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Advancing neuroscience through real-time processing of big data:Transition from open-loop to closed-loop paradigms

Zoological Research 2024年第3期45卷 518-519页

作者： Yu-Fan Wang jiu-lin du Institute of Neuroscience State Key Laboratory of NeuroscienceCenter for Excellence in Brain Science and Intelligence TechnologyChinese Academy of SciencesShanghai 200031China University of Chinese Academy of Sciences Beijing 100049China School of Life Science and Technology ShanghaiTech UniversityShanghai 200031China

The brain functions as a closed-loop system that continuously generates behavior in response to the external environment and adjusts actions based on the *** research methodologies in neuroscience,especially those employed in brain imaging experiments,have mainly adopted an open-loop paradigm(Grosenick et al.,2015).Functional neural circuits are analyzed offline and subsequently tested through manipulation of neuronal activities within specific regions or with genetic *** establishing a closed-loop research paradigm,functional ensembles can be detected and tested in real time with temporal *** functional ensembles,rather than brain regions or genetically labeled neural populations,serve as fundamental units of neural networks,offering valuable insights into the dissection of neural *** closed-loop research paradigm also enables the capture of high-dimensional activities of internal brain dynamics and precise elucidation of physiological processes such as learning,decision-making,and sleep.

关键词： neural loop Transition

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Improvement in Tol2 transposon for efficient large-cargo capacity transgene applications in cultured cells and zebrafish(Danio rerio)

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Zoological Research 2024年第3期45卷 567-574页

作者： Peng-Cheng Wang Hao Deng Rang Xu jiu-lin du Rongkun Tao Department of Pediatric Cardiology Xinhua HospitalShanghai Jiao Tong University School of MedicineShanghai 200092China Institute of Neuroscience State Key Laboratory of NeuroscienceCenter for Excellence in Brain Science and Intelligence TechnologyChinese Academy of SciencesShanghai 200031China Medical School Kunming University of Science and TechnologyKunmingYunnan 650500China Scientific Research Center Xinhua HospitalShanghai Jiao Tong University School of MedicineShanghai 200092China University of Chinese Academy of Sciences Beijing 100049China School of Life Science and Technology ShanghaiTech UniversityShanghai 200031China

Most viruses and transposons serve as effective carriers for the introduction of foreign DNA up to 11 kb into vertebrate ***,their activity markedly diminishes with payloads exceeding 11 *** the payload capacity of transposons could facilitate more sophisticated cargo designs,improving the regulation of expression and minimizing mutagenic risks associated with molecular therapeutics,metabolic engineering,and transgenic animal *** this study,we improved the Tol2 transposon by increasing protein expression levels using a translational enhancer(QBI SP163,ST)and enhanced the nuclear targeting ability using the nuclear localization protein H2B(SHT).The modified Tol2 and ST transposon efficiently integrated large DNA cargos into human cell cultures(H1299),comparable to the well-established super PiggyBac ***,mRNA from ST and SHT showed a significant increase in transgene delivery efficiency of large DNA payloads(8 kb,14 kb,and 24 kb)into zebrafish(Danio rerio).This study presents a modified Tol2 transposon as an enhanced nonviral vector for the delivery of large DNA payloads in transgenic applications.

关键词： Zebrafish Tol2 transposase Transgene Large payload Synthetic biology

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One-step generation of zebrafish carrying a conditional knockoutknockin visible switch via CRISPR/Cas9-mediated intron targeting

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Science China(Life Sciences) 2020年第1期63卷 59-67页

作者： Jia Li Hong-Yu Li Shan-Ye Gu Hua-Xing Zi Lai Jiang jiu-lin du Institute of Neuroscience State Key Laboratory of NeuroscienceCAS Center for Excellence in Brain Science and Intelligence TechnologyShanghai Research Center for Brain Science and Brain-Inspired IntelligenceChinese Academy of SciencesShanghai 200031China University of Chinese Academy of Sciences Beijing 100049China Department of Anesthesiology and Surgical Intensive Care Unit Xinhua HospitalShanghai Jiao Tong University School of MedicineShanghai 200092China School of Life Science and Technology Shanghai Tech UniversityShanghai 200031China

The zebrafish has become a popular vertebrate animal model in biomedical ***,it is still challenging to make conditional gene knockout(CKO)models in zebrafish due to the low efficiency of homologous recombination(HR).Here we report an efficient non-HR-based method for generating zebrafish carrying a CKO and knockin(KI)switch(zCKOIS)coupled with dual-color fluorescent *** this strategy,we generated hey2^zCKOIS which served as a hey2 KI reporter with EGFP *** Cre induction in targeted cells,the hey2^zCKOIS was switched to a non-functional CKO allele hey2^zCKOIS-inv associated with Tag RFP expression,enabling visualization of the CKO ***,simplification of the design,and the visibility and combination of both CKO and KI alleles make our z CKOIS strategy an applicable CKO approach for zebrafish.

关键词： NHEJ non-HR knockin conditional knockout visible switch zCKOIS zebrafish

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Mapping functional and anatomical whole-brain connectome of larval zebrafish

Mapping functional and anatomical whole-brain connectome of ...

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中国神经科学学会第十二届全国学术会议

作者： jiu-lin du Institute of Neuroscience State Key Laboratory of Neuroscience Center for Excellence in Brain Science and Intelligence Technology Chinese Academy of Sciences

Zebrafish is a relatively new vertebrate animal model with a conserved brain architecture and rich repertoire of behaviors. due to the transparency and simplicity of the brain, larval zebrafish has emerged as an ideal model for dissecting brain functions at a whole-brain scale based on a strategy from synapses, neurons, circuitries to behaviors. For this purpose, our lab has developed multiple in vivo assays tailored for system neuroscience research with larval zebrafish and studied neural mechanisms underlying visuomotor processing and behavior generation. Using in vivo light-sheet calcium imaging, we can simultaneously monitor the activity of all neurons(10 neurons) in the larval brain and animal behaviors in response to sensory stimuli and search for relevant functional neuronal ensembles. Optogenetics can be then performed to manipulate the activity of the neuronal ensemble to study the causality between the ensemble and behavior. In vivo whole-cell recording of neurons in the ensemble can be further performed to explore underlying synaptic and circuit mechanisms. Furthermore, by combining CRISPR/Cas9-based knockin, single-cell electroporation and virus-based tracing, the brain-wide projections and even synaptic partners of the neuronal ensemble can be figured out. By using this strategy, the brain-wide anatomical and functional connectomes of neural circuits underlying behaviors can be systematically elucidated.

关键词： neural circuit calcium imaging optogenetics sensory behavior connectome zebrafish

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Tackling neural circuits of visual behaviors by using larval zebrafish

Tackling neural circuits of visual behaviors by using larval...

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中国神经科学学会第十二届全国学术会议

作者： Bai-bing Zhang jiu-lin du Institute of Neuroscience State Key Laboratory of Neuroscience Center for Excellence in Brain Science and Intelligence Technology Chinese Academy of Sciences University of Chinese Academy of Sciences School of Life Science and Technology Shanghai Tech University

Visual behaviors are essential for animal’s survival. Elucidating the neural circuit underlying visual behaviors is one of the big challenges in Neuroscience research. Zebrafish larvae with the transparent brain, conserved but relatively simple brain structure and variable visual behaviors, is as an ideal model for exploring neural circuit mechanisms of visual behaviors. At the beginning, I developed an efficient knockin approach to label and manipulate specific neurons in zebrafish. Combining with in vivo functional assays, I then devoted to figure out neural circuits of visual behaviors. Conventional transgenic method for zebrafish is complex and time consuming, and the constructed promoter cannot fully recapture the original expression pattern, making conventional method unsuitable for functional research. Therefore, in my first study,collaborated with a postdoc, I used the CRISPR/Cas9 system and established an intron targeting-mediated and homologous recombination-independent knockin approach, with which Gal4 or EGFP was efficiently integrated into cell type-specific genes in a multicistronic or fused manner, and the integrity of the coding sequence and regulatory elements of targeted genes are maintained. This method provides us an applicable knockin strategy with the advantage of readiness, high efficiency and maintenance of targeted gene integrity, and has been highly appreciated in the zebrafish community. This work was published on CELL RESEARCH（2015）. With this method, I generated many specific Gal4 lines labeling neuromodulatory systems that are very helpful for my later studies. Visual cues with distinct ethological meaning will induce different visual behaviors. However, the mechanism of how appropriate behaviors are selected remains poorly understood. In my second study, collaborated with two senior students, we used a defined zebrafish escape circuit as a model and found that behavior selection can be implemented at the visuomotor transformation stage

关键词： CRISPR/Cas9 system zebrafish knockin strategy visual-induced escape behavior dopaminergic neuron glycinergic neuron light-preference behavior habenula eminentia thalami retinal ganglion cells neural circuit zebrafish

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Dopaminergic gating of visual flow to escape circuit via hindbrain interneurons

Dopaminergic gating of visual flow to escape circuit via hin...

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中国神经科学学会第十届全国学术会议

作者： Yuan-yuan Yao jiu-lin du Institute of Neuroscience and State Key Laboratory of Neuroscience Shanghai Institutes for Biological Sciences Chinese Academy of Sciences

Gating is an important strategy for neural signal processing and behavior ***,its underlying mechanism is largely *** zebrafish as a model organism,we find that visual input to the escape circuit is gated at the synapses made by optic tectal neurons on Mauthner cells,command-like neurons for escape *** is the gating signal to control visual information transmission to the escape circuit via dopamine *** dopaminergic gating is achieved by downstream inhibition,as evidenced by the fact that dis-inhibition can mimic and enhancing inhibition can attenuate dopaminergic *** combining electrophysiological,genetic and two-photon ablation methods,we further identify dopamine neurons in the caudal hypothalamus and two clusters of glycinergic interneurons in the hindbrain as a gating ***,by differentially regulating the firing activity of the gating module,this gating prevents behavior-irrelevant visual inputs but permits behavior-relevant ones to go through the sensorimotor circuit for initiating escape ***,our findings provide a cellular and circuit mechanism for sensory processing that involves dopaminergic modulation of interneurons.

关键词： Information gating dopaminergic intemeuron escape behavior visual input zebrafish

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Reciprocal regulation between resting microglial dynamics and neuronal activity in vivo

Reciprocal regulation between resting microglial dynamics an...

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中国神经科学学会第十届全国学术会议

作者： Ying LI Xu-Fei du jiu-lin du Institute of Neuroscience and State Key Laboratory of Neuroscience Shanghai Institutes for Biological SciencesChinese Academy of Sciences

Objective Microglia are the primary immune cells in the *** physiological conditions,they typically stay in a "resting" state,with ramified processes continuously extending to and retracting from surrounding neural *** and how such highly dynamic resting microglia functionally interact with surrounding neurons are still *** In vivo time-lapse imaging of both microghal morphology and neuronal activity in the optic tectum of larval zebrafish was *** We found that neuronal activity steers resting microglial processes and facilitates their contact with highly active *** process requires the activation of pannexin-1 hemichannels on ***,such resting microglianeuron contact reduces both spontaneous and visually evoked activities of contacted *** Our findings reveal an instructive role for neuronal activity in resting microghal motility and suggest the function for microglia in homeostatic regulation of neuronal activity in the healthy brain.

关键词： resting microglia neuronal activity contact pannexin-1 Rac in vivo imaging zebrafish

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Circadian rhythm regulates diurnal developmental synaptogenesis via the hypocretin system in vivo

Circadian rhythm regulates diurnal developmental synaptogene...

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中国神经科学学会第十届全国学术会议

作者： Xu-Fei du Ying LI jiu-lin du Institute of Neuroscience and State Key Laboratory of Neuroscience Shanghai Institutes for Biological SciencesChinese Academy of Sciences

Objective Orcadian rhythm orchestrates a wide variety of physiological and behavioral processes of animals to adapt to cyclic environmental *** the adult brain,circadian clock is capable of regulating the structural plasticity of *** remains unknown whether it can regulate developmental synaptogenesis,a fundamental process for building neural circuits in the developing *** Using in vivo two-photon time-lapse imaging of all the presynaptic puncta on the axon arbor of individual retinal ganglion cells in transgenic zebrafish larvae,we examined the dynamics of retinotectal synaptogenesis across day-night cycles during 4-6 days ***（1） We found that the synaptogenic rate undergoes a diurnal oscillation with a peak at daytime and a trough at nighttime.（2） This diurnal oscillation persists under constant darkness or constant light,indicating the involvement of circadian clock in regulating synaptogenesis.（3） This circadian rhythrnicity in synaptogenesis is due to a higher rate of synapse formation at subjective day than at subjective night,but not the difference of synapse elimination rate.（4）Mechanistically,the hypocretin signaling is required for this circadian regulation of *** Our study reveals an endogenous circadian regulation of synaptogenesis in the developing brain and adds a new line of evidence for the circadian control of developmental processes in living animals.

关键词： synaptogenesis circadian rhythm development hypocretin transgenic zebrafish retinotectal system two-photon imaging in vivo time-lapse imaging

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Developmental pruning of early-stage myelin segments during CNS myelination in vivo

Developmental pruning of early-stage myelin segments during ...

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中国神经科学学会第十届全国学术会议

作者： Peng LIU jiu-lin du Cheng HE Department of Neurobiology and Key Laboratory of Molecular Neurobiology of Ministry of Education Second Military Medical University Institute of Neuroscience and State Key Laboratory of Neuroscience Shanghai Institutes for Biological SciencesChinese Academy of Sciences

Objective Axon-wrapping myelin sheath is essential for proper functioning of the nervous *** is formed from membranous sheets that are elaborated by oligodendrocytes（OLs） in the central nervous system（CNS）.Although much is known about the development and migration of OL and its progenitor cell in vitro and in vivo,how CNS myelin is dynamically formed through OL processes is still *** To examine the process of CNS myelination in vivo,we performed long-term time-lapse confocal imaging of the same OLs in the spinal cord of intact zebrafish larvae during 2-7 days post-fertilization（dpf） with intervals of 2- 12 *** OLs were mosaically labeled by membrane-tethered red fluorescent protein（mRFP） under the control of zebrafish SoxlO regulatory *** We find that,besides extension of some early-stage axon-wrapping myelin segments,28%segments undergo pruning in an axon diameter-correlated manner during early ***-regulating neuregulin-ErbB signaling impairs the pruning but not ***,chimera analyses also indicate that ErbB receptors are specifically required in *** Thus our results indicate myelin segments pruning as a novel growth pattern during development of oligodendrocyte in vivo.

关键词： oligodendrocyte myelin segments pruning NRG1 ErbB zebrafish

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Local Calcium Activity Regulates Pathfinding of Brain Endothelial Tip Cells

Local Calcium Activity Regulates Pathfinding of Brain Endoth...

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中国神经科学学会第十二届全国学术会议

作者： Ting-Ting LIU Xu-Fei du jiu-lin du Institute of Neuroscience State Key Laboratory of Neuroscience Center for Excellence in Brain Science and Intelligence Technology Chinese Academy of Sciences University of Chinese Academy of Sciences

Pathfinding of endothelial tip cell plays a critical role in the formation of brain vasculature. It is achieved by coordinated and highly dynamic behaviors of multiple tip cell branches. However, the underlying mechanism of endothelial tip cell’s pathfinding remains unknown. Especially, it is still an open question how endothelial tip cells retract redundant branches and stabilize and extend the remaining one. Here, using the developing brain vasculature in intact larval zebrafish as a model, we found that the pattern of compartmentalized local Ca2+ activity in the branch of endothelial tip cells determines the fate of the branch. In vivo Ca2+ imaging showed that tip cell branches displayed spontaneous local Ca2+ activities. Combining local caging/uncaging and optogenetics, we found that high and low frequency of local Ca2+ activities triggers retraction and extension, respectively. Furthermore, we found that the Piezo1, a mechanosensitive and Ca2+ permeable non-selective cationic channel, play an important role in the local Ca2+ activity of endothelial tip cells. Our study reveals a critical function of subcellular Ca2+ signaling in controlling endothelial tip cell branch dynamics.

关键词： endothelial tip cell brain vasculature zebrafish Ca2+ retraction extension Piezo1

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