Abstract
Skeletal development and mineralization are essential processes driven by the coordinated action of neural signals, circulating molecules and local factors. Our previous studies revealed that the novel neuropeptide Pth4, synthesized by hypothalamic cells, was involved in bone metabolism via phosphate regulation in adult zebrafish. Here, we investigate the role of pth4 during skeletal development using single-cell resolution, two-photon laser ablation of Pth4:eGFP-expressing cells and confocal imaging in vivo. Using a stable transgenic Pth4:eGFP zebrafish line, we identify Pth4:eGFP-expressing cells as post-mitotic neurons. After targeted ablation of eGFP-expressing cells in the hypothalamus, the experimental larvae exhibited impaired mineralization of the craniofacial bones whereas cartilage development was normal. In addition to a decrease in pth4 transcript levels, we noted altered expression of phex and entpd5, genes associated with phosphate homeostasis and mineralization, as well as a delay in the expression of osteoblast differentiation markers such as sp7 and sparc. Taken together, these results suggest that Pth4-expressing hypothalamic neurons participate in the regulation of bone metabolism, possibly through regulating phosphate balance during zebrafish development.
MeSH terms
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Animals
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Animals, Genetically Modified
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Bone Density
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Bone and Bones / metabolism
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Bone and Bones / pathology
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Calcification, Physiologic / genetics*
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Calcinosis / genetics*
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Calcinosis / pathology
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Embryo, Nonmammalian
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Gene Expression Regulation, Developmental
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Genes, Reporter
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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Hypothalamus / growth & development
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Hypothalamus / injuries
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Hypothalamus / metabolism*
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Larva
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Laser Therapy
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Neurons / metabolism*
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Neurons / pathology
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Osteoblasts / metabolism*
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Osteoblasts / pathology
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Osteogenesis / genetics
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Osteonectin / genetics
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Osteonectin / metabolism
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PHEX Phosphate Regulating Neutral Endopeptidase / genetics
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PHEX Phosphate Regulating Neutral Endopeptidase / metabolism
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Parathyroid Hormone-Related Protein / genetics*
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Parathyroid Hormone-Related Protein / metabolism
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Phosphates / metabolism
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Pyrophosphatases / genetics
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Pyrophosphatases / metabolism
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Signal Transduction
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Sp7 Transcription Factor
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Transcription Factors / genetics
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Transcription Factors / metabolism
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Xenopus Proteins / genetics*
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Xenopus Proteins / metabolism
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Zebrafish
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Zebrafish Proteins / genetics
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Zebrafish Proteins / metabolism
Substances
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Osteonectin
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Parathyroid Hormone-Related Protein
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Phosphates
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Pth4 protein, zebrafish
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Sp7 Transcription Factor
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Transcription Factors
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Xenopus Proteins
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Zebrafish Proteins
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sp7 protein, zebrafish
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Green Fluorescent Proteins
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PHEX Phosphate Regulating Neutral Endopeptidase
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Pyrophosphatases
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entpd5a protein, zebrafish