Characterization of developmental Na+ uptake in rainbow trout larvae supports a significant role for Nhe3b

Developing freshwater fish must compensate for the loss of ions, including sodium (Na+), to the environment. In this study, we used a radiotracer flux approach and pharmacological inhibitors to investigate the role of sodium/hydrogen exchange proteins (Nhe) in Na+ uptake in rainbow trout (Oncorhynchus mykiss) reared from fertilization in soft water (0.1 mM Na+). For comparison, a second group of embryos/larvae reared in hard water (2.2 mM Na+, higher pH and [Ca2 +]) were also included in the experiment but were fluxed in soft water, only. Unidirectional rates of Na+ uptake increased throughout development and were significantly higher in embryos/larvae reared in soft water. However, the mechanisms of Na+ uptake in both groups of larvae were not significantly different, either in larvae immediately post-hatch or later in development: the broad spectrum Na+ channel blocker amiloride inhibited 85–90% of uptake and the Nhe-inhibitor EIPA also caused near maximal inhibitions of Na+ uptake. These data indicated Na+ uptake was Nhe-mediated in soft water. A role of Nhe3b (but not Nhe2 or Nhe3a) in Na+ uptake in soft water was also supported through gene expression analyses: expression of nhe3b increased throughout development in whole embryos/larvae in both groups and was significantly higher in those reared in soft water. This pattern of expression correlated well with measurements of Na+ uptake. Together these data indicate that in part, rainbow trout embryos/larvae reared in low Na+ soft water maintained Na+ homeostasis by an EIPA sensitive component of Na+ uptake, and support a primary role for Nhe3b.