Abstract：
Cl- is a major anion in the bodily fluids of vertebrates, and maintaining its homeostasis is essential for normal physiological functions. Fishes inhabiting freshwater (FW) passively lose body fluid ions, including Cl-, to the external environment because of the electrochemical gradient of ions across the body surface. Therefore, FW fishes have to actively absorb Cl- from the surroundings to maintain ion homeostasis in their bodily fluids. Hormonal control is vital for modulating ion uptake in fish. Vitamin D is involved in the regulation of Ca2+ uptake and acid secretion in fish. In the present study, we found that the levels of bioactive vitamin D, 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3), significantly increased in zebrafish embryos and adults after exposure to water containing low levels of Cl-. Moreover, the administration of 1α,25(OH)2D3 treatment (20 μg/L) in zebrafish embryos, and intraperitoneal (i.p.) injection of 1α,25(OH)2D3 (5 μg/kg body mass) in zebrafish adults, resulting the increased Cl- content in bodily fluid in zebrafish. Na+-Cl- cotransporter 2b (NCC2b) and Cl- channel 2c (CLC2c) are specifically expressed during Cl- uptake by ionocytes in zebrafish. Our results indicated that the mRNA and protein expression of NCC2b and CLC2c considerably increased in the zebrafish with exogenous 1α,25(OH)2D3 treatment. Additionally, exogenous 1α,25(OH)2D3 administration increased the number of NCC2b- and CLC2c-expressing cells in yolk skins of zebrafish embryos and the gill filaments of zebrafish adults. Transcript signals of vitamin D receptors (VDRs) were identified in NCC2b-expressing cells. Knockdown of VDRa and VDRb significantly reduced the expression of NCC2b and CLC2c and the number of NCC2b- and CLC2c-expressing cells. These results indicate that vitamin D can affect Cl- uptake in zebrafish and extend our knowledge of the role of vitamin D in fish physiology.
摘要：
Cl-是脊椎动物体液中的主要阴离子,维持其体内平衡对于正常的生理功能至关重要。生活在淡水（FW）中的鱼类被动失去体液离子，包括Cl-,由于离子在整个身体表面的电化学梯度，因此到达外部环境。因此,FW鱼类必须从周围环境中主动吸收Cl-以维持其体液中的离子稳态。激素控制对于调节鱼类的离子吸收至关重要。维生素D参与调节鱼的Ca2+摄取和酸分泌。在本研究中,我们发现生物活性维生素D的水平，1α,25-二羟维生素D3（1α，25(OH)2D3),斑马鱼胚胎和成人暴露于含低水平Cl-的水后显著增加。此外，维生素D(1α，斑马鱼胚胎中的25（OH）2D3处理（20μg/L），腹膜内（i.p.）注射1α，斑马鱼成虫中的25（OH）2D3（5μg/kg体重），导致斑马鱼体液中Cl-含量增加。Na+-Cl-协同转运蛋白2b(NCC2b)和Cl-通道2c(CLC2c)在斑马鱼离子细胞摄取Cl-期间特异性表达。我们的结果表明，在外源性1α斑马鱼中，NCC2b和CLC2c的mRNA和蛋白表达显著增加。25（OH）2D3处置。此外,外源1α，25（OH）2D3的施用增加了斑马鱼胚胎的卵黄皮和斑马鱼成体的g丝中NCC2b和CLC2c表达细胞的数量。在表达NCC2b的细胞中鉴定了维生素D受体(VDR)的转录物信号。VDRa和VDRb的敲除显著降低了NCC2b和CLC2c的表达以及NCC2b和CLC2c表达细胞的数量。这些结果表明，维生素D可以影响斑马鱼的Cl-摄取，并扩展了我们对维生素D在鱼类生理中的作用的认识。
