Fully bioresorbable hybrid opto-electronic neural implant system for simultaneous electrophysiological recording and optogenetic stimulation.

Bioresorbable neural implants based on emerging classes of biodegradable materials offer a promising solution to the challenges of secondary surgeries for removal of implanted devices required for existing neural implants. In this study, we introduce a fully bioresorbable flexible hybrid opto-electronic system for simultaneous electrophysiological recording and optogenetic stimulation. The flexible and soft device, composed of biodegradable materials, has a direct optical and electrical interface with the curved cerebral cortex surface while exhibiting excellent biocompatibility. Optimized to minimize light transmission losses and photoelectric artifact interference, the device was chronically implanted in the brain of transgenic mice and performed to photo-stimulate the somatosensory area while recording local field potentials. Thus, the presented hybrid neural implant system, comprising biodegradable materials, promises to provide monitoring and therapy modalities for versatile applications in biomedicine.

KDS2010, a reversible MAO-B inhibitor, extends the lifetime of neural probes by preventing glial scar formation.

Implantable neural probes have been extensively utilized in the fields of neurocircuitry, systems neuroscience, and brain-computer interface. However, the long-term functionality of these devices is hampered by the formation of glial scar and astrogliosis at the surface of electrodes. In this study, we administered KDS2010, a recently developed reversible MAO-B inhibitor, to mice through ad libitum drinking in order to prevent glial scar formation and astrogliosis. The administration of KDS2010 allowed long-term recordings of neural signals with implantable devices, which remained stable over a period of 6 months and even restored diminished neural signals after probe implantation. KDS2010 effectively prevented the formation of glial scar, which consists of reactive astrocytes and activated microglia around the implant. Furthermore, it restored neural activity by disinhibiting astrocytic MAO-B dependent tonic GABA inhibition induced by astrogliosis. We suggest that the use of KDS2010 is a promising approach to prevent glial scar formation around the implant, thereby enabling long-term functionality of neural devices.

Production of Highly Uniform Midbrain Organoids from Human Pluripotent Stem Cells.

Brain organoids have been considered as an advanced platform for in vitro disease modeling and drug screening, but numerous roadblocks exist, such as lack of large-scale production technology and lengthy protocols with multiple manipulation steps, impeding the industrial translation of brain organoid technology. Here, we describe the high-speed and large-scale production of midbrain organoids using a high-throughput screening-compatible platform within 30 days. Micro midbrain organoids (µMOs) exhibit a highly uniform morphology and gene expression pattern with minimal variability. Notably, µMOs show dramatically accelerated maturation, resulting in the generation of functional µMOs within only 30 days of differentiation. Furthermore, individual µMOs display highly consistent responsiveness to neurotoxin, suggesting their usefulness as an in vitro high-throughput drug toxicity screening platform. Collectively, our data indicate that µMO technology could represent an advanced and robust platform for in vitro disease modeling and drug screening for human neuronal diseases.

A wireless, solar-powered, optoelectronic system for spatial restriction-free long-term optogenetic neuromodulations.

Numerous wireless optogenetic systems have been reported for practical tether-free optogenetics in freely moving animals. However, most devices rely on battery-powered or coil-powered systems requiring periodic battery replacement or bulky, high-cost charging equipment with delicate antenna design. This leads to spatiotemporal constraints, such as limited experimental duration due to battery life or animals' restricted movement within specific areas to maintain wireless power transmission. In this study, we present a wireless, solar-powered, flexible optoelectronic device for neuromodulation of the complete freely behaving subject. This device provides chronic operation without battery replacement or other external settings including impedance matching technique and radio frequency generators. Our device uses high-efficiency, thin InGaP/GaAs tandem flexible photovoltaics to harvest energy from various light sources, which powers Bluetooth system to facilitate long-term, on-demand use. Observation of sustained locomotion behaviors for a month in mice via secondary motor cortex area stimulation demonstrates the notable capabilities of our device, highlighting its potential for space-free neuromodulating applications.

A neural probe for concurrent real-time measurement of multiple neurochemicals with electrophysiology in multiple brain regions in vivo.

Real-time monitoring of various neurochemicals with high spatial resolution in multiple brain regions in vivo can elucidate neural circuits related to various brain diseases. However, previous systems for monitoring neurochemicals have limitations in observing multiple neurochemicals without crosstalk in real time, and these methods cannot record electrical activity, which is essential for investigating neural circuits. Here, we present a real-time bimodal (RTBM) neural probe that uses monolithically integrated biosensors and multiple shanks to study the connectivity of neural circuits by measuring multiple neurochemicals and electrical neural activity in real time. Using the RTBM probe, we demonstrate concurrent measurements of four neurochemicals-glucose, lactate, choline, and glutamate without cross-talking each other-and electrical activity in real time in vivo. Additionally, we show the functional connectivity between the medial prefrontal cortex and mediodorsal thalamus through the simultaneous measurement of chemical and electrical signals. We expect that our device will contribute to not only elucidating the role of neurochemicals in neural circuits related to brain functions but also developing drugs for various brain diseases related to neurochemicals.

Symmetry Breaking of Human Pluripotent Stem Cells (hPSCs) in Micropattern Generates a Polarized Spinal Cord-Like Organoid (pSCO) with Dorsoventral Organization.

Axis formation and related spatial patterning are initiated by symmetry breaking during development. A geometrically confined culture of human pluripotent stem cells (hPSCs) mimics symmetry breaking and cell patterning. Using this, polarized spinal cord organoids (pSCOs) with a self-organized dorsoventral (DV) organization are generated. The application of caudalization signals promoted regionalized cell differentiation along the radial axis and protrusion morphogenesis in confined hPSC colonies. These detached colonies grew into extended spinal cord-like organoids, which established self-ordered DV patterning along the long axis through the spontaneous expression of polarized DV patterning morphogens. The proportions of dorsal/ventral domains in the pSCOs can be controlled by the changes in the initial size of micropatterns, which altered the ratio of center-edge cells in 2D. In mature pSCOs, highly synchronized neural activity is separately detected in the dorsal and ventral side, indicating functional as well as structural patterning established in the organoids. This study provides a simple and precisely controllable method to generate spatially ordered organoids for the understanding of the biological principles of cell patterning and axis formation during neural development.

Integration of reconfigurable microchannels into aligned three-dimensional neural networks for spatially controllable neuromodulation.

Anisotropically organized neural networks are indispensable routes for functional connectivity in the brain, which remains largely unknown. While prevailing animal models require additional preparation and stimulation-applying devices and have exhibited limited capabilities regarding localized stimulation, no in vitro platform exists that permits spatiotemporal control of chemo-stimulation in anisotropic three-dimensional (3D) neural networks. We present the integration of microchannels seamlessly into a fibril-aligned 3D scaffold by adapting a single fabrication principle. We investigated the underlying physics of elastic microchannels' ridges and interfacial sol-gel transition of collagen under compression to determine a critical window of geometry and strain. We demonstrated the spatiotemporally resolved neuromodulation in an aligned 3D neural network by local deliveries of KCl and Ca2+ signal inhibitors, such as tetrodotoxin, nifedipine, and mibefradil, and also visualized Ca2+ signal propagation with a speed of ~3.7 µm/s. We anticipate that our technology will pave the way to elucidate functional connectivity and neurological diseases associated with transsynaptic propagation.

A CMOS Microelectrode Array System With Reconfigurable Sub-Array Multiplexing Architecture Integrating 24,320 Electrodes and 380 Readout Channels.

This article presents a CMOS microelectrode array (MEA) system with a reconfigurable sub-array multiplexing architecture using the time-division multiplexing (TDM) technique. The system consists of 24,320 TiN electrodes with 17.7 µm-pitch pixels and 380 column-parallel readout channels including a low-noise amplifier, a programmable gain amplifier, and a 10-b successive approximation register analog to digital converter. Readout channels are placed outside the pixel for high spatial resolution, and a flexible structure to acquire neural signals from electrodes selected by configuring in-pixel memory is realized. In this structure, a single channel can handle 8 to 32 electrodes, guaranteeing a temporal resolution from 5 kS/s to 20 kS/s for each electrode. A 128 × 190 MEA system was fabricated in a 110-nm CMOS process, and each readout channel consumes 81 µW at 1.5-V supply voltage featuring input-referred noise of 1.48 µVrms without multiplexing and 5.4 µVrms with multiplexing at the action-potential band (300 Hz-10 kHz).

Neural probe system for behavioral neuropharmacology by bi-directional wireless drug delivery and electrophysiology in socially interacting mice.

Assessing the neurological and behavioral effects of drugs is important in developing pharmacological treatments, as well as understanding the mechanisms associated with neurological disorders. Herein, we present a miniaturized, wireless neural probe system with the capability of delivering drugs for the real-time investigation of the effects of the drugs on both behavioral and neural activities in socially interacting mice. We demonstrate wireless drug delivery and simultaneous monitoring of the resulting neural, behavioral changes, as well as the dose-dependent and repeatable responses to drugs. Furthermore, in pairs of mice, we use a food competition assay in which social interaction was modulated by the delivery of the drug, and the resulting changes in their neural activities are analyzed. During modulated food competition by drug injection, we observe changes in neural activity in mPFC region of a participating mouse over time. Our system may provide new opportunities for the development of studying the effects of drugs on behaviour and neural activity.

SYNCRIP controls miR-137 and striatal learning in animal models of methamphetamine abstinence.

Abstinence from prolonged psychostimulant use prompts stimulant withdrawal syndrome. Molecular adaptations within the dorsal striatum have been considered the main hallmark of stimulant abstinence. Here we explored striatal miRNA-target interaction and its impact on circulating miRNA marker as well as behavioral dysfunctions in methamphetamine (MA) abstinence. We conducted miRNA sequencing and profiling in the nonhuman primate model of MA abstinence, followed by miRNA qPCR, LC-MS/MS proteomics, immunoassays, and behavior tests in mice. In nonhuman primates, MA abstinence triggered a lasting upregulation of miR-137 in the dorsal striatum but a simultaneous downregulation of circulating miR-137. In mice, aberrant increase in striatal miR-137-dependent inhibition of SYNCRIP essentially mediated the MA abstinence-induced reduction of circulating miR-137. Pathway modeling through experimental deduction illustrated that the MA abstinence-mediated downregulation of circulating miR-137 was caused by reduction of SYNCRIP-dependent miRNA sorting into the exosomes in the dorsal striatum. Furthermore, diminished SYNCRIP in the dorsal striatum was necessary for MA abstinence-induced behavioral bias towards egocentric spatial learning. Taken together, our data revealed circulating miR-137 as a potential blood-based marker that could reflect MA abstinence-dependent changes in striatal miR-137/SYNCRIP axis, and striatal SYNCRIP as a potential therapeutic target for striatum-associated cognitive dysfunction by MA withdrawal syndrome.

Astrocytic urea cycle detoxifies Aß-derived ammonia while impairing memory in Alzheimer's disease.

Alzheimer's disease (AD) is one of the foremost neurodegenerative diseases, characterized by beta-amyloid (Aß) plaques and significant progressive memory loss. In AD, astrocytes are proposed to take up and clear Aß plaques. However, how Aß induces pathogenesis and memory impairment in AD remains elusive. We report that normal astrocytes show non-cyclic urea metabolism, whereas Aß-treated astrocytes show switched-on urea cycle with upregulated enzymes and accumulated entering-metabolite aspartate, starting-substrate ammonia, end-product urea, and side-product putrescine. Gene silencing of astrocytic ornithine decarboxylase-1 (ODC1), facilitating ornithine-to-putrescine conversion, boosts urea cycle and eliminates aberrant putrescine and its toxic byproducts ammonia and H2O2 and its end product GABA to recover from reactive astrogliosis and memory impairment in AD. Our findings implicate that astrocytic urea cycle exerts opposing roles of beneficial Aß detoxification and detrimental memory impairment in AD. We propose ODC1 inhibition as a promising therapeutic strategy for AD to facilitate removal of toxic molecules and prevent memory loss.

Cell-line dependency in cerebral organoid induction: cautionary observations in Alzheimer's disease patient-derived induced pluripotent stem cells.

The cerebral organoid (CO) model has been used in the study of various neurodegenerative diseases owing to its physiological implications. However, the CO model may only be representative of certain clinical findings in affected patients, while some features are not recapitulated. In this study, we found that neurons in the CO model from patients with Alzheimer's disease were less responsive to depolarization, in contrast to previous reports. This difference may be partly attributed to the variations in brain spatial identity depending on the genetic background of the induced pluripotent stem cells. Our current observation raises concerns that the phenotypes observed in the CO model need to be carefully evaluated for their clinical implications.

Production of human spinal-cord organoids recapitulating neural-tube morphogenesis.

Human spinal-cord-like tissues induced from human pluripotent stem cells are typically insufficiently mature and do not mimic the morphological features of neurulation. Here, we report a three-dimensional culture system and protocol for the production of human spinal-cord-like organoids (hSCOs) recapitulating the neurulation-like tube-forming morphogenesis of the early spinal cord. The hSCOs exhibited neurulation-like tube-forming morphogenesis, cellular differentiation into the major types of spinal-cord neurons as well as glial cells, and mature synaptic functional activities, among other features of the development of the spinal cord. We used the hSCOs to screen for antiepileptic drugs that can cause neural-tube defects. hSCOs may also facilitate the study of the development of the human spinal cord and the modelling of diseases associated with neural-tube defects.

Interference-free, lightweight wireless neural probe system for investigating brain activity during natural competition.

Competition is one of the most fundamental, yet complex, conflicts between social animals, and previous studies have indicated that the medial prefrontal cortex (mPFC) region of a brain is involved in social interactions. However, because we do not have a lightweight, wireless recording system that is free of interference, it is still unclear how the neural activity of the mPFC region is involved in the diverse, interacting behaviors that comprise competition. Herein, we present an interference-free, lightweight, wireless neural probe system that we applied to two mice to measure mPFC neural activities during a food competition test. In the test, we categorized 18 behavioral repertoires expressed by the mice. From the analysis of the neural signals during each repetition of the test, we found that the mPFC neural activity had the most positive correlation with goal-driven competitive behaviors, such as guarding resources and behaviors related to the extortion of resources. Remarkably, we found that the neural activity associated with guarding behavior was higher than that of extorting behavior, and this highlighted the importance of resource-guarding behavior for winning the competition, i.e., 'winning a trophy is hard, but keeping it is harder'. Our approach in which a wireless system is used will enable in-depth studies of the brains of mice in their natural social interactions.

Multiplex SNP Genotyping Using SWITCH: Sequence-Specific Nanoparticle with Interpretative Toehold-Mediated Sequence Decoding in Hydrogel.

Single nucleotide polymorphisms (SNPs) that can alter phenotypes of individuals play a pivotal role in disease development and, more importantly, responses to therapy. However, SNP genotyping has been challenging due to the similarity of SNP alleles and their low concentration in biological samples. Sequence-specific nanoparticle with interpretative toehold-mediated sequence decoding in hydrogel (SWITCH) for multiplex SNP genotyping is presented. The encoding with gold nanoparticle probes transduces each SNP target to ≈1000 invaders with prominently different sequences between wild and mutant types, featuring polymerase chain reaction (PCR)-free amplification. Subsequently, the toehold-mediated DNA replacement in hydrogel microparticles decodes the invaders via SNP-specific fluorescence signals. The 4-plex detection of the warfarin-associated SNP targets spiked in commercially validated human serum (S1-100ML, Merck) is successfully demonstrated with excellent specificity. This work is the first technology development presenting PCR-free, multiplex SNP genotyping with a single reporting fluorophore, to the best of knowledge.

A Multimodal Multi-Shank Fluorescence Neural Probe for Cell-Type-Specific Electrophysiology in Multiple Regions across a Neural Circuit.

Cell-type-specific, activity-dependent electrophysiology can allow in-depth analysis of functional connectivity inside complex neural circuits composed of various cell types. To date, optics-based fluorescence recording devices enable monitoring cell-type-specific activities. However, the monitoring is typically limited to a single brain region, and the temporal resolution is significantly low. Herein, a multimodal multi-shank fluorescence neural probe that allows cell-type-specific electrophysiology from multiple deep-brain regions at a high spatiotemporal resolution is presented. A photodiode and an electrode-array pair are monolithically integrated on each tip of a minimal-form-factor silicon device. Both fluorescence and electrical signals are successfully measured simultaneously in GCaMP6f expressing mice, and the cell type from sorted neural spikes is identified. The probe's capability of combined electro-optical recordings for cell-type-specific electrophysiology at multiple brain regions within a neural circuit is demonstrated. The new experimental paradigm to enable the precise investigation of functional connectivity inside and across complex neural circuits composed of various cell types is expected.

Interpreting the Entire Connectivity of Individual Neurons in Micropatterned Neural Culture With an Integrated Connectome Analyzer of a Neuronal Network (iCANN).

The function of a neural circuit can be determined by the following: (1) characteristics of individual neurons composing the circuit, (2) their distinct connection structure, and (3) their neural circuit activity. However, prior research on correlations between these three factors revealed many limitations. In particular, profiling and modeling of the connectivity of complex neural circuits at the cellular level are highly challenging. To reduce the burden of the analysis, we suggest a new approach with simplification of the neural connection in an array of honeycomb patterns on 2D, using a microcontact printing technique. Through a series of guided neuronal growths in defined honeycomb patterns, a simplified neuronal circuit was achieved. Our approach allowed us to obtain the whole network connectivity at cellular resolution using a combination of stochastic multicolor labeling via viral transfection. Therefore, we were able to identify several types of hub neurons with distinct connectivity features. We also compared the structural differences between different circuits using three-node motif analysis. This new model system, iCANN, is the first experimental model of neural computation at the cellular level, providing neuronal circuit structures for the study of the relationship between anatomical structure and function of the neuronal network.

Effect of carbon nanomaterial dimension on the functional activity and degeneration of neurons.

Despite growing concerns regarding the threat of airborne nanoparticle-mediated brain degeneration, the underlying pathological mechanisms remain unclear. Carbon nanomaterials, the main components of airborne nanoparticles, have multi-dimensional structures. Therefore, the dimensional effect of carbon-based nanomaterials on the regulation of neural function in brain disorders requires additional clarification. Herein, we report the interaction between zero-to three-dimensional carbon nanostructures and the amyloid-beta protein, which can either activate or interrupt neuronal functions, depending on the dimension of the carbon nanostructures. The carbon nanomaterials induced significant cellular activation by short-term exposure, while prolonged exposure eventually caused neuronal cell death. Such dimension-dependent activation or degeneration was more evident in the higher-dimension carbon nanomaterials, as confirmed by the increases in neurotransmitter secretion and synapse-related protein levels to more than five times at 72 h of monitoring and calcium signaling in the neurons. The inclusion of amyloid-beta proteins ameliorated the cytotoxic effects of carbon nanomaterials in higher-dimensional carbon nanomaterials by regulating 333 genes. We found that the ɑ-synuclein gene is the key factor in carbon-induced abnormal neuronal function. Therefore, through biological analyses and in vitro feasibility studies, this new insight may contribute toward understanding the pathological mechanism and finding a new target for therapy in human brain pathologies.

Excitatory synapses and gap junctions cooperate to improve Pv neuronal burst firing and cortical social cognition in Shank2-mutant mice.

NMDA receptor (NMDAR) and GABA neuronal dysfunctions are observed in animal models of autism spectrum disorders, but how these dysfunctions impair social cognition and behavior remains unclear. We report here that NMDARs in cortical parvalbumin (Pv)-positive interneurons cooperate with gap junctions to promote high-frequency (>80 Hz) Pv neuronal burst firing and social cognition. Shank2-/- mice, displaying improved sociability upon NMDAR activation, show impaired cortical social representation and inhibitory neuronal burst firing. Cortical Shank2-/- Pv neurons show decreased NMDAR activity, which suppresses the cooperation between NMDARs and gap junctions (GJs) for normal burst firing. Shank2-/- Pv neurons show compensatory increases in GJ activity that are not sufficient for social rescue. However, optogenetic boosting of Pv neuronal bursts, requiring GJs, rescues cortical social cognition in Shank2-/- mice, similar to the NMDAR-dependent social rescue. Therefore, NMDARs and gap junctions cooperate to promote cortical Pv neuronal bursts and social cognition.

Bimodal neural probe for highly co-localized chemical and electrical monitoring of neural activities in vivo.

Investigation of the chemical and electrical signals of cells in vivo is critical for studying functional connectivity and brain diseases. Most previous studies have observed either the electrical signals or the chemical signals of cells because recording electrical signals and neurochemicals are done by fundamentally different methods. Herein, we present a bimodal MEMS neural probe that is monolithically integrated with an array of microelectrodes for recording electrical activity, microfluidic channels for sampling extracellular fluid, and a microfluidic interface chip for multiple drug delivery and sample isolation from the localized region at the cellular level. In this work, we successfully demonstrated the functionality of our probe by monitoring and modulating bimodal (electrical and chemical) neural activities through the delivery of chemicals in a co-localized brain region in vivo. We expect our bimodal probe to provide opportunities for a variety of in-depth studies of brain functions as well as for the investigation of neural circuits related to brain diseases.