Nanoscale Vertical Resolution in Optical Printing of Inorganic Nanoparticles.

Direct optical printing of functional inorganics shows tremendous potential as it enables the creation of intricate two-dimensional (2D) patterns and affordable design and production of various devices. Although there have been recent advancements in printing processes using short-wavelength light or pulsed lasers, the precise control of the vertical thickness in printed 3D structures has received little attention. This control is vital to the diverse functionalities of inorganic thin films and their devices, as they rely heavily on their thicknesses. This lack of research is attributed to the technical intricacy and complexity involved in the lithographic processes. Herein, we present a generalized optical 3D printing process for inorganic nanoparticles using maskless digital light processing. We develop a range of photocurable inorganic nanoparticle inks encompassing metals, semiconductors, and oxides, combined with photolinkable ligands and photoacid generators, enabling the direct solidification of nanoparticles in the ink medium. Our process creates complex and large-area patterns with a vertical resolution of ∼50 nm, producing 50-nm-thick 2D films and several micrometer-thick 3D architectures with no layer height difference via layer-by-layer deposition. Through fabrication and operation of multilayered switching devices with Au electrodes and Ag-organic resistive layers, the feasibility of our process for cost-effective manufacturing of multilayered devices is demonstrated.

Systematic investigation of allelic regulatory activity of schizophrenia-associated common variants.

Genome-wide association studies (GWASs) have successfully identified 145 genomic regions that contribute to schizophrenia risk, but linkage disequilibrium makes it challenging to discern causal variants. We performed a massively parallel reporter assay (MPRA) on 5,173 fine-mapped schizophrenia GWAS variants in primary human neural progenitors and identified 439 variants with allelic regulatory effects (MPRA-positive variants). Transcription factor binding had modest predictive power, while fine-map posterior probability, enhancer overlap, and evolutionary conservation failed to predict MPRA-positive variants. Furthermore, 64% of MPRA-positive variants did not exhibit expressive quantitative trait loci signature, suggesting that MPRA could identify yet unexplored variants with regulatory potentials. To predict the combinatorial effect of MPRA-positive variants on gene regulation, we propose an accessibility-by-contact model that combines MPRA-measured allelic activity with neuronal chromatin architecture.

Generalised optical printing of photocurable metal chalcogenides.

Optical three-dimensional (3D) printing techniques have attracted tremendous attention owing to their applicability to mask-less additive manufacturing, which enables the cost-effective and straightforward creation of patterned architectures. However, despite their potential use as alternatives to traditional lithography, the printable materials obtained from these methods are strictly limited to photocurable resins, thereby restricting the functionality of the printed objects and their application areas. Herein, we report a generalised direct optical printing technique to obtain functional metal chalcogenides via digital light processing. We developed universally applicable photocurable chalcogenidometallate inks that could be directly used to create 2D patterns or micrometre-thick 2.5D architectures of various sizes and shapes. Our process is applicable to a diverse range of functional metal chalcogenides for compound semiconductors and 2D transition-metal dichalcogenides. We then demonstrated the feasibility of our technique by fabricating and evaluating a micro-scale thermoelectric generator bearing tens of patterned semiconductors. Our approach shows potential for simple and cost-effective architecturing of functional inorganic materials.

Detection method for transparent window cleaning device, image processing approach.

Recent years, there has been an increase in the number of high-rise buildings, and subsequently, the interest in external wall cleaning methods has similarly increased. While a number of exterior wall cleaning robots are being developed, a method to detect contaminants on the exterior walls is still required. The exteriors of most high-rise buildings today take the form of a window curtain-wall made of translucent glass. Detecting dust on translucent glass is a significant challenge. Here, we have attempted to overcome this challenge using image processing, inspired by the fact that people typically use just the 'naked eye' to recognize dust on windows. In this paper, we propose a method that detects dust through simple image processing techniques and estimates its density. This method only uses processing techniques that are not significantly restricted by global brightness and background, making it easily applicable in outdoor conditions. Dust separation was performed using a median filter, and dust density was estimated through a mean shift analysis technique. This dust detection method can perform dust separation and density estimation using only an image of the dust on a translucent window with blurry background.

FosGFP expression does not capture a sensory learning-related engram in superficial layers of mouse barrel cortex.

Immediate-early gene (IEG) expression has been used to identify small neural ensembles linked to a particular experience, based on the principle that a selective subset of activated neurons will encode specific memories or behavioral responses. The majority of these studies have focused on "engrams" in higher-order brain areas where more abstract or convergent sensory information is represented, such as the hippocampus, prefrontal cortex, or amygdala. In primary sensory cortex, IEG expression can label neurons that are responsive to specific sensory stimuli, but experience-dependent shaping of neural ensembles marked by IEG expression has not been demonstrated. Here, we use a fosGFP transgenic mouse to longitudinally monitor in vivo expression of the activity-dependent gene c-fos in superficial layers (L2/3) of primary somatosensory cortex (S1) during a whisker-dependent learning task. We find that sensory association training does not detectably alter fosGFP expression in L2/3 neurons. Although training broadly enhances thalamocortical synaptic strength in pyramidal neurons, we find that synapses onto fosGFP+ neurons are not selectively increased by training; rather, synaptic strengthening is concentrated in fosGFP- neurons. Taken together, these data indicate that expression of the IEG reporter fosGFP does not facilitate identification of a learning-specific engram in L2/3 in barrel cortex during whisker-dependent sensory association learning.

Programmable multilayer printing of a mechanically-tunable 3D hydrogel co-culture system for high-throughput investigation of complex cellular behavior.

Hydrogels are widely used as a 3D cell culture platform, as they can be tailored to provide suitable microenvironments to induce cellular phenotypes with physiological significance. Hydrogels are especially deemed attractive as a co-culture platform, in which two or more different types of cells are cultured together in close proximity, since the spatial distribution of different cell types can be rendered possible by advanced microfabrication schemes. Herein, programmable multilayer photolithography is employed to develop a 3D hydrogel-based co-culture system in an efficient and scalable manner, which consists of an inner microgel array containing one cell type covered by an outer hydrogel overlay containing another cell type. In particular, the mechanical properties of microgel array and hydrogel overlay are independently controlled in a wide range, with elastic moduli ranging from 1.7 to 31.6 kPa, allowing the high-throughput investigation of both individual hydrogel mechanics and mechanical gradients generated at their interface. Utilizing this system, phenotypical changes (i.e. proliferation, spheroid formation and Mφ polarization) of macrophages encapsulated in microgel array, in response to complex mechanical microenvironment and co-cultured fibroblasts, are comprehensively explored.

Dynamic multimodal holograms of conjugated organogels via dithering mask lithography.

Polymeric materials have been used to realize optical systems that, through periodic variations of their structural or optical properties, interact with light-generating holographic signals. Complex holographic systems can also be dynamically controlled through exposure to external stimuli, yet they usually contain only a single type of holographic mode. Here, we report a conjugated organogel that reversibly displays three modes of holograms in a single architecture. Using dithering mask lithography, we realized two-dimensional patterns with varying cross-linking densities on a conjugated polydiacetylene. In protic solvents, the organogel contracts anisotropically to develop optical and structural heterogeneities along the third dimension, displaying holograms in the form of three-dimensional full parallax signals, both in fluorescence and bright-field microscopy imaging. In aprotic solvents, these heterogeneities diminish as organogels expand, recovering the two-dimensional periodicity to display a third hologram mode based on iridescent structural colours. Our study presents a next-generation hologram manufacturing method for multilevel encryption technologies.

An automated homecage system for multiwhisker detection and discrimination learning in mice.

Automated, homecage behavioral training for rodents has many advantages: it is low stress, requires little interaction with the experimenter, and can be easily manipulated to adapt to different experimental conditions. We have developed an inexpensive, Arduino-based, homecage training apparatus for sensory association training in freely-moving mice using multiwhisker air current stimulation coupled to a water reward. Animals learn this task readily, within 1-2 days of training, and performance progressively improves with training. We examined the parameters that regulate task acquisition using different stimulus intensities, directions, and reward valence. Learning was assessed by comparing anticipatory licking for the stimulus compared to the no-stimulus (blank) trials. At high stimulus intensities (>9 psi), animals showed markedly less participation in the task. Conversely, very weak air current intensities (1-2 psi) were not sufficient to generate rapid learning behavior. At intermediate stimulus intensities (5-6 psi), a majority of mice learned that the multiwhisker stimulus predicted the water reward after 24-48 hrs of training. Both exposure to isoflurane and lack of whiskers decreased animals' ability to learn the task. Following training at an intermediate stimulus intensity, mice were able to transfer learning behavior when exposed to a lower stimulus intensity, an indicator of perceptual learning. Mice learned to discriminate between two directions of stimulation rapidly and accurately, even when the angular distance between the stimuli was <15 degrees. Switching the reward to a more desirable reward, aspartame, had little effect on learning trajectory. Our results show that a tactile association task in an automated homecage environment can be monitored by anticipatory licking to reveal rapid and progressive behavioral change. These Arduino-based, automated mouse cages enable high-throughput training that facilitate analysis of large numbers of genetically modified mice with targeted manipulations of neural activity.

Multi-Color Luminescence Transition of Upconversion Nanocrystals via Crystal Phase Control with SiO2 for High Temperature Thermal Labels.

Upconversion nanocrystals (UCNs)-embedded microarchitectures with luminescence color transition capability and enhanced luminescence intensity under extreme conditions are suitable for developing a robust labeling system in a high-temperature thermal industrial process. However, most UCNs based labeling systems are limited by the loss of luminescence owing to the destruction of the crystalline phase or by a predetermined luminescence color without color transition capability. Herein, an unusual crystal phase transition of UCNs to a hexagonal apatite phase in the presence of SiO2 nanoparticles is reported with the enhancements of 130-fold green luminescence and 52-fold luminance as compared to that of the SiO2-free counterpart. By rationally combining this strategy with an additive color mixing method using a mask-less flow lithography technique, single to multiple luminescence color transition, scalable labeling systems with hidden letters-, and multi-luminescence colored microparticles are demonstrated for a UCNs luminescence color change-based high temperature labeling system.

Hydrogel-Based Colorimetric Assay for Multiplexed MicroRNA Detection in a Microfluidic Device.

Although microRNA (miRNA) expression levels provide important information regarding disease states owing to their unique dysregulation patterns in tissues, translation of miRNA diagnostics into point-of-care (POC) settings has been limited by practical challenges. Here, we developed a hydrogel-based microfluidic platform for colorimetric profiling of miRNAs, without the use of complex external equipment for fluidics and imaging. For sensitive and reliable measurement without the risk of sequence bias, we employed a gold deposition-based signal amplification scheme and dark-field imaging, and seamlessly integrated a previously developed miRNA assay scheme into this platform. The assay demonstrated a limit of detection of 260 fM, along with multiplexing of small panels of miRNAs in healthy and cancer samples. We anticipate this versatile platform to facilitate a broad range of POC profiling of miRNAs in cancer-associated dysregulation with high-confidence by exploiting the unique features of hydrogel substrate in an on-chip format and colorimetric analysis.

Highly Stable Upconverting Nanocrystal-Polydiacetylenes Nanoplates for Orthogonal Dual Signaling-Based Detection of Cyanide.

Although the unique optical signaling properties of polydiacetylene (PDA) have been exploited in diverse bio-chemosensors, the practical application of most PDA sensor systems is limited by their instability in harsh environments and fluorescence signal weakness. Herein, a universal design principle for a highly stable PDA sensor system with a practical dual signaling capability is developed to detect cyanide (CN) ions, which are commonly found in drinking water. Effective metal intercalation and enhanced hydrophobic intermolecular interactions between PDA-metal supramolecules are used to construct highly stacked PDA-metal nanoplates that feature unusual optical stability upon exposure to strong acids, bases, organic solvents, and thermal/mechanical stresses, and can selectively detect CN anions, concomitantly undergoing a specific supramolecular structure change. To realize the practical dual signaling capability of the PDA sensor system, upconverting nanocrystals (UCNs) are incorporated into highly stacked PDA-metal nanoplates, and practical dual signaling (orthogonal changes in luminescence and visible color) is demonstrated using a portable detection system. The presented universal design principle is expected to be suitable for the development of other highly stable and selective PDA sensor systems with practical dual signaling capability.

Recyclable High-Performance Polymer Electrolyte Based on a Modified Methyl Cellulose-Lithium Trifluoromethanesulfonate Salt Composite for Sustainable Energy Systems.

Although energy-storage devices based on Li ions are considered as the most prominent candidates for immediate application in the near future, concerns with regard to their stability, safety, and environmental impact still remain. As a solution, the development of all-solid-state energy-storage devices with enhanced stability is proposed. A new eco-friendly polymer electrolyte has been synthesized by incorporating lithium trifluoromethanesulfonate into chemically modified methyl cellulose (LiTFS-LiSMC). The transparent and flexible electrolyte exhibits a good conductivity of near 1 mS cm-1 . An all-solid-state supercapacitor fabricated from 20 wt % LiTFS-LiSMC shows comparable specific capacitances to a standard liquid-electrolyte supercapacitor and an excellent stability even after 20 000 charge-discharge cycles. The electrolyte is also compatible with patterned carbon, which enables the simple fabrication of micro-supercapacitors. In addition, the LiTFS-LiSMC electrolyte can be recycled and reused more than 20 times with negligible change in its performance. Thus, it is a promising material for sustainable energy-storage devices.

Shank3 Exons 14-16 Deletion in Glutamatergic Neurons Leads to Social and Repetitive Behavioral Deficits Associated With Increased Cortical Layer 2/3 Neuronal Excitability.

Shank3, an abundant excitatory postsynaptic scaffolding protein, has been associated with multiple brain disorders, including autism spectrum disorders (ASD) and Phelan-McDermid syndrome (PMS). However, how cell type-specific Shank3 deletion affects disease-related neuronal and brain functions remains largely unclear. Here, we investigated the impacts of Shank3 deletion in glutamatergic neurons on synaptic and behavioral phenotypes in mice and compared results with those previously obtained from mice with global Shank3 mutation and GABAergic neuron-specific Shank3 mutation. Neuronal excitability was abnormally increased in layer 2/3 pyramidal neurons in the medial prefrontal cortex (mPFC) in mice with a glutamatergic Shank3 deletion, similar to results obtained in mice with a global Shank3 deletion. In addition, excitatory synaptic transmission was abnormally increased in layer 2/3 neurons in mice with a global, but not a glutamatergic, Shank3 deletion, suggesting that Shank3 in glutamatergic neurons are important for the increased neuronal excitability, but not for the increased excitatory synaptic transmission. Neither excitatory nor inhibitory synaptic transmission was altered in the dorsal striatum of Shank3-deficient glutamatergic neurons, a finding that contrasts with the decreased excitatory synaptic transmission in global and Shank3-deficient GABAergic neurons. Behaviorally, glutamatergic Shank3-deficient mice displayed abnormally increased direct social interaction and repetitive self-grooming, similar to global and GABAergic Shank3-deficient mice. These results suggest that glutamatergic and GABAergic Shank3 deletions lead to distinct synaptic and neuronal changes in cortical layer 2/3 and dorsal striatal neurons, but cause similar social and repetitive behavioral abnormalities likely through distinct mechanisms.

Fluorescence-Based Quantitative Synapse Analysis for Cell Type-Specific Connectomics.

Anatomical methods for determining cell type-specific connectivity are essential to inspire and constrain our understanding of neural circuit function. We developed genetically-encoded reagents for fluorescence-synapse labeling and connectivity analysis in brain tissue, using a fluorogen-activating protein (FAP)-coupled or YFP-coupled, postsynaptically-localized neuroligin-1 (NL-1) targeting sequence (FAP/YFPpost). FAPpost expression did not alter mEPSC or mIPSC properties. Sparse AAV-mediated expression of FAP/YFPpost with the cell-filling, red fluorophore dTomato (dTom) enabled high-throughput, compartment-specific detection of putative synapses across diverse neuron types in mouse somatosensory cortex. We took advantage of the bright, far-red emission of FAPpost puncta for multichannel fluorescence alignment of dendrites, FAPpost puncta, and presynaptic neurites in transgenic mice with saturated labeling of parvalbumin (PV), somatostatin (SST), or vasoactive intestinal peptide (VIP)-expressing neurons using Cre-reporter driven expression of YFP. Subtype-specific inhibitory connectivity onto layer 2/3 (L2/3) neocortical pyramidal (Pyr) neurons was assessed using automated puncta detection and neurite apposition. Quantitative and compartment-specific comparisons show that PV inputs are the predominant source of inhibition at both the soma and the dendrites and were particularly concentrated at the primary apical dendrite. SST inputs were interleaved with PV inputs at all secondary-order and higher-order dendritic branches. These fluorescence-based synapse labeling reagents can facilitate large-scale and cell-type specific quantitation of changes in synaptic connectivity across development, learning, and disease states.

Shank3 Mice Carrying the Human Q321R Mutation Display Enhanced Self-Grooming, Abnormal Electroencephalogram Patterns, and Suppressed Neuronal Excitability and Seizure Susceptibility.

Shank3, a postsynaptic scaffolding protein involved in regulating excitatory synapse assembly and function, has been implicated in several brain disorders, including autism spectrum disorders (ASD), Phelan-McDermid syndrome, schizophrenia, intellectual disability, and mania. Here we generated and characterized a Shank3 knock-in mouse line carrying the Q321R mutation (Shank3 Q321R mice) identified in a human individual with ASD that affects the ankyrin repeat region (ARR) domain of the Shank3 protein. Homozygous Shank3 Q321R/Q321R mice show a selective decrease in the level of Shank3a, an ARR-containing protein variant, but not other variants. CA1 pyramidal neurons in the Shank3 Q321R/Q321R hippocampus show decreased neuronal excitability but normal excitatory and inhibitory synaptic transmission. Behaviorally, Shank3 Q321R/Q321R mice show moderately enhanced self-grooming and anxiolytic-like behavior, but normal locomotion, social interaction, and object recognition and contextual fear memory. In addition, these mice show abnormal electroencephalogram (EEG) patterns and decreased susceptibility to induced seizures. These results indicate that the Q321R mutation alters Shank3 protein stability, neuronal excitability, repetitive and anxiety-like behavior, EEG patterns, and seizure susceptibility in mice.

Early Correction of N-Methyl-D-Aspartate Receptor Function Improves Autistic-like Social Behaviors in Adult Shank2-/- Mice.

BACKGROUND: Autism spectrum disorder involves neurodevelopmental dysregulations that lead to visible symptoms at early stages of life. Many autism spectrum disorder-related mechanisms suggested by animal studies are supported by demonstrated improvement in autistic-like phenotypes in adult animals following experimental reversal of dysregulated mechanisms. However, whether such mechanisms also act at earlier stages to cause autistic-like phenotypes is unclear. METHODS: We used Shank2-/- mice carrying a mutation identified in human autism spectrum disorder (exons 6 and 7 deletion) and combined electrophysiological and behavioral analyses to see whether early pathophysiology at pup stages is different from late pathophysiology at juvenile and adult stages and whether correcting early pathophysiology can normalize late pathophysiology and abnormal behaviors in juvenile and adult mice. RESULTS: Early correction of a dysregulated mechanism in young mice prevents manifestation of autistic-like social behaviors in adult mice. Shank2-/- mice, known to display N-methyl-D-aspartate receptor (NMDAR) hypofunction and autistic-like behaviors at postweaning stages after postnatal day 21 (P21), show the opposite synaptic phenotype-NMDAR hyperfunction-at an earlier preweaning stage (∼P14). Moreover, this NMDAR hyperfunction at P14 rapidly shifts to NMDAR hypofunction after weaning (∼P24). Chronic suppression of the early NMDAR hyperfunction by the NMDAR antagonist memantine (P7-P21) prevents NMDAR hypofunction and autistic-like social behaviors from manifesting at later stages (∼P28 and P56). CONCLUSIONS: Early NMDAR hyperfunction leads to late NMDAR hypofunction and autistic-like social behaviors in Shank2-/- mice, and early correction of NMDAR dysfunction has the long-lasting effect of preventing autistic-like social behaviors from developing at later stages.

GABA Neuronal Deletion of Shank3 Exons 14-16 in Mice Suppresses Striatal Excitatory Synaptic Input and Induces Social and Locomotor Abnormalities.

Shank3 is an excitatory postsynaptic scaffolding protein implicated in multiple brain disorders, including autism spectrum disorders (ASD) and Phelan-McDermid syndrome (PMS). Although previous neurobiological studies on Shank3 and Shank3-mutant mice have revealed diverse roles of Shank3 in the regulation of synaptic, neuronal and brain functions, whether Shank3 expression in specific cell types distinctly contributes to mouse phenotypes remains largely unclear. In the present study, we generated two Shank3-mutant mouse lines (exons 14-16) carrying global and GABA neuron-specific deletions and characterized their electrophysiological and behavioral phenotypes. These mouse lines show similar decreases in excitatory synaptic input onto dorsolateral striatal neurons. In addition, the abnormal social and locomotor behaviors observed in global Shank3-mutant mice are strongly mimicked by GABA neuron-specific Shank3-mutant mice, whereas the repetitive and anxiety-like behaviors are only partially mimicked. These results suggest that GABAergic Shank3 (exons 14-16) deletion has strong influences on striatal excitatory synaptic transmission and social and locomotor behaviors in mice.

Sexually dimorphic behavior, neuronal activity, and gene expression in Chd8-mutant mice.

Autism spectrum disorders (ASDs) are four times more common in males than in females, but the underlying mechanisms are poorly understood. We characterized sexually dimorphic changes in mice carrying a heterozygous mutation in Chd8 (Chd8+/N2373K) that was first identified in human CHD8 (Asn2373LysfsX2), a strong ASD-risk gene that encodes a chromatin remodeler. Notably, although male mutant mice displayed a range of abnormal behaviors during pup, juvenile, and adult stages, including enhanced mother-seeking ultrasonic vocalization, enhanced attachment to reunited mothers, and isolation-induced self-grooming, their female counterparts do not. This behavioral divergence was associated with sexually dimorphic changes in neuronal activity, synaptic transmission, and transcriptomic profiles. Specifically, female mice displayed suppressed baseline neuronal excitation, enhanced inhibitory synaptic transmission and neuronal firing, and increased expression of genes associated with extracellular vesicles and the extracellular matrix. Our results suggest that a human CHD8 mutation leads to sexually dimorphic changes ranging from transcription to behavior in mice.

Rapid Light-Driven Color Transition of Novel Photoresponsive Polydiacetylene Molecules.

We developed new photoresponsive polydiacetylene (PR-PDA) molecules by incorporating a photocleavable moiety, 6-nitropiperonyl alcohol (NP) or 4,5-dimethoxy-2-nitrobenzyl alcohol (DMN), into a self-assembling diacetylene molecule. Inducing steric disordering of the assembled PDA molecules by the cleavage of the photoresponsive moiety under 365 nm UV irradiation results in color transition from blue to red and development of red fluorescence, allowing convenient photo patterning. Further writing and erasing of fluorescence patterns are demonstrated toward novel secure information communication and anticounterfeiting applications.

Modulatory Functionalization of Gold Nanorods Using Supramolecular Assemblies.

Supramolecular-assembly-mediated functionalization of gold nanorods (GNRs) has been developed by reversible phase transfer between water and oils, which offers a facile method for fabricating robust GNRs with surface-charge tunability. In this regard, trimethylammonium (TMA) GNRs were initially prepared from conventional cetyltrimethylammonium bromide (CTAB) GNRs by means of a ligand-exchange reaction in the presence of an excess amount of TMA ligands. To further expand their functionality and potential applications, electrostatic assemblies of positively charged TMA-GNRs with negatively charged oleate ions were prepared. These assemblies (OA-GNRs) can undergo facile phase transfer from water to hexane. Interestingly, the reversible electrostatic assembly between the TMA and OA ions fabricated onto GNRs can be easily disrupted by treatment with HCl, which removes the OA ions from the GNRs to re-form the TMA-GNRs, which can be made soluble in aqueous media again. In addition, OA-GNRs can be further used for the synthesis of negatively charged GNRs such as 11-mercaptoundecanoic acid (MUA) GNRs, which are hard to prepare directly from CTAB-GNRs. This versatile method for phase transfer and functionalization on GNRs is expected to broaden the scope of their applications in sensing, biomedical imaging, photothermal therapies, and drug delivery systems.