Dual Oxidase Mutant Retards Mauthner-Cell Axon Regeneration at an Early Stage via Modulating Mitochondrial Dynamics in Zebrafish.

Dual oxidase (duox)-derived reactive oxygen species (ROS) have been correlated with neuronal polarity, cerebellar development, and neuroplasticity. However, there have not been many comprehensive studies of the effect of individual duox isoforms on central-axon regeneration in vivo. Here, we explored this question in zebrafish, an excellent model organism for central-axon regeneration studies. In our research, mutation of the duox gene with CRISPR/Cas9 significantly retarded the single-axon regeneration of the zebrafish Mauthner cell in vivo. Using deep transcriptome sequencing, we found that the expression levels of related functional enzymes in mitochondria were down-regulated in duox mutant fish. In vivo imaging showed that duox mutants had significantly disrupted mitochondrial transport and redox state in single Mauthner-cell axon. Our research data provide insights into how duox is involved in central-axon regeneration by changing mitochondrial transport.

In vivo imaging of evoked calcium responses indicates the intrinsic axonal regenerative capacity of zebrafish.

Calcium is an important messenger in the neuronal system, but its specific role in axonal regeneration has not been fully investigated. To clarify it, we constructed a noninvasive in vivo calcium-imaging model of zebrafish Mauthner cells and monitored subcellular calcium dynamics during axonal regeneration. Using the calcium indicator GCamp6f, we observed that the regenerative length correlated with the peak amplitude of the evoked calcium response before axotomy, which suggested that the evoked calcium response might serve as a useful indicator of evoked neuronal activity and axonal regenerative capacity. To investigate this possibility, we overexpressed an inward rectifying potassium channel protein, Kir2.1a, to decrease the Mauthner neuronal activity and found that the inhibition of the calcium response correlated with decreased axonal regeneration. In contrast, treatment of pentylenetetrazol and knockout of the sodium voltage-gated channel α subunit 1 gene increased the calcium response and thus enhanced axonal regeneration. Our results therefore increased the understanding of the correlation between the neural activity and the vertebrate axonal regeneration.-Chen, M., Huang, R.-C., Yang, L.-Q., Ren, D.-L., Hu, B. In vivo imaging of evoked calcium responses indicates the intrinsic axonal regenerative capacity of zebrafish.

Hypoxia Delays Oligodendrocyte Progenitor Cell Migration and Myelin Formation by Suppressing Bmp2b Signaling in Larval Zebrafish.

Hypoxia in newborns tends to result in developmental deficiencies in the white matter of the brain. As previous studies of the effects of hypoxia on neuronal development in rodents and human infants have been unable to use in vivo imaging, insight into the dynamic development of oligodendrocytes (OLs) in the central nervous system under hypoxia is limited. Here, we developed a visual model to study OL development using sublethal postnatal hypoxia in zebrafish larvae. We observed that hypoxia significantly suppressed OL progenitor cell migration toward the dorsum using in vivo imaging. Further, we found that hypoxia affected myelination, as indicated by thinner myelin sheaths and by a downregulation of myelin basic protein expression. Bmp2b protein expression was also significantly downregulated following hypoxia onset. Using gain of function and loss of function experiments, we demonstrated that the Bmp2b protein was associated with the regulation of OL development. Thus, our work provides a visual hypoxia model within which to observe OL development in vivo, and reveals the underlying mechanisms involved in these processes.

Circadian genes period1b and period2 differentially regulate inflammatory responses in zebrafish.

The circadian clock has been shown to regulate various immune processes in different animals. Our previous report demonstrated that the innate immune responses in zebrafish show significant rhythmicity that could be regulated by melatonin. Here, we used diurnal zebrafish to determine the role of circadian genes in the inflammatory responses. Our results indicate that circadian genes exhibit rhythmic oscillations in zebrafish leukocytes, and mutations of the clock genes period1b (per1b) and period2 (per2) considerably affect these oscillations. Using a wounded zebrafish inflammation model, we found that under constant dark conditions (DD), the expression of pro-inflammatory cytokines is significantly downregulated in per1b gene mutant zebrafish and significantly upregulated in the per2 gene mutant zebrafish. Furthermore, using real-time imaging technology, we found that the per1b gene markedly disturbs the rhythmic recruitment of neutrophils toward the injury, whereas the per2 gene does not show a significant effect. Taken together, our results reveal differential functions of the circadian genes per1b and per2 in the inflammatory responses, serving as evidence that circadian rhythms play a vital role in immune processes.