Zimmer Alex M, Dymowska Agnieszka K, Kumai Yusuke, Goss Greg G, Perry Steve F, Kwong Raymond W M
Department of Biology, University of Ottawa, Ottawa, Ontario, Canada.
Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada; College of Marine Sciences, University of South Florida, Saint Petersburg, USA.
Comp Biochem Physiol A Mol Integr Physiol. 2018 Dec;226:1-10. doi: 10.1016/j.cbpa.2018.06.012. Epub 2018 Jun 15.
Na uptake in larval zebrafish (Danio rerio) is coordinated by three mechanisms: Na/H-exchanger 3b (NHE3b) expressed in H-ATPase-rich (HR) cells, an unidentified Na channel coupled to electrogenic H-ATPase expressed in HR cells, and Na-Cl-cotransporter (NCC) expressed in NCC cells. Recently, acid-sensing ion channels (ASICs) were proposed to be the putative Na channel involved in H-ATPase-mediated Na uptake in adult zebrafish and rainbow trout. In the present study, we hypothesized that ASICs also play this role in Na uptake in larval zebrafish. In support of this hypothesis, immunohistochemical analyses revealed that ASIC4b was expressed in HR cells on the yolk sac skin at 4 days post-fertilization (dpf). However, neither treatment with the ASIC-specific blocker 4,6-diamidino-2-phenylindole (DAPI) nor morpholino knockdown of ASIC4b reduced Na uptake in circumneutral conditions at 4 dpf. However, because ASIC4b knockdown led to significant increases in the mRNA expression of nhe3b and ncc and a significant increase in HR cell density, it is possible that Na influx was sustained by increased participation of non-ASIC4b pathways. Moreover, when fish were reared in acidic water (pH = 4), ASIC4b knockdown led to a stimulation of Na uptake at 3 and 4 dpf, results which also were inconsistent with an essential role for ASIC-mediated Na uptake, even under conditions known to constrain Na uptake via NHE3b. Thus, while ASIC4b clearly is expressed in HR cells, the current functional experiments cannot confirm its involvement in Na uptake in larval zebrafish.
斑马鱼幼体（Danio rerio）的钠摄取由三种机制协调：富含H - ATP酶（HR）的细胞中表达的钠/氢交换体3b（NHE3b）、与HR细胞中表达的电生H - ATP酶偶联的一种未鉴定的钠通道，以及NCC细胞中表达的钠 - 氯共转运体（NCC）。最近，酸敏感离子通道（ASICs）被认为是参与成年斑马鱼和虹鳟鱼中H - ATP酶介导的钠摄取的假定钠通道。在本研究中，我们假设ASICs在斑马鱼幼体的钠摄取中也发挥这一作用。支持这一假设的是，免疫组织化学分析显示，受精后4天（dpf）时，ASIC4b在卵黄囊皮肤的HR细胞中表达。然而，无论是用ASIC特异性阻滞剂4,6 - 二脒基 - 2 - 苯基吲哚（DAPI）处理，还是用吗啉代寡核苷酸敲低ASIC4b，都不会降低4 dpf时中性条件下的钠摄取。然而，由于敲低ASIC4b导致nhe3b和ncc的mRNA表达显著增加以及HR细胞密度显著增加，钠内流可能是由非ASIC4b途径的参与增加所维持的。此外，当鱼在酸性水（pH = 4）中饲养时，敲低ASIC4b会导致3和4 dpf时钠摄取的刺激，即使在已知通过NHE3b限制钠摄取的条件下，这些结果也与ASIC介导钠摄取的重要作用不一致。因此，虽然ASIC4b显然在HR细胞中表达，但目前的功能实验无法证实其参与斑马鱼幼体的钠摄取。
