NIN-LIKE PROTEIN3.2 inhibits repressor Aux/IAA14 expression and enhances root biomass in maize seedlings under low nitrogen.

Plants generally enhance their root growth in the form of greater biomass and/or root length to boost nutrient uptake in response to short-term low nitrogen (LN). However, the underlying mechanisms of short-term LN-mediated root growth remain largely elusive. Our genome-wide association study, haplotype analysis, and phenotyping of transgenic plants showed that the crucial nitrate signaling component NIN-LIKE PROTEIN3.2 (ZmNLP3.2), a positive regulator of root biomass, is associated with natural variations in root biomass of maize (Zea mays L.) seedlings under LN. The monocot-specific gene AUXIN/INDOLE-3-ACETIC ACID14 (ZmAux/IAA14) exhibited opposite expression patterns to ZmNLP3.2 in ZmNLP3.2 knockout and overexpression lines, suggesting that ZmNLP3.2 hampers ZmAux/IAA14 transcription. Importantly, ZmAux/IAA14 knockout seedlings showed a greater root dry weight (RDW), whereas ZmAux/IAA14 overexpression reduced RDW under LN compared with wild-type plants, indicating that ZmAux/IAA14 negatively regulates the RDW of LN-grown seedlings. Moreover, in vitro and vivo assays indicated that AUXIN RESPONSE FACTOR19 (ZmARF19) binds to and transcriptionally activates ZmAux/IAA14, which was weakened by the ZmNLP3.2-ZmARF19 interaction. The zmnlp3.2 ZmAux/IAA14-OE seedlings exhibited further reduced RDW compared to ZmAux/IAA14 overexpression lines when subjected to LN treatment, corroborating the ZmNLP3.2-ZmAux/IAA14 interaction. Thus, our study reveals a ZmNLP3.2-ZmARF19-ZmAux/IAA14 module regulating root biomass in response to nitrogen limitation in maize.

Acute cannabinoids impair working memory through astroglial CB1 receptor modulation of hippocampal LTD.

Impairment of working memory is one of the most important deleterious effects of marijuana intoxication in humans, but its underlying mechanisms are presently unknown. Here, we demonstrate that the impairment of spatial working memory (SWM) and in vivo long-term depression (LTD) of synaptic strength at hippocampal CA3-CA1 synapses, induced by an acute exposure of exogenous cannabinoids, is fully abolished in conditional mutant mice lacking type-1 cannabinoid receptors (CB(1)R) in brain astroglial cells but is conserved in mice lacking CB(1)R in glutamatergic or GABAergic neurons. Blockade of neuronal glutamate N-methyl-D-aspartate receptors (NMDAR) and of synaptic trafficking of glutamate α-amino-3-hydroxy-5-methyl-isoxazole propionic acid receptors (AMPAR) also abolishes cannabinoid effects on SWM and LTD induction and expression. We conclude that the impairment of working memory by marijuana and cannabinoids is due to the activation of astroglial CB(1)R and is associated with astroglia-dependent hippocampal LTD in vivo.

Copolymerization Involving Sulfur-Containing Monomers.

Incorporating sulfur (S) atoms into polymer main chains endows these materials with many attractive features, including a high refractive index, mechanical properties, electrochemical properties, and adhesive ability to heavy metal ions. The copolymerization involving S-containing monomers constitutes a facile method for effectively constructing S-containing polymers with diverse structures, readily tunable sequences, and topological structures. In this review, we describe the recent advances in the synthesis of S-containing polymers via copolymerization or multicomponent polymerization techniques concerning a variety of S-containing monomers, such as dithiols, carbon disulfide, carbonyl sulfide, cyclic thioanhydrides, episulfides and elemental sulfur (S8). Particularly, significant focus is paid to precise control of the main-chain sequence, stereochemistry, and topological structure for achieving high-value applications.

Distinct processing of the state prediction error signals in frontal and parietal correlates in learning the environment model.

Goal-directed reinforcement learning constructs a model of how the states in the environment are connected and prospectively evaluates action values by simulating experience. State prediction error (SPE) is theorized as a crucial signal for learning the environment model. However, the underlying neural mechanisms remain unclear. Here, using electroencephalogram, we verified in a two-stage Markov task two neural correlates of SPEs: an early negative correlate transferring from frontal to central electrodes and a late positive correlate over parietal regions. Furthermore, by investigating the effects of explicit knowledge about the environment model and rewards in the environment, we found that, for the parietal correlate, rewards enhanced the representation efficiency (beta values of regression coefficient) of SPEs, whereas explicit knowledge elicited a larger SPE representation (event-related potential activity) for rare transitions. However, for the frontal and central correlates, rewards increased activities in a content-independent way and explicit knowledge enhanced activities only for common transitions. Our results suggest that the parietal correlate of SPEs is responsible for the explicit learning of state transition structure, whereas the frontal and central correlates may be involved in cognitive control. Our study provides novel evidence for distinct roles of the frontal and the parietal cortices in processing SPEs.

Aberrant activated Notch1 promotes prostate enlargement driven by androgen signaling via disrupting mitochondrial function in mouse.

The prostate is a vital accessory gonad in the mammalian male reproductive system. With the ever-increasing proportion of the population over 60 years of age worldwide, the incidence of prostate diseases, such as benign prostatic hyperplasia (BPH) and prostate cancer (PCa), is on the rise and is gradually becoming a significant medical problem globally. The notch signaling pathway is essential in regulating prostate early development. However, the potential regulatory mechanism of Notch signaling in prostatic enlargement and hyperplasia remains unclear. In this study, we proved that overactivation of Notch1 signaling in mouse prostatic epithelial cells (OEx) led to prostatic enlargement via enhancing proliferation and inhibiting apoptosis of prostatic epithelial cells. Further study showed that N1ICD/RBPJ directly up-regulated the androgen receptor (AR) and enhanced prostatic sensitivity to androgens. Hyper-proliferation was not found in orchidectomized OEx mice without androgen supply but was observed after Dihydrotestosterone (DHT) supplementation. Our data showed that the number of mitochondrion in prostatic epithelial cells of OEx mice was increased, but the mitochondrial function was impaired, and the essential activity of the mitochondrial respiratory electron transport chain was significantly weakened. Disordered mitochondrial number and metabolic function further resulted in excessive accumulation of reactive oxygen species (ROS). Importantly, anti-oxidant N-Acetyl-L-Cysteine (NAC) therapy could alleviate prostatic hyperplasia caused by the over-activation of Notch1 signaling. Furthermore, we observed the incremental Notch signaling activity in progenitor-like club cells in the scRNA-seq data set of human BPH patients. Moreover, the increased number of TROP2+ progenitors and Club cells was also confirmed in our OEx mice. In conclusion, our study revealed that over-activated Notch1 signaling induces prostatic enlargement by increasing androgen receptor sensitivity, disrupting cellular mitochondrial metabolism, increasing ROS, and a higher number of progenitor cells, all of which can be effectively rescued by NAC treatment.

Giant Carrier Mobility in a Room-Temperature Ferromagnetic VSi2N4 Monolayer.

Using density functional theory (DFT), we investigate that two possible phases of VSi2N4 (VSN) may be realized, one called the "H phase" corresponding to what is known from calculation and herein the other new "T phase" being stabilized by a biaxial tensile strain of 3%. Significantly, the H phase is predicted to display a giant carrier mobility of 1 × 106 cm2 V-1 s-1, which exceeds that for most 2D magnetic materials, with a Curie temperature (TC) exceeding room temperature and a band gap of 2.01 eV at the K point. Following the H-T phase transition, the direct band gap shifts to the Γ point and increases to 2.59 eV. The Monte Carlo (MC) simulations also indicate that TC of the T phase VSN can be effectively modulated by strain, reaching room temperature under a biaxial strain of -4%. These results show that VSN should be a promising functional material for future nanoelectronics.

Multipiezo Effect in Altermagnetic V2SeTeO Monolayer.

Strain engineering has been used as an efficient method to modulate various properties of quantum materials and electronic devices. One may establish piezo effects based on a disciplined response to the strain in multifunctional nanosystems. Inspired by a recent theoretical proposal on the interesting piezomagnetism and C-paired valley polarization in the V2Se2O monolayer, we predict a stable altermagnetic Janus monolayer V2SeTeO using density functional theory calculations. It exhibits a novel "multipiezo" effect combining piezoelectricity, piezovalley, and piezomagnetism. Most interestingly, the valley polarization and the net magnetization under strain in V2SeTeO exceed these in V2Se2O, along with the additional large piezoelectric coefficient. The "multipiezo" effect makes Janus monolayer V2SeTeO as a tantalizing material for potential applications in nanoelectronics, optoelectronics, spintronics, and valleytronics.

Augmented Fluorescence Signaling on a Single BaTiO3 Microbead Optical Booster toward High-Sensitive Biosensing.

In this work, we report a new generation of single microbead bioassay that employs a single BaTiO3 microbead as an optical booster for target biomarker enrichment and optical enhancement toward protein and nucleic acid analysis. The single BaTiO3 microbead can not only concentrate the target molecules by nearly 104-fold but also act as an optical booster to prominently enhance the target-induced fluorescence signal by the whispering gallery mode for improving the excitation efficiency and the microlens effect for promoting the signal collecting efficiency, respectively. Compared with using a conventional single microbead, this optical booster exhibits nearly 2 orders of magnitude higher sensitivity without the assistance of any signal amplification techniques or costly instruments. Moreover, this single microbead optical booster is capable of detecting different kinds of protein and nucleic acid biomarkers in a simple mix-and-read manner, holding great potential for early clinical diagnosis.

Field Detection of Uranyl in Coastal Water of China Using a Portable Device via DNA Photocleavage.

The urgent need for field detection of uranium in seawater is 2-fold: to provide prompt guidance for uranium extraction and to prevent human exposure to nuclear radiation. However, current methods for this purpose are largely hindered by bulky instrumentation, high costs of developed materials, and severe matrix interferences, which limit their further application in the field. Herein, we demonstrated a portable and label-free strategy for the field detection of uranyl in seawater based on the efficient photocleavage of DNA. Further experiments confirmed the generation of ultraviolet (UV) light-induced reactive oxygen species (ROS), such as O2•- and •OH, which fragmented oligomeric DNA in the presence of uranyl and UV light. Detailed studies showed that DNA significantly enhances uranyl absorption in the UV-visible region, leading to the generation of more ROS. A fluorescence system for the selective detection of uranyl in seawater was established by immobilizing two complementary oligonucleotides with the fluorescent dye SYBR Green I. The strategy of UV-induced photocleavage offers high selectivity, excellent interference immunity, and high sensitivity for uranyl, with a detection limit of 6.8 nM. Additionally, the fluorescence can be visually detected using a 3D-printed miniaturized device integrated with a smartphone. This method has been successfully applied to the on-site detection of uranyl in seawater in 18 Chinese coastal cities and along the coast of Hainan Island within 3 min for a single sample. The sample testing and field analysis results indicate that this strategy has promising potential for real-time monitoring of trace uranyl in China's coastal waters. It is expected to be utilized for the rapid assessment of nuclear contamination and nuclear engineering construction.

Predictive value of peripheral blood biomarkers in patients with non-small-cell lung cancer responding to anti-PD-1-based treatment.

BACKGROUND: The introduction of the anti-PD-1 antibody has greatly improved the clinical outcomes of patients with non-small cell lung cancer (NSCLC). In this study, we retrospectively analyzed the efficacy of PD-1 antibody-based therapy in patients with locally advanced inoperable or metastatic NSCLC and reported an association between peripheral blood biomarkers and clinical response in these patients. METHODS: This single-center study included medical record data of patients with NSCLC treated with the PD-1 antibody as a first-line or subsequent line of treatment, either as monotherapy or in combination with chemotherapy. The patients were enrolled from 2020 to 2022. We dynamically evaluated multiple Th1 and Th2 cytokines in the blood serum and analyzed the phenotype of T cells from the peripheral blood to explore the correlation between cytokine levels, T cell phenotypes, and clinical response. RESULTS: A total of 88 patients with stage IIIA-IV NSCLC were enrolled, out of which 60 (68.18%) achieved a partial response (PR), 13 (14.77%) had stable disease (SD), and 15 (17.05%) experienced disease progression (PD). The disease control rate was 82.95%. Our results suggested a significant reduction (P = 0.002, P < 0.005) in lymphocyte absolute counts after treatment in patients with PD. Higher levels of IFN-γ (P = 0.023, P < 0.05), TNF-α (P = 0.00098, P < 0.005), IL-4 (P = 0.0031, P < 0.005), IL-5 (P = 0.0015, P < 0.005), and IL-10 (P = 0.036, P < 0.05) were detected in the peripheral blood before treatment in the PR group compared to the PD group. Moreover, patients with high levels of IL-5, IL-13, IL-4, IL-6, IFN-γ, and TNF-α (> 10 ng/mL) had superior progression-free survival compared to those with low levels (< 10 ng/mL). Furthermore, PD-1 expression on CD8+ T cells was higher in patients who showed a PR than in those who did not show a response (SD + PD; P = 0.042, P < 0.05). CONCLUSIONS: The findings of this study imply that the decrease in absolute blood lymphocyte counts after treatment is correlated with disease progression. Serum cytokine levels may predict the effectiveness and survival rates of anti-PD-1 blockade therapy in patients with NSCLC. In addition, PD-1 expression on CD8+ T cells was positively associated with better clinical response. Our findings highlight the potential of peripheral blood biomarkers to predict the effectiveness of PD-1-targeted treatments in patients with NSCLC. Larger prospective studies are warranted to further clarify the value of these biomarkers.

The efficacy and safety of adding PD-1 blockade to induction chemotherapy and concurrent chemoradiotherapy (IC-CCRT) for locoregionally advanced nasopharyngeal carcinoma: an observational, propensity score-matched analysis.

BACKGROUND: Despite the success of PD-1 blockade in recurrent/metastatic nasopharyngeal carcinoma (NPC), its effect for locoregionally advanced NPC (LANPC) remains unclear. This study aimed to evaluate the benefit of adding PD-1 blockade to the current standard treatment (gemcitabine and cisplatin IC  plus cisplatin CCRT ) for LANPC patients. METHODS: From January 2020 to November 2022, 347 patients with non-metastatic high-risk LANPC (stage III-IVA, excluding T3-4N0) were included. Of the 347 patients, 268 patients were treated with standard treatment (IC-CCRT), and 79 received PD-1 blockade plus IC-CCRT (PD-1 group). For the PD-1 group, PD-1 blockade was given intravenously once every 3 weeks for up to 9 cycles (3 induction and 6 adjuvant). The primary endpoint was disease-free survival (DFS) (i.e. freedom from local/regional/distant failure or death). The propensity score matching (PSM) with the ratio of 1:2 was performed to control confounding factors. RESULTS: After PSM analysis, 150 patients receiving standard treatment and 75 patients receiving additional PD-1 blockade remained in the current analysis. After three cycles of IC, the PD-1 group had significantly higher rates of complete response (defined as disappearance of all target lesions; 24% vs. 9%; P = 0.006) and complete biological response (defined as undetectable cell-free Epstein-Barr virus DNA, cfEBV DNA; 79% vs. 65%; P = 0.046) than that in the standard group. And the incidence of grade 3-4 toxicity during IC was 47% in the PD-1 group and 41% in the standard group, with no significant difference (P = 0.396). During follow-up period, additional PD-1 blockade to standard treatment improved 3-year DFS from 84 to 95%, with marginal statistical significance (HR, 0.28; 95%CI, 0.06-1.19; P = 0.064). CONCLUSION: Additiaonl PD-1 blockade to gemcitabine and cisplatin IC and adjuvant treatment results in significant improvement in tumor regression, cfEBV DNA clearance, superior DFS, and comparable toxicity profiles in high-risk LANPC patients.

Diagnostic value of a novel salivary gland ultrasound scoring system in IgG4-related sialadenitis.

OBJECTIVES: To develop a novel ultrasound scoring system for the major salivary glands in patients with immunoglobulin G4-related sialadenitis (IgG4-RS) and assess its diagnostic value in a multicenter cohort of Chinese patients. METHODS: Twenty clinicians (rheumatologists, stomatologists, and radiologists) participated. The study was conducted in four steps: (1) defining the ultrasonography (US) elements, (2) developing a novel ultrasound scoring system for US of the salivary glands, (3) evaluation of inter- and intra-reader reliabilities using the new ultrasound scoring system, and (4) assessing the diagnostic value of this novel ultrasound scoring system in IgG4-RS patients in a Chinese multicenter cohort. RESULTS: A novel ultrasound scoring system for the salivary glands was developed, with total scores ranging from 0 to 34. The inter- and intra-reader reliabilities of the ultrasound scoring system were excellent (0.972 and 0.940, respectively). A total of 470 people were recruited in this study; 187 patients were diagnosed with IgG4-RS, and the remaining 283 people were diagnosed with non-IgG4-RS. Patients with IgG4-RS had significantly higher US scores than the non-IgG4-RS group (mean US score=16 vs. 4, P < 0.001). The calculated area under the curve (AUC) for the total US score was 0.852 (95% CI: 0.814-0.891). The total US scores≥9 showed a sensitivity of 75.4% and a specificity of 91.9%. Association analysis showed a positive correlation between total US scores and serum IgG4 levels and hypocomplementemia (r=0.221, r=0.349; P = 0.002) and a negative correlation between total US scores and serum C3 and C4 levels (r=-0.210, r=-0.303; P = 0.005, P < 0.001). CONCLUSIONS: A novel semiquantitative ultrasound scoring system for patients with IgG4-RS was developed, with good diagnostic performance. The inter- and intra-reader reliabilities were excellent. US scores were correlated with IgG4, C3, and C4 levels and hypocomplementemia.

Stacking architecture for collimated backlight using cylindrical lens sheet with linear light sources or edge-lit/direct-lit BLU.

Highly collimated and directional backlights are essential for realizing advanced display technologies such as autostereoscopic 3D displays. Previously reported collimated backlights, either edge-lit or direct-lit, in general still suffer unsatisfactory form factors, directivity, uniformity, or crosstalk etc. In this work, we report a simple stacking architecture for the highly collimated and uniform backlights, by combining linear light source arrays and carefully designed cylindrical lens arrays. Experiments were conducted to validate the design and simulation, using the conventional edge-lit backlight or the direct-lit mini-LED (mLED) arrays as light sources, the NiFe (stainless steel) barrier sheets, and cylindrical lens arrays fabricated by molding. Highly collimated backlights with small angular divergence of ±1.45°âˆ¼±2.61°, decent uniformity of 93-96%, and minimal larger-angle sidelobes in emission patterns were achieved with controlled divergence of the light source and optimization of lens designs. The architecture reported here provides a convenient way to convert available backlight sources into a highly collimated backlight, and the use of optically reflective barrier also helps recycle light energy and enhance the luminance. The results of this work are believed to provide a facile approach for display technologies requiring highly collimated backlights.

Up-regulation of inflammatory, oxidative stress, and apoptotic mediators via inflammatory, oxidative stress, and apoptosis-associated pathways in bovine endometritis.

Endometritis is the inflammation of the endothelial lining of the uterine lumen and is multifactorial in etiology. Escherichia (E.) coli is a Gram-negative bacteria, generally considered as a primary causative agent for bovine endometritis. Bovine endometritis is characterized by the activation of Toll-like receptors (TLRs) by E. coli, which in turn triggers inflammation, oxidative stress, and apoptosis. The objective of this study was to investigate the gene expression of inflammatory, oxidative stress, and apoptotic markers related to endometritis in the uteri of cows. Twenty uterine tissues were collected from the abattoir. Histologically, congestion, edema, hyperemia, and hemorrhagic lesions with massive infiltration of neutrophil and cell necrosis were detected markedly (P < 0.05) in infected uterine samples. Additionally, we identify E. coli using the ybbW gene (177 base pairs; E. coli-specific gene) from infected uterine samples. Moreover, qPCR and western blot results indicated that TLR2, TLR4, proinflammatory mediators, and apoptosis-mediated genes upregulated except Bcl-2, which is antiapoptotic, and there were downregulations of oxidative stress-related genes in the infected uterine tissue. The results of our study suggested that different gene expression regimes related to the immune system reflex were activated in infected uteri. This research gives a novel understanding of active immunological response in bovine endometritis.

Sub-ppt level detection of carbon monoxide and nitrous oxide enabled by mid-infrared doubly resonant photoacoustic spectroscopy.

We report highly sensitive detection of carbon monoxide (CO) and nitrous oxide (N2O) using doubly resonant photoacoustic spectroscopy paired with a quantum cascade laser (QCL) at 4.57 µm. The butterfly-packaged QCL is used to exploit the CO absorption line at 2190.02 cm-1 and the N2O absorption line at 2191.42 cm-1 by scanning the injection current. Leveraging the simultaneous acoustic and optical resonances and adopting a lower photoacoustic detection frequency, we achieve a minimum detection limit of 0.85 part-per-trillion (ppt) for CO over the 500 s averaging time, and 0.7 ppt for N2O over the 200 s averaging time. Our approach demonstrates record sensitivity for CO and N2O detection compared to state-of-the-art optical gas sensors.

Spin-Chirality-Driven Multiferroicity in van der Waals Monolayers.

Driven by the expected contribution of two-dimensional multiferroic systems with strong magnetoelectric coupling to the development of multifunctional nanodevices, here we propose, by means of first-principles calculations, vanadium-halide monolayers as a new class of spin-chirality-driven van der Waals multiferroics. The frustrated 120-deg magnetic structure in the triangular lattice induces a ferroelectric polarization perpendicular to the spin-spiral plane, whose sign is switched by a spin-chirality change. It follows that, in the presence of an applied electric field perpendicular to the monolayers, one magnetic chirality can be stabilized over the other, thereby allowing the long-sought electrical control of spin textures. Moreover, we demonstrate the remarkable role of spin-lattice coupling on magnetoelectricity, which adds to the expected contribution of spin-orbit interaction determined by an anion. Indeed, such compounds exhibit sizeable spin-driven structural distortions, thereby promoting the investigation of multifunctional spin-electric-lattice couplings.

π Phase Interlayer Shift and Stacking Fault in the Kagome Superconductor CsV_{3}Sb_{5}.

The stacking degree of freedom is a crucial factor in tuning material properties and has been extensively investigated in layered materials. The kagome superconductor CsV_{3}Sb_{5} was recently discovered to exhibit a three-dimensional CDW phase below T_{CDW}∼94 K. Despite the thorough investigation of in-plane modulation, the out-of-plane modulation has remained ambiguous. Here, our polarization- and temperature-dependent Raman measurements reveal the breaking of C_{6} rotational symmetry and the presence of three distinct domains oriented at approximately 120° to each other. The observations demonstrate that the CDW phase can be naturally explained as a 2c staggered order phase with adjacent layers exhibiting a relative π phase shift. Further, we discover a first-order structural phase transition at approximately 65 K and suggest that it is a stacking order-disorder phase transition due to stacking fault, supported by the thermal hysteresis behavior of a Cs-related phonon mode. Our findings highlight the significance of the stacking degree of freedom in CsV_{3}Sb_{5} and offer structural insights to comprehend the entanglement between superconductivity and CDW.

Aberrant activation of Notch1 signaling in the mouse uterine epithelium promotes hyper-proliferation by increasing estrogen sensitivity.

In mammals, the endometrium undergoes dynamic changes in response to estrogen and progesterone to prepare for blastocyst implantation. Two distinct types of endometrial epithelial cells, the luminal (LE) and glandular (GE) epithelial cells play different functional roles during this physiological process. Previously, we have reported that Notch signaling plays multiple roles in embryo implantation, decidualization, and postpartum repair. Here, using the uterine epithelial-specific Ltf-iCre, we showed that Notch1 signaling over-activation in the endometrial epithelium caused dysfunction of the epithelium during the estrous cycle, resulting in hyper-proliferation. During pregnancy, it further led to dysregulation of estrogen and progesterone signaling, resulting in infertility in these animals. Using 3D organoids, we showed that over-activation of Notch1 signaling increased the proliferative potential of both LE and GE cells and reduced the difference in transcription profiles between them, suggesting disrupted differentiation of the uterine epithelium. In addition, we demonstrated that both canonical and non-canonical Notch signaling contributed to the hyper-proliferation of GE cells, but only the non-canonical pathway was involved with estrogen sensitivity in the GE cells. These findings provided insights into the effects of Notch1 signaling on the proliferation, differentiation, and function of the uterine epithelium. This study demonstrated the important roles of Notch1 signaling in regulating hormone response and differentiation of endometrial epithelial cells and provides an opportunity for future studies in estrogen-dependent diseases, such as endometriosis.

Altered pupil responses to social and non-social stimuli in Shank3 mutant dogs.

Pupillary response, an important process in visual perception and social and emotional cognition, has been widely studied for understanding the neural mechanisms of neuropsychiatric disorders. However, there have been few studies on pupil response to social and non-social stimuli in animal models of neurodevelopmental disorders including autism spectrum disorder (ASD) and attention deficit hyperactivity disorder. Here, we developed a pupilometer using a robust eye feature-detection algorithm for real-time pupillometry in dogs. In a pilot study, we found that a brief light flash induced a less-pronounced and slower pupil dilation response in gene-edited dogs carrying mutations in Shank3; mutations of its ortholog in humans were repeatedly identified in ASD patients. We further found that obnoxious, loud firecracker sound of 120 dB induced a stronger and longer pupil dilation response in Shank3 mutant dogs, whereas a high reward food induced a weaker pupillary response in Shank3 mutants than in wild-type control dogs. In addition, we found that Shank3 mutants showed compromised pupillary synchrony during dog-human interaction. These findings of altered pupil response in Shank3 mutant dogs recapitulate the altered sensory responses in ASD patients. Thus, this study demonstrates the validity and value of the pupilometer for dogs, and provides an effective paradigm for studying the underlying neural mechanisms of ASD and potentially other psychiatric disorders.

Modeling SHANK3-associated autism spectrum disorder in Beagle dogs via CRISPR/Cas9 gene editing.

Despite intensive studies in modeling neuropsychiatric disorders especially autism spectrum disorder (ASD) in animals, many challenges remain. Genetic mutant mice have contributed substantially to the current understanding of the molecular and neural circuit mechanisms underlying ASD. However, the translational value of ASD mouse models in preclinical studies is limited to certain aspects of the disease due to the apparent differences in brain and behavior between rodents and humans. Non-human primates have been used to model ASD in recent years. However, a low reproduction rate due to a long reproductive cycle and a single birth per pregnancy, and an extremely high cost prohibit a wide use of them in preclinical studies. Canine model is an appealing alternative because of its complex and effective dog-human social interactions. In contrast to non-human primates, dog has comparable drug metabolism as humans and a high reproduction rate. In this study, we aimed to model ASD in experimental dogs by manipulating the Shank3 gene as SHANK3 mutations are one of most replicated genetic defects identified from ASD patients. Using CRISPR/Cas9 gene editing, we successfully generated and characterized multiple lines of Beagle Shank3 (bShank3) mutants that have been propagated for a few generations. We developed and validated a battery of behavioral assays that can be used in controlled experimental setting for mutant dogs. bShank3 mutants exhibited distinct and robust social behavior deficits including social withdrawal and reduced social interactions with humans, and heightened anxiety in different experimental settings (n = 27 for wild-type controls and n = 44 for mutants). We demonstrate the feasibility of producing a large number of mutant animals in a reasonable time frame. The robust and unique behavioral findings support the validity and value of a canine model to investigate the pathophysiology and develop treatments for ASD and potentially other psychiatric disorders.