Effect of electric field direction on neuronal activity: an ex-vivo study.

The effect of electrical stimulation on neurons depends on the spatiotemporal properties of the applied electric field as well as on the biophysical properties of the neural tissue, which includes geometric and electrical characteristics of the cells, and the neural circuit dynamics. In this work, we characterize the effect of electric field direction on neural response in cortical layers. This can, for instance, enable more efficient (e.g., with reduced currents) and/or more selective stimulation. We stimulated mice brain slices using a recently developed brain slice platform to study transcranial currents in an ex-vivo model, where electrodes are separated from the brain slice to inject electric fields at a distance. By rotating the electrode array with respect to the slice, we changed the direction of electric field with respect to the cortical column. Our results demonstrate that in somatosensory cortex, the maximum local field potential (LFP) response is attained when the electric field is oriented parallel to the cortical column. For the same field intensity, when the field is oriented perpendicular to the cortical column, the LFP response is absent. This confirms that electric field direction is an important quantity to determine the effect of neuronal stimulation.

Adaptive frequency-domain filtering for neural signal preprocessing.

Electrical interference from various sources is a common issue for experimental extracellular electrophysiology recordings collected using multi-electrode array neural recording systems. This interference deteriorates the signal-to-noise ratio (SNR) of the raw electrophysiology signals and hampers the accuracy of data post-processing using techniques such as spike-sorting. Traditional signal processing methods to digitally remove electrical interference during post-processing include bandpass filtering to limit the signal to the relevant spectral range of the biological data, e.g., the spikes band (300 Hz - 7 kHz), targeted notch filtering to remove power line interference from standard alternating current mains electricity and common reference removal to minimize noise common to all electrodes. These methods require a priori knowledge of the frequency of the interfering signal source to address the unique electromagnetic interference environment of each experimental setup. We discuss an adaptive method for automatically removing narrow-band electrical interference through a spectral peak detection and removal (SPDR) step that can be applied during post-processing of the recorded data, based on the intuition that tall, narrowband signals localized in the signal spectrum correspond to interference, rather than the activity of neurons. A spectral peak prominence (SPP) threshold is used to detect these peaks in the frequency domain, which will then be removed via notch filtering. We applied this method to simulated waveforms and also experimental electrophysiology data collected from cerebral organoids to demonstrate its effectiveness for removing unwanted interference without significantly distorting the neural signals. We discuss that proper selection of the SPP threshold is required to avoid over-filtering, which can result in distortion of the electrophysiology data. We also compare the firing-rate activity in the filtered electrophysiology with fluorescence calcium imaging, a secondary cellular activity marker, to quantify signal distortion and provide bounds on SNR-based optimization of the SPP threshold. The adaptive filtering technique demonstrated in this paper is a powerful method that can automatically detect and remove interband interference in recorded neural signals, potentially enabling data collection in more naturalistic settings where external interference signals are difficult to eliminate.

Optimized virtual optical waveguides enhance light throughput in scattering media.

Ultrasonically-sculpted gradient-index optical waveguides enable non-invasive light confinement inside scattering media. The confinement level strongly depends on ultrasound parameters (e.g., amplitude, frequency), and medium optical properties (e.g., extinction coefficient). We develop a physically-accurate simulator, and use it to quantify these dependencies for a radially-symmetric virtual optical waveguide. Our analysis provides insights for optimizing virtual optical waveguides for given applications. We leverage these insights to configure virtual optical waveguides that improve light confinement fourfold compared to previous configurations at five mean free paths. We show that virtual optical waveguides enhance light throughput by 50% compared to an ideal external lens, in a medium with bladder-like optical properties at one transport mean free path. We corroborate these simulation findings with real experiments: we demonstrate, for the first time, that virtual optical waveguides recycle scattered light, and enhance light throughput by 15% compared to an external lens at five transport mean free paths.

Focused Epicranial Brain Stimulation by Spatial Sculpting of Pulsed Electric Fields Using High Density Electrode Arrays.

Transcranial electrical neuromodulation of the central nervous system is used as a non-invasive method to induce neural and behavioral responses, yet targeted non-invasive electrical stimulation of the brain with high spatial resolution remains elusive. This work demonstrates a focused, steerable, high-density epicranial current stimulation (HD-ECS) approach to evoke neural activity. Custom-designed high-density (HD) flexible surface electrode arrays are employed to apply high-resolution pulsed electric currents through skull to achieve localized stimulation of the intact mouse brain. The stimulation pattern is steered in real time without physical movement of the electrodes. Steerability and focality are validated at the behavioral, physiological, and cellular levels using motor evoked potentials (MEPs), intracortical recording, and c-fos immunostaining. Whisker movement is also demonstrated to further corroborate the selectivity and steerability. Safety characterization confirmed no significant tissue damage following repetitive stimulation. This method can be used to design novel therapeutics and implement next-generation brain interfaces.

2D optical confinement in an etchless stratified trench waveguide.

We demonstrate novel trapezoidal and rectangular stratified trench optical waveguide designs that feature low-loss two-dimensional confinement of guided optical modes that can be realized in continuous polymer thin film layers formed in a trench mold. The design is based on geometrical bends in a thin film core to enable two-dimensional confinement of light in the transverse plane, without any variation in the core thickness. Incidentally, the waveguide design would completely obviate the need for etching the waveguide core, avoiding the scattering loss due to the etched sidewall roughness. This new design exhibits an intrinsic leakage loss due to coupling of light out of the trench, which can be minimized by choosing an appropriate waveguide geometry. Finite-difference eigenmode simulation demonstrates a low intrinsic leakage loss of less than 0.15 dB/cm. We discuss the principle of operation of these stratified trench waveguides and present the design and numerical simulations of a specific realization of this waveguide geometry. The design considerations and tradeoffs in propagation loss and confinement compared with traditional ridge waveguides are discussed.

Large-scale multimodal surface neural interfaces for primates.

Deciphering the function of neural circuits can help with the understanding of brain function and treating neurological disorders. Progress toward this goal relies on the development of chronically stable neural interfaces capable of recording and modulating neural circuits with high spatial and temporal precision across large areas of the brain. Advanced innovations in designing high-density neural interfaces for small animal models have enabled breakthrough discoveries in neuroscience research. Developing similar neurotechnology for larger animal models such as nonhuman primates (NHPs) is critical to gain significant insights for translation to humans, yet still it remains elusive due to the challenges in design, fabrication, and system-level integration of such devices. This review focuses on implantable surface neural interfaces with electrical and optical functionalities with emphasis on the required technological features to realize scalable multimodal and chronically stable implants to address the unique challenges associated with nonhuman primate studies.

Enhanced spectral-domain optical coherence tomography (SD-OCT) using in situ ultrasonic virtual tunable optical waveguides.

A conventional optical lens can enhance lateral resolution in optical coherence tomography (OCT) by focusing the input light onto the sample. However, the typical Gaussian beam profile of such a lens will impose a tradeoff between the depth of focus (DOF) and the lateral resolution. The lateral resolution is often compromised to achieve a mm-scale DOF. We have experimentally shown that using a cascade system of an ultrasonic virtual tunable optical waveguide (UVTOW) and a short focal-length lens can provide a large DOF without severely compromising the lateral resolution compared to an external lens with the same effective focal length. In addition, leveraging the reconfigurability of UVTOW, we show that the focal length of the cascade system can be tuned without the need for mechanical translation of the optical lens. We compare the performance of the cascade system with a conventional optical lens to demonstrate enhanced DOF without compromising the lateral resolution as well as reconfigurability of UVTOW for OCT imaging.

CMU Array: A 3D nanoprinted, fully customizable high-density microelectrode array platform.

Microelectrode arrays provide the means to record electrophysiological activity critical to brain research. Despite its fundamental role, there are no means to customize electrode layouts to address specific experimental or clinical needs. Moreover, current electrodes demonstrate substantial limitations in coverage, fragility, and expense. Using a 3D nanoparticle printing approach that overcomes these limitations, we demonstrate the first in vivo recordings from electrodes that make use of the flexibility of the 3D printing process. The customizable and physically robust 3D multi-electrode devices feature high electrode densities (2600 channels/cm2 of footprint) with minimal gross tissue damage and excellent signal-to-noise ratio. This fabrication methodology also allows flexible reconfiguration consisting of different individual shank lengths and layouts, with low overall channel impedances. This is achieved, in part, via custom 3D printed multilayer circuit boards, a fabrication advancement itself that can support several biomedical device possibilities. This effective device design enables both targeted and large-scale recording of electrical signals throughout the brain.

A Semi-Automated System for Wafer-Scale Optical Waveguide Characterization.

Integrated photonic waveguide systems are used in biomedical sensing and require robust, high-throughput methods of characterization. Here, we demonstrate a semi-automated robotic system to characterize waveguides at the wafer-scale with minimal human intervention based on imaging the outscattered light to measure the propagation loss. We demonstrate automated input coupling efficiency optimization using closed-loop control of the input fiber position. The automated characterization system collects and combines multiple images of the waveguide to measure the propagation loss. This system allows high-throughput characterization of integrated photonic waveguides and lays the foundation for a fully automated and high throughput system to characterize photonic waveguides at the wafer scale.Clinical Relevance- This method enables high precision, high throughput characterization of optoelectrical neural probes to maximize the yield of surgical implantation and electrophysiology recording.

Parylene Photonic Microimager for Implantable Imaging.

We have recently introduced a fully flexible, compact photonic platform, Parylene photonics. Here, we demonstrate a Parylene photonic waveguide array microimager with a light source localization accuracy of 17.04 µm along the x-axis and 30.07 µm along the y-axis over a 200 µm×1000 µm region. We show the feasibility of fluorescent imaging from mouse brain tissue using the microimager array.Clinical Relevance- Implantable microimagers can be used for clinical intraoperative monitoring as well as structural and functional imaging with cell-type specificity in research.

Flexible optoelectric neural interfaces.

Understanding the neural basis of brain function and dysfunction and designing effective therapeutics require high resolution targeted stimulation and recording of neural activity. Optical methods have been recently developed for neural stimulation as well as functional and structural imaging. These methods call for implantable devices to deliver light into the neural tissue at depth with high spatiotemporal resolution. To address this need, rigid and flexible neurophotonic implants have been recently designed. This article reviews the state-of-the-art flexible passive and active penetrating optical neural probes developed for light delivery with minimal damage to the tissue. Passive and active flexible neurophotonic implants are compared and insights about future directions are provided.

Bioelectrical interfaces with cortical spheroids in three-dimensions.

Objective.Three-dimensional (3D) neuronal spheroid culture serves as a powerful model system for the investigation of neurological disorders and drug discovery. The success of such a model system requires techniques that enable high-resolution functional readout across the entire spheroid. Conventional microelectrode arrays and implantable neural probes cannot monitor the electrophysiology (ephys) activity across the entire native 3D geometry of the cellular construct.Approach.Here, we demonstrate a 3D self-rolled biosensor array (3D-SR-BA) integrated with a 3D cortical spheroid culture for simultaneousin vitroephys recording, functional Ca2+imaging, while monitoring the effect of drugs. We have also developed a signal processing pipeline to detect neural firings with high spatiotemporal resolution from the ephys recordings based on established spike sorting methods.Main results.The 3D-SR-BAs cortical spheroid interface provides a stable, high sensitivity recording of neural action potentials (<50µV peak-to-peak amplitude). The 3D-SR-BA is demonstrated as a potential drug screening platform through the investigation of the neural response to the excitatory neurotransmitter glutamate. Upon addition of glutamate, the neural firing rates increased notably corresponding well with the functional Ca2+imaging.Significance.Our entire system, including the 3D-SR-BA integrated with neuronal spheroid culture, enables simultaneous ephys recording and functional Ca2+imaging with high spatiotemporal resolution in conjunction with chemical stimulation. We demonstrate a powerful toolset for future studies of tissue development, disease progression, and drug testing and screening, especially when combined with native spheroid cultures directly extracted from humans.

Effect of skull thickness and conductivity on current propagation for noninvasively injected currents.

Objective.When currents are injected into the scalp, e.g. during transcranial current stimulation, the resulting currents generated in the brain are substantially affected by the changes in conductivity and geometry of intermediate tissue. In this work, we introduce the concept of 'skull-transparent' currents, for which the changing conductivity does not significantly alter the field while propagating through the head.Approach.We establish transfer functions relating scalp currents to head potentials in accepted simplified models of the head, and find approximations for which skull-transparency holds. The current fields resulting from specified current patterns are calculated in multiple head models, including MRI heads and compared with homogeneous heads to characterize the transparency. Experimental validation is performed by measuring the current field in head phantoms.Main results.The main theoretical result is derived from observing that at high spatial frequencies, in the transfer function relating currents injected into the scalp to potential generated inside the head, the conductivity terms form a multiplicative factor and do not otherwise influence the transfer function. This observation is utilized to design injected current waveforms that maintain nearly identical focusing patterns independently of the changes in skull conductivity and thickness for a wide range of conductivity and thickness values in an idealized spherical head model as well as in a realistic MRI-based head model. Experimental measurements of the current field in an agar-based head phantom confirm the transparency of these patterns.Significance.Our results suggest the possibility that well-chosen patterns of current injection result in precise focusing inside the brain even withouta prioriknowledge of exact conductivities of intermediate layers.

Overcoming the tradeoff between confinement and focal distance using virtual ultrasonic optical waveguides.

A conventional optical lens can be used to focus light into the target medium from outside, without disturbing the medium. The focused spot size is proportional to the focal distance in a conventional lens, resulting in a tradeoff between penetration depth in the target medium and spatial resolution. We have shown that virtual ultrasonically sculpted gradient-index (GRIN) optical waveguides can be formed in the target medium to steer light without disturbing the medium. Here, we demonstrate that such virtual waveguides can relay an externally focused Gaussian beam of light through the medium beyond the focal distance of a single external physical lens, to extend the penetration depth without compromising the spot size. Moreover, the spot size can be tuned by reconfiguring the virtual waveguide. We show that these virtual GRIN waveguides can be formed in transparent and turbid media, to enhance the confinement and contrast ratio of the focused beam of light at the target location. This method can be extended to realize complex optical systems of external physical lenses and in situ virtual waveguides, to extend the reach and flexibility of optical methods.

Parylene photonics: a flexible, broadband optical waveguide platform with integrated micromirrors for biointerfaces.

Targeted light delivery into biological tissue is needed in applications such as optogenetic stimulation of the brain and in vivo functional or structural imaging of tissue. These applications require very compact, soft, and flexible implants that minimize damage to the tissue. Here, we demonstrate a novel implantable photonic platform based on a high-density, flexible array of ultracompact (30 µm × 5 µm), low-loss (3.2 dB/cm at λ = 680 nm, 4.1 dB/cm at λ = 633 nm, 4.9 dB/cm at λ = 532 nm, 6.1 dB/cm at λ = 450 nm) optical waveguides composed of biocompatible polymers Parylene C and polydimethylsiloxane (PDMS). This photonic platform features unique embedded input/output micromirrors that redirect light from the waveguides perpendicularly to the surface of the array for localized, patterned illumination in tissue. This architecture enables the design of a fully flexible, compact integrated photonic system for applications such as in vivo chronic optogenetic stimulation of brain activity.

Ultrasonically sculpted virtual relay lens for in situ microimaging.

We demonstrate in situ non-invasive relay imaging through a medium without inserting physical optical components. We show that a virtual optical graded-index (GRIN) lens can be sculpted in the medium using in situ reconfigurable ultrasonic interference patterns to relay images through the medium. Ultrasonic wave patterns change the local density of the medium to sculpt a graded refractive index pattern normal to the direction of light propagation, which modulates the phase front of light, causing it to focus within the medium and effectively creating a virtual relay lens. We demonstrate the in situ relay imaging and resolving of small features (22 µm) through a turbid medium (optical thickness = 5.7 times the scattering mean free path), which is normally opaque. The focal distance and the numerical aperture of the sculpted optical GRIN lens can be tuned by changing the ultrasonic wave parameters. As an example, we experimentally demonstrate that the axial focal distance can be continuously scanned over a depth of 5.4 mm in the modulated medium and that the numerical aperture can be tuned up to 21.5%. The interaction of ultrasonic waves and light can be mediated through different physical media, including turbid media, such as biological tissue, in which the ultrasonically sculpted GRIN lens can be used for relaying images of the underlying structures through the turbid medium, thus providing a potential alternative to implanting invasive endoscopes.

High Density, Double-Sided, Flexible Optoelectronic Neural Probes With Embedded µLEDs.

Optical stimulation and imaging of neurons deep in the brain require implantable optical neural probes. External optical access to deeper regions of the brain is limited by scattering and absorption of light as it propagates through tissue. Implantable optoelectronic probes capable of high-resolution light delivery and high-density neural recording are needed for closed-loop manipulation of neural circuits. Micro-light-emitting diodes (µLEDs) have been used for optical stimulation, but predominantly on rigid silicon or sapphire substrates. Flexible polymer neural probes would be preferable for chronic applications since they cause less damage to brain tissue. Flexible µLED neural probes have been recently implemented by flip-chip bonding of commercially available µLED chips onto flexible substrates. Here, we demonstrate a monolithic design for flexible optoelectronic neural interfaces with embedded gallium nitride µLEDs that can be microfabricated at wafer-scale. Parylene C is used as the substrate and insulator due to its biocompatibility, compliance, and optical transparency. We demonstrate one-dimensional and two-dimensional individually-addressable µLED arrays. Our µLEDs have sizes as small as 22 × 22 µm in arrays of up to 32 µLEDs per probe shank. These devices emit blue light at a wavelength of 445 nm, suitable for stimulation of channelrhodopsin-2, with output powers greater than 200 µW at 2 mA. Our flexible optoelectronic probes are double-sided and can illuminate brain tissue from both sides. Recording electrodes are co-fabricated with µLEDs on the front- and backside of the optoelectronic probes for electrophysiology recording of neuronal activity from the volumes of tissue on the front- and backside simultaneously with bi-directional optical stimulation.

In situ 3D reconfigurable ultrasonically sculpted optical beam paths.

We demonstrate that optical beams can be spatially and temporally shaped in situ by forming 3D reconfigurable interference patterns of ultrasound waves in the medium. In this technique, ultrasonic pressure waves induce a modulated refractive index pattern that shapes the optical beam as it propagates through the medium. Using custom-designed cylindrical ultrasonic arrays, we demonstrate that complex patterns of light can be sculpted in the medium, including dipole and quadrupole shapes. Additionally, through a combination of theory and experiment, we demonstrate that these optical patterns can be scanned in radial and azimuthal directions. Moreover, we show that light can be selectively confined to different extrema of the spatial ultrasound pressure profile by temporally synchronizing lightwave and ultrasound. Finally, we demonstrate that this technique can also be used to define spatial patterns of light in turbid media. The notion of in situ 3D sculpting of optical beam paths using ultrasound interference patterns can find intriguing applications in biological imaging and manipulation, holography, and microscopy.

Ultrasonic sculpting of virtual optical waveguides in tissue.

Optical imaging and stimulation are widely used to study biological events. However, scattering processes limit the depth to which externally focused light can penetrate tissue. Optical fibers and waveguides are commonly inserted into tissue when delivering light deeper than a few millimeters. This approach, however, introduces complications arising from tissue damage. In addition, it makes it difficult to steer light. Here, we demonstrate that ultrasound can be used to define and steer the trajectory of light within scattering media by exploiting local pressure differences created by acoustic waves that result in refractive index contrasts. We show that virtual light pipes can be created deep into the tissue (>18 scattering mean free paths). We demonstrate the application of this technology in confining light through mouse brain tissue. This technology is likely extendable to form arbitrary light patterns within tissue, extending both the reach and the flexibility of light-based methods.