SPECIAL ISSUE REGULAR PAPER Effects of elevated temperature on osmoregulation and stress responses in Atlantic salmon Salmo salar smolts in fresh water and seawater Luis Vargas-Chacoff 1,2,3 | Amy M. Regish 2 | Andrew Weinstock 2 | Stephen D. McCormick 2,4 1 Instituto de Ciencias Marinas y Limnológicas, Laboratorio de Fisiología de Peces, Universidad Austral de Chile, Valdivia, Chile 2 U.S. Geological Survey, Leetown Science Center, S.O. Conte Anadromous Fish Research Laboratory, Turners Falls, Massachusetts 3 Centro Fondap-IDEAL, Universidad Austral de Chile, Valdivia, Chile 4 Department of Biology, University of Massachusetts, Amherst, Massachusetts Correspondence Luis Vargas-Chacoff, Instituto de Ciencias Marinas y Limnológicas, Laboratorio de Fisiología de Peces, Universidad Austral de Chile, Valdivia, Chile. Email: luis.vargas@uach.cl Funding information Fondap-IDEAL, Grant/Award Number: Fondap-IDEAL 15150003; Fondecyt, Grant/ Award Number: Fondecyt 1160877 Smolting in Atlantic salmon Salmo salar is a critical life-history stage that is preparatory for downstream migration and entry to seawater that is regulated by abiotic variables including photoperiod and temperature. The present study was undertaken to determine the interaction of temperature and salinity on salinity tolerance, gill osmoregulatory proteins and cellular and endocrine stress in S. salar smolts. Fish were exposed to rapid changes in temperature (from 14 to 17, 20 and 24  C) in fresh water (FW) and seawater (SW), with and without prior acclima- tion and sampled after 2 and 8 days. Fish exposed simultaneously to SW and 24  C experienced 100% mortality, whereas no mortality occurred in any of the other groups. The highest tempera- ture also resulted in poor ion regulation in SW with or without prior SW acclimation, whereas no substantial effect was observed in FW. Gill Na + –K + -ATPase (NKA) activity increased in SW fish compared to FW fish and decreased with high temperature in both FW and SW. Gill Nkaα1a abundance was high in FW and Nkaα1b and Na + –K + -2Cl- cotransporter high in SW, but all three were lower at the highest temperature. Gill Hsp70 levels were elevated in FW and SW at the highest temperature and increased with increasing temperature 2 days following direct transfer to SW. Plasma cortisol levels were elevated in SW at the highest temperature. Our results indicate that there is an important interaction of salinity and elevated temperature on osmoregulatory performance and the cellular stress response in S. salar, with an apparent threshold for osmoregulatory failure in SW above 20  C. KEYWORDS heat shock protein, ion transport, salinity, Salmo salar, smolts 1 | INTRODUCTION As part of their anadromous life history, Atlantic salmon Salmo salar L. 1758 migrates from fresh water (FW) to seawater (SW) as juveniles. This normally occurs after undergoing preparatory changes in behav- iour, morphology and physiology that are adaptive for downstream migration and seawater entry (McCormick, 2013). This parr–smolt transformation occurs in spring and is mediated primarily through photoperiod and temperature cues. One of the hallmarks of the parr– smolt transformation is a large increase in the capacity for ion regulation in seawater that is adaptive for rapid movements through estuaries and into the open ocean. There has been substantial research on the ion-transport mecha- nisms and control of the ability of salmonids to move from fresh water to seawater. The gill plays a principal role in the maintenance of ion homeostasis in both FW and SW-acclimated fish (Evans et al., 2005). In order to maintain the internal ionic balance, the gills have special- ized cells called ionocytes (also known as mitochondrion-rich or chlo- ride cells) that are involved in chloride and sodium secretion in SW and uptake in FW (Hiroi & McCormick, 2012; Marshall & Grosell, 2006). There are three major ion-transport proteins involved in sodium and chloride secretion by the SW gill. Na + –K + -ATPase (NKA) pumps three sodium ions out of the cell while pumping in two potas- sium ions, making the inside of the chloride cell both low in sodium and negatively charged (Marshall & Grosell, 2006). The sodium gradi- ent is then used by the Na + –K + -2Cl - co-transporter (NKCC) to bring chloride into the cell (Cutler & Cramb, 2002). Chloride subsequently leaves the cells on a favourable electrical gradient through an apical Received: 14 November 2017 Accepted: 4 May 2018 DOI: 10.1111/jfb.13683 FISH 550 © 2018 The Fisheries Society of the British Isles wileyonlinelibrary.com/journal/jfb J Fish Biol. 2018;93:550–559.