Metabolic responses to salinity changes in the subantarctic notothenioid teleost Eleginops maclovinus L. Vargas-Chacoff, F. Moneva, R. Oyarzún, D. Martínez, E. Saavedra, I. Ruiz-Jarabo, J. L. P. Muñoz, C. Bertrán & J. M. Mancera Polar Biology ISSN 0722-4060 Volume 39 Number 7 Polar Biol (2016) 39:1297-1308 DOI 10.1007/s00300-015-1854-1 1 23 Your article is protected by copyright and all rights are held exclusively by Springer- Verlag Berlin Heidelberg. This e-offprint is for personal use only and shall not be self- archived in electronic repositories. If you wish to self-archive your article, please use the accepted manuscript version for posting on your own website. You may further deposit the accepted manuscript version in any repository, provided it is only made publicly available 12 months after official publication or later and provided acknowledgement is given to the original source of publication and a link is inserted to the published article on Springer's website. The link must be accompanied by the following text: "The final publication is available at link.springer.com”. 1 23 Author's personal copy Polar Biol (2016) 39:1297–1308 DOI 10.1007/s00300-015-1854-1 ORIGINAL PAPER Metabolic responses to salinity changes in the subantarctic notothenioid teleost Eleginops maclovinus 1,2 1 1 1 1 L. Vargas-Chacoff • F. Moneva • R. Oyarzu´n • D. Martı´nez • E. Saavedra • 3 4 1 5 I. Ruiz-Jarabo • J. L. P. Mun˜oz • C. Bertra´n • J. M. Mancera Received: 2 March 2015 / Revised: 25 November 2015 / Accepted: 30 November 2015 / Published online: 8 December 2015 Ó Springer-Verlag Berlin Heidelberg 2015 Abstract Eleginops maclovinus is an endemic, sub- affected by salinity changes, probably due to its role as a antarctic Notothenioidei species. This study examined the supplier of energetic substrates. The gills and kidney, influence of different environmental salinities (5, 15, and osmoregulatory tissues, maintained their energy metabo- 45 psu; and 32 psu as a control) on energy metabolism in E. lism levels with minor modifications. In conclusion, E. maclovinus over a period of 14 days. Metabolite contents maclovinus exhibits metabolic adjustments to adapt to and enzymatic activities related to carbohydrate, amino different salinities, showing the best responses in isosmotic acid, and lipid metabolisms were evaluated in metabolic environmental salinities. (liver) and osmoregulatory (gill and kidney) tissues. At extreme salinities (5 and 45 psu), the liver showed a high Keywords Carbohydrates Á Eleginops maclovinus Á consumption of energy reserves, mainly as amino acids and Gills Á Kidney Á Lipids Á Liver Á Metabolism carbohydrates. Carbohydrate metabolism in the gills did not change under different salinities, but increased lactate levels were found, suggesting that this tissue may use Introduction lactate as an energy substrate. Amino acid metabolism in the gills decreased at 5 psu but increased at 45 psu, and Euryhaline teleosts can acclimate to a variety of salinity lipid metabolism increased at 5 and 15 psu during the first conditions. This adaptive capacity depends on several days of the trial, indicating a possible use of lipids as factors, including energy supply and demand in relation to energy. Kidney carbohydrate catabolism and amino acid osmoregulatory processes (Soengas et al. 2007). Impor- metabolism increased after 14 days at 45 psu, while lipid tantly, successful acclimation to changes in salinity may metabolism did not vary in relation to salinity changes. require metabolic reorganization to meet increased ener- Together, these results suggest that the liver is most getic demands (Boeuf and Payan 2001; Soengas et al. 2007; Vargas-Chacoff et al. 2009a). To adapt to different salinities, teleosts present & L. Vargas-Chacoff osmoregulatory and metabolic adjustments in a number of [email protected] tissues, such as the gills, kidney, and liver. For example, 1 Instituto de Ciencias Marinas y Limnolo´gicas, Universidad sodium and chloride transport across the gill epithelium Austral de Chile, Valdivia, Chile adjusts to environmental salinity, exercising ion uptake in 2 Centro Fondap de Investigacio´n de Altas Latitudes (IDEAL), hypoosmotic water and ion excretion in hyperosmotic Universidad Austral de Chile, Valdivia, Chile water (McCormick 2001). In turn, the kidney plays an 3 Centre of Marine Sciences (CCMar), Universidade do active role in the extrusion of divalent ions and the elim- Algarve, Campus de Gambelas, Faro, Portugal ination of excess water in hyperosmotic and hypoosmotic 4 Centro I-Mar, Universidad de los Lagos, Puerto Montt, Chile environments, respectively (Bijvelds et al. 1998; Kelly and Woo 1999; Kelly et al. 1999; Beyenbach 2000). Adaptive 5 Departamento de Biologı´a, Facultad de Ciencias del Mar y Ambientales, Campus de Excelencia Internacional del Mar modifications to salinity require energy (i.e. ATP) and may (CEI-MAR), Universidad de Ca´diz, Ca´diz, Spain alter overall energy requirements, and the kidney has an 123 Author's personal copy 1298 Polar Biol (2016) 39:1297–1308 additional function in changing intermediary metabolism temperature (12.0 ± 0.5 °C) conditions. Specimens of E. (Soengas et al. 1994, 2007; Kelly and Woo 1999; Arjona maclovinus (150 ± 5 g) were fed daily at 1 % of their et al. 2009). Finally, the liver is the main site of glycogen/ body weight with commercial dry pellets (Skretting glucose turnover, ammoniagenesis, fatty acid synthesis, Nutrece Defense 100) containing 48 % protein, 22 % fat, and gluconeogenesis in teleosts (Perago´n et al. 1998). Liver 13 % carbohydrates, 8 % moisture, and 8.5 % ash. metabolism may become enhanced during osmotic adap- Before subjecting fish to salinity challenges, eight fish tation to provide fuel for metabolic and osmoregulatory were sampled as pre-transfer controls. Then, the SW-ac- processes, especially in osmoregulatory tissues such as the climated fish were divided as previously mentioned into gills and kidney (Vargas-Chacoff et al. 2009a, b; Costas four groups and subjected to one of the following salinities: et al. 2011). low salinity water (LSW; 5 psu); medium salinity water Eleginops maclovinus (Valenciennes 1830) is a sub- (MSW; 15 psu); SW (32 psu, control group); or high antarctic Notothenioidei endemic to the Patagonian region salinity water (HSW; 45 psu). Samples were taken 1, 3, 7, of South America. Due to its high degree of euryhalinity, and 14 days post-transfer (dpt). this species can be found in environments with different All experimental procedures complied with guidelines salinities, including the open sea, coastal waters, estuaries, of the Comisio´n Nacional de Ciencias y Tecnologı´ade and freshwater rivers (Pave´s et al. 2005; Pequen˜o et al. Chile (CONICYT) and the Universidad Austral de Chile 2010). Moreover, Reyes and Hune (2013) found E. for the use of laboratory animals. maclovinus inhabiting intertidal pools isolated from the tidal flux for extended periods. In these pools, evaporation Tissue sampling can increase the salinity up to 3–4 times higher than sea- water (Nordlie 1985; Panfili et al. 2004, 2006; Tine et al. Fish were netted and subjected to lethal doses of 2-phe- 2007). Another report in the osmoregulatory system of E. noxyethanol (1 ml l-) and cervical dislocation before tis- maclovinus studied the influence of acclimation to four sues were sampled. The fish were weighed, and then a gill different environmental salinities (5, 15, 32, and 45 psu) arch (dried with absorbent paper), a portion of the kidney, over 14 days (Vargas-Chacoff et al. 2014a), finding that and the complete liver were extracted from each fish, fro- specimens used allostatic changes to accommodate their zen in liquid nitrogen, and stored at -80 °C. osmotic system (McEwen and Wingfield 2003). However, modifications in the intermediary metabolism Tissue metabolites and enzymatic activities related to osmoregulation have been poorly studied in subantarctic teleosts (Magnoni et al. 2013) and, to the best Frozen liver, kidney, and gill samples were finely minced of our knowledge, these regulatory mechanisms have never in an ice-cooled Petri dish and divided into two aliquots to been analysed in E. maclovinus. Therefore, the objective of assess metabolite levels and enzymatic activities. To assess the present study was to examine the influence of four metabolite levels, the frozen tissues were homogenized by different environmental salinities (5, 15, and 45 psu, plus a ultrasonic disruption with 7.5 volumes of ice-cooled 0.6°N control group at 32 psu) on the energy metabolism of perchloric acid, neutralized using 1 M potassium bicar- different tissues (liver, gills, and kidney) in juvenile E. bonate, and centrifuged for 30 min at 13,0009g (Eppen- maclovinus using a 14-day acclimation time-course dorf 5415R). The resulting supernatant was used to experiment. determine tissue metabolite levels. Tissue lactate and triglyceride levels were spectrophotometrically measured using commercial kits (Spinreact, Lactate Ref. 1001330 Materials and methods and Triglycerides Ref. 1001311). Tissue glycogen con- centrations were assessed using the Keppler and Decker Fish and experimental design (1974) method. Glucose levels were obtained after glyco- gen breakdown (i.e. after subtracting free glucose levels) The same specimen and experimental procedures used in using a commercial kit (Spinreact, Glucose-HK Ref. Vargas-Chacoff et al. (2014a) were applied in the present 1001200). Total a-amino acid levels were assessed using study. Briefly, juvenile specimens of E. maclovinus