Our approach improves the confidence of parameter estimation by reducing the degrees of freedom in the non-linear fitting model.
Animal models of stroke are used extensively to study the mechanisms involved in the acute and chronic phases of recovery following stroke. A translatable animal model that closely mimics the mechanisms of a human stroke is essential in understanding recovery processes as well as developing therapies that improve functional outcomes. We describe a photothrombosis stroke model that is capable of targeting a single distal pial branch of the middle cerebral artery with minimal damage to the surrounding parenchyma in awake head-fixed mice. Mice are implanted with chronic cranial windows above one hemisphere of the brain that allow optical access to study recovery mechanisms for over a month following occlusion. Additionally, we study the effect of laser spot size used for occlusion and demonstrate that a spot size with small axial and lateral resolution has the advantage of minimizing unwanted photodamage while still monitoring macroscopic changes to cerebral blood flow during photothrombosis. We show that temporally guiding illumination using real-time feedback of blood flow dynamics also minimized unwanted photodamage to the vascular network. Finally, through quantifiable behavior deficits and chronic imaging we show that this model can be used to study recovery mechanisms or the effects of therapeutics longitudinally.
Wavefront-shaping devices incorporated into optical microscopy systems are capable of correcting sample-induced aberrations and recovering diffraction-limited imaging performance. The widespread dissemination and application of adaptive optical techniques, however, requires easy integration of adaptive optical modules, both in terms of hardware and software, into existing microscopes. We built an adaptive optical module with reduced complexity and simplified integration by utilizing a novel segmented deformable mirror and a standalone control software program. We demonstrated its ability to improve image brightness and resolution at depth in the mouse, zebrafish, and fly brains in vivo.
A solar simulator capable of producing an irradiance of 300 suns is reported. Technical challenges were not limited to optical design; developing a methodology to measuring 300 suns was difficult. This document reports on the design of the custom fabricated solar simulator, the measurement methodology for high-powered solar concentrator measurements and empirical results validating the desired simulated power density as well as the irradiance stability, spectral accuracy, beam uniformity and irradiance beam size.
Wavelength conversion (WC) imaging is a methodology that employs temperature sensitive detectors to convert photoinduced
termperature into a detectable optical signal. One specific method is to use molecular detectors such as
thermochromic liquid crystals (TLC), which exhibits thermochromism to observe the surface temperature of an area by
observing the apparent color in the visible spectrum. Utilizing this methodology, an ultra-broadband room temperature
imaging system was envisioned and realized using off the shelf thermochromic liquid crystals. The thermochromic
properties of the sensor were characterized to show a thermochromic coefficient α = 10%/°K and a noise equivalent
power (NEP) of 64 μW. With the TLC camera, images of both pulsed and continuous wave (CW) sources spanning 0.6
μm to 150 μm wavelengths were captured to demonstrate its potential as a portable, low-cost, and ultra-broadband
imaging tool.
THz optics has experienced a tremendous increase in interest among the scientific community as
better THz sources and detection schemes are discovered. With recent studies in THz modulation
experiment using optically excited Si opens a new possibility in constructing THz optics using an
optically controlled THz SLM. Thus, various patterns, such as zone lenses, could be optically
constructed and tuned in real time for used in THz beam correction. For example, optically
constructed Fresnel zone lenses on high-resistivity Si can be actively tuned for focal length and
chromatic aberration, which are just some possible applications of this methodology. In this paper,
we will present results for an optically controlled single pixeled THz semiconductor SLM for use as
a modulator and discuss extensions to applications in zone lenses, diffraction gratings, and other
optical components.
KEYWORDS: Terahertz radiation, Modulation, Signal attenuation, Absorption, Modulators, Silicon, Semiconductors, Telecommunications, Spectroscopy, Near field optics
Optically controlled modulation of broadband THz radiation with a comparably uniform spatial distribution is demonstrated in a Si-based semiconductor structure with moderate doping. Using THz Time-Domain Spectroscopy a maximum intensity modulation of more than 99% was demonstrated for a spectrum ranging from 50GHz to 3.5THz with 3dB attenuation already for optical excitation as low as only 5mW. The uniformity of the modulation was measured and compared to the THz beam profile.
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