Explore the vast range of titles available.

The brain blood vasculature consists of a highly ramified vessel network that is tailored to meet its physiological functions. How the brain vasculature is formed has long been fascinating biologists. Here we report that the developing vasculature in the zebrafish midbrain undergoes not only angiogenesis but also extensive vessel pruning, which is driven by changes in blood flow. This pruning process shapes the initial exuberant interconnected meshwork into a simplified architecture. Using in vivo long-term serial confocal imaging of the same zebrafish larvae during 1.5-7.5 d post-fertilization, we found that the early formed midbrain vasculature consisted of many vessel loops and higher order segments. Vessel pruning occurred preferentially at loop-forming segments via a process mainly involving lateral migration of endothelial cells (ECs) from pruned to unpruned segments rather than EC apoptosis, leading to gradual reduction in the vasculature complexity with development. Compared to unpruned ones, pruned segments exhibited a low and variable blood flow, which further decreased irreversibly prior to the onset of pruning. Local blockade of blood flow with micro-bead obstruction led to vessel pruning, whereas increasing blood flow by noradrenergic elevation of heartbeat impeded the pruning process. Furthermore, the occurrence of vessel pruning could be largely predicted by haemodynamics-based numerical simulation of vasculature refinement. Thus, changes of blood flow drive vessel pruning via lateral migration of ECs, leading to the simplification of the vasculature and possibly efficient routing of blood flow in the developing brain.

The circadian clock orchestrates a wide variety of physiological and behavioral processes, enabling animals to adapt to daily environmental changes, particularly the day-night cycle. However, the circadian clock’s role in the developmental processes remains unclear. Here, we employ the in vivo long-term time-lapse imaging of retinotectal synapses in the optic tectum of larval zebrafish and reveal that synaptogenesis, a fundamental developmental process for neural circuit formation, exhibits circadian rhythm. This rhythmicity arises primarily from the synapse formation rather than elimination and requires the hypocretinergic neural system. Disruption of this synaptogenic rhythm, by impairing either the circadian clock or the hypocretinergic system, affects the arrangement of the retinotectal synapses on axon arbors and the refinement of the postsynaptic tectal neuron’s receptive field. Thus, our findings demonstrate that the developmental synaptogenesis is under hypocretin-dependent circadian regulation, suggesting an important role of the circadian clock in neural development. Whether the circadian clock regulates early developmental processes is poorly understood. Here, the authors report the circadian rhythm of synapse formation during early brain development by using the retinotectal system of larval zebrafish as an in vivo model.

Integrating morphological, functional and molecular information of individual neurons is critical for classifying neuronal cell types and probing circuit mechanisms of brain functions. Despite the emergence of extensive single-neuronal morphology datasets largely via random sparse labeling, it remains challenging to map arbitrarily selected neuron’s morphology in vivo, especially in conjunction with its functional and molecular characteristics. Here, we report a genetically encoded Photo-inducible single-cell labeling system (Pisces) that enables simple, rapid and long-term in vivo labeling of the entire morphology of arbitrary neurons, as exemplified in intact larval zebrafish. Pisces allows sequential tracing of multiple neurons within individual animals, facilitating brain-wide projectome mapping. Importantly, combined with in vivo calcium imaging, and fluorescence in situ hybridization or single-cell RNA sequencing, Pisces allows linking individual neurons’ morphology characterization with their functional and/or gene expression investigation, respectively. This strategy promises to advance the construction of single-neuronal multimodal atlases and expedite the elucidation of neural circuitries underlying brain functions. Understanding brain function requires integrating neuronal structure, activity, and genes. Here, authors developed an optogenetic tool they name Pisces, to enable complete labeling of individual neurons’ morphology with functional and molecular profiling, allowing multimodal single-cell analysis in vivo in zebrafish.

PRKAG2 cardiac syndrome is an autosomal dominant inherited disease resulted from mutations in the PRK- AG2 gene that encodes γ2 regulatory subunit of AMP-activated protein kinase. Affected patients usually develop ventricular tachyarrhythmia and experience progressive heart failure that is refractory to medical treatment and requires cardiac transplantation. In this study, we identify a H530R mutation in PRKAG2 from patients with familial Wolff-Parkinson-White syndrome. By generating H530R PRKAG2 transgenic and knock-in mice, we show that both models recapitulate human symptoms including cardiac hypertrophy and glycogen storage, confirming that the H530R mutation is causally related to PRKAG2 cardiac syndrome. We further combine adeno-associated virus-9 （AAV9） and the CRISPR/Cas9 gene-editing system to disrupt the mutant PRKAG2 allele encoding H530R while leav- ing the wild-type allele intact. A single systemic injection of AAV9-Cas9/sgRNA at postnatal day 4 or day 42 substantially restores the morphology and function of the heart in H530R PRKAG2 transgenic and knock-in mice. Together, our work suggests that in vivo CRISPR/Cas9 genome editing is an effective tool in the treatment of PRKAG2 cardiac syndrome and other dominant inherited cardiac diseases by selectively disrupting disease-causing mutations.

N-methyl-D-aspartate receptors (NMDARs) are a major class of glutamate receptors crucial for neural development and function. Here, we report that NMDARs expressed on neurons regulate brain vascular development via neurovascular communication. Dysfunction of neuronal NMDARs impairs the formation of the zebrafish brain vasculature and abrogates the neural activity-induced enhancement of the brain vascular development. These defects are attributed to the reduced growth of vascular endothelial tip cells (ETCs) situating at the leading edge of brain angiogenic sprouts. At the molecular level, NMDAR dysfunction down-regulates neuronal expression of vascular endothelial growth factor and subsequent global Ca 2+ activities of ETCs, thereby impairing ETC growth. Thus, our study uncovers an important role of NMDARs in brain vascular development, expanding the functional repertoire of NMDARs and the mechanistic understanding of the interplay between nervous and vascular systems during development. Neurovascular interaction is critical for normal brain vascular development. Here, the authors show that neuronal NMDARs regulate brain vascular development via VEGF-induced global Ca2+ activities of endothelial tip cells.

Background Tryptophan is an essential amino acid involved in critical cellular processes in vertebrates, serving as a precursor for serotonin and kynurenine, which are key neuromodulators to influence neural and immune functions. Systematic and quantitative measurement of tryptophan is vital to understanding these processes. Results Here, we utilized a robust and highly responsive green ratiometric indicator for tryptophan (GRIT) to quantitatively measure tryptophan dynamics in bacteria, mitochondria of mammalian cell cultures, human serum, and intact zebrafish. At the cellular scale, these quantitative analyses uncovered differences in tryptophan dynamics across cell types and organelles. At the whole-organism scale, we revealed that inflammation-induced tryptophan concentration increases in zebrafish brain led to elevated serotonin and kynurenine levels, prolonged sleep duration, suggesting a novel metabolic connection between immune response and behavior. Moreover, GRIT’s application in detecting reduced serum tryptophan levels in patients with inflammation symptoms suggests its potential as a high-throughput diagnostic tool. Conclusions In summary, this study introduces GRIT as a powerful method for studying tryptophan metabolism and its broader physiological implications, paving the way for new insights into the metabolic regulation of health and disease across multiple biological scales.

Glutamatergic retinal waves, the spontaneous patterned neural activities propagating among developing retinal ganglion cells (RGCs), instruct the activity-dependent refinement of visuotopic maps. However, its initiation and underlying mechanism remain largely elusive. Here using larval zebrafish and multiple in vivo approaches, we discover that bipolar cells (BCs) are responsible for the generation of glutamatergic retinal waves. The wave originates from BC axon terminals (ATs) and propagates laterally to nearby BCs and vertically to downstream RGCs and the optic tectum. Its initiation is triggered by the activation of and consequent glutamate release from BC ATs, and is mediated by the N -methyl- D -aspartate subtype of glutamate receptors (NMDARs) expressed at these ATs. Intercellular asymmetry of NMDAR expression at BC ATs enables the preferential initiation of waves at the temporal retina, where BC ATs express more NMDARs. Thus, our findings indicate that glutamatergic retinal waves are initiated by BCs through a presynaptic NMDA autoreceptor-dependent process. Retinal waves are important for visual system development. However, the mechanism involved in their generation remains largely unknown. Here using in vivo two-photon imaging the authors identify the presence of retinal waves in zebrafish larvae and find that they are initiated at bipolar cells via presynaptic NMDARs.

Polarized membrane addition is crucial for axon development and elongation during neuronal morphogenesis. This process is believed to be regulated by directed membrane trafficking of Rab10-containing post-Golgi carriers. However, the mechanisms underlying the biogenesis of these carriers remain unclear. Here, we report that Rab10 interaction with myosin Vb (MYO5B) determines the formation of Rab10 carriers and is important for axon development. Rab10 interacts with the exon D-encoded domain of MYO5B. Downregulating the expression of MYO5B (+D) or blocking its interaction with Rab10 impairs the fission of Rab10 vesicles from trans-Golgi membranes, causes a decrease in the number of Rab10 transport carriers and inhibits axon development in cultured hippocampal neurons. Furthermore, the MYO5B–Rab10 system is required for axon development of vertebrate neocortical neurons or zebrafish retinal ganglion cells in vivo . Thus, specific interaction between Rab10 and MYO5B controls the formation of Rab10 vesicles, which is required for axon development. Polarized membrane addition during axon development requires post-Golgi Rab10 carriers, whose biogenesis mechanisms remain unknown. This work shows that specific interaction between Rab10 and MYO5B controls formation of the Rab10 carriers, and this process is essential for neuronal polarization.

As noted by Darwin, chickens have the greatest phenotypic diversity of all birds, but an interesting evolutionary difference between domestic chickens and their wild ancestor, the Red Junglefowl, is their comparatively weaker vi- sion. Existing theories suggest that diminished visual prowess among domestic chickens reflect changes driven by the relaxation of functional constraints on vision, but the evidence identifying the underlying genetic mechanisms respon- sible for this change has not been definitively characterized. Here, a genome-wide analysis of the domestic chicken and Red Junglefowl genomes showed significant enrichment for positively selected genes involved in the development of vision. There were significant differences between domestic chickens and their wild ancestors regarding the level of mRNA expression for these genes in the retina. Numerous additional genes involved in the development of vision also showed significant differences in mRNA expression between domestic chickens and their wild ancestors, particularly for genes associated with phototransduction and photoreceptor development, such as RHO （rhodopsin）, GUCAIA, PDE6B and NR2E3. Finally, we characterized the potential role of the VIT gene in vision, which experienced positive selection and downregulated expression in the retina of the village chicken. Overall, our results suggest that positive selection, rather than relaxation of purifying selection, contributed to the evolution of vision in domestic chickens. The progenitors of domestic chickens harboring weaker vision may have showed a reduced fear response and vigi- lance, making them easier to be unconsciously selected and/or domesticated.

The retinotectal synapse in larval zebrafish, combined with live time-lapse imaging, provides an advantageous model for study of the development and remodelling of central synapses in vivo . In previous studies, these synapses were labelled by transient expression of fluorescence-tagged synaptic proteins, which resulted in the dramatic variation of labelling patterns in each larva. Here, using GAL4-Upstream Activating Sequence (GAL4-UAS) methodology, we generated stable transgenic lines, which express EGFP-tagged synaptophysin (a presynaptic protein) in retinal ganglion cells (RGCs), to reliably label the pre-synaptic site of retinotectal synapses. This tool avoids the variable labelling of RGCs that occurs in transient transgenic larvae. We obtained several stable transgenic lines that differ consistently in the number of labelled RGCs. Using stable lines that consistently had a single labelled RGC, we could trace synaptogenic dynamics on an individual RGC axonal arbor across different developmental stages. In the stable lines that consistently had multiple labelled RGCs, we could simultaneously monitor both pre- and post-synaptic compartments by combining transient labelling of post-synaptic sites on individual tectal neurons. These tools allowed us to investigate molecular events underlying synaptogenesis and found that the microRNA-132 ( miR-132 ) is required for developmental synaptogenesis. Thus, these transgenic zebrafish stable lines provide appropriate tools for studying central synaptogenesis and underlying molecular mechanisms in intact vertebrate brain.