Influence of testosterone administration on osmoregulation and energy metabolism of gilthead sea bream Sparus auratus

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Abstract

The osmoregulatory and metabolic role of testosterone (T) in the euryhaline teleost Sparus auratus was examined. Fish were implanted with a slow-release coconut oil implant alone (control) or containing T (2 or 5 μg g−1 body weight) and sampled 1, 3, and 7 days after implantation. Gill Na+,K+-ATPase activity increased in fish treated with the lower dose of T after 7 days of treatment. Kidney Na+,K+-ATPase activity enhanced at first day post-implantation in the group treated with the higher dose of T but the values diminished by day 3. Plasma levels of metabolites (glucose, lactate, triglyceride, and protein) increased after T treatment. This higher availability of plasma metabolites was reflected in several metabolic changes within different tissues of T-treated fish such as (i) increased glycogen levels and capacity for gluconeogenesis, ketogenesis, glucose exporting, and amino acid catabolism in the liver, (ii) enhanced lipogenic capacity in the gills, (iii) increased glycogen levels and capacity for oxidizing amino acids in the kidney, and (iv) enhanced levels of glycogen, aceotacetate, glucose and triglycerides, and higher capacity of phosphorylating glucose in the brain. These results provide evidence regarding an osmoregulatory and metabolic role for T in S. auratus that could be related to changes in both processes during sexual maturation.

Introduction

The most important androgens in fish are 11-ketotestosterone (11-KT)1 and testosterone (T), being both involved in the control of spermatogenesis, and development of the secondary sexual characters (Borg, 1994, Weltzien et al., 2004). The changes in plasma levels of both androgens during the reproductive cycle of male gilthead seabream have been recently described (Chaves-Pozo, 2005). 11-KT levels increased during spermatogenesis peaking (ca. 0.6 ng ml−1) at the end of such process (just prior spawning) indicating the clear implication of this androgen in driving the spermatogenetic development in male teleost (Borg, 1994). During the rest of the reproductive cycle, 11-KT levels were kept low (< 0.3 ng ml−1). As T levels are concerned, they showed little changes during spermatogenesis (0.4–0.5 ng ml−1), peaked during post-spawning (ca. 1 ng ml−1), and kept relatively high (ca. 0.7 ng ml−1) during resting. The significance of peak T levels at post-spawning is unclear. Possible mechanisms would involve the gonadotroph cell development of the pituitary (Cavaco et al., 2001), the inhibition of subsequent spermatogenetic processes to initiate the resting period (T inhibits spermatogenesis induced by 11-KT; Cavaco et al., 2001), and/or the initial step in the preparation of the gonad for the next reproductive cycle (Prat et al., 1990). In addition to their role in reproduction, sex steroids are known to be involved in growth (Sparks et al., 2003), digestion and food utilization (Ince et al., 1982), gut transport (Reshkin et al., 1989), and shifts in body composition (Dasmahapatra and Medda, 1982, Haux and Norberg, 1985).

It is well established that sexual maturation and treatment with exogenous sex steroids have a negative effect on the adaptability of several salmonids to seawater (McCormick and Naiman, 1985, McCormick, 1995, Le François and Blier, 2000). In non-salmonid species, there are few studies focused on the role of sex steroids on the adaptability to different salinities, the majority of them dealing with estrogen effects. Negative influence of 17β-estradiol (E2) treatment on gill Na+,K+-ATPase activity and hypoosmotic capacity have been reported in tilapia (Vijayan et al., 2001) and mummichog (Mancera et al., 2004). However, in gilthead sea bream Sparus auratus a clear stimulatory effect of E2 on gill Na+,K+-ATPase activity was observed after long-term treatment (Guzmán et al., 2004). To our knowledge, the evidence about a direct effect of T on gill Na+,K+-ATPase activity has been only reported in tilapia (Sunny and Oommen, 2000, Sunny and Oommen, 2002) in which gill Na+,K+-ATPase and Ca+2-ATPase activities were significantly enhanced by as early as 30 min after T treatment and also after 5 days of treatment.

Gonadal steroids exert significant effects on intermediary metabolism in vertebrates. In fish, there are several studies regarding effects of estrogens on fish energy metabolism (Mommsen and Walsh, 1988) generally addressing a metabolic reallocation of reserves from liver to the gonad. In this way, in a previous study we showed several changes in the liver, gills and brain carbohydrate metabolism of E2-treated gilthead sea bream (Sangiao-Alvarellos et al., 2005b). Considering that changes in androgen levels during the reproductive season have been shown to correlate with changes in energy allocation (Leonard et al., 2002), a role for androgens in energy metabolism seems to be reasonable. Accordingly, treatment with T in fish increased oxygen consumption (Sparks et al., 2003, Ros et al., 2004), and induced several changes in intermediary metabolism suggesting an anabolic role for this hormone in the liver (Peter and Oommen, 1989, Woo et al., 1993, Singh and Gupta, 2002, Sunny et al., 2002a). A possible metabolic role in other tissues has been less studied (Gupta et al., 1993). Therefore, the precise role of androgens in energy metabolism of teleosts remains to be thoroughly investigated.

Our group has analysed the influence of different hormones in the adaptation of gilthead sea bream to hyperosmotic and hypoosmotic environments (Mancera et al., 2002, Mancera et al., 1994, Laiz-Carrión et al., 2002, Laiz-Carrión et al., 2003, Laiz-Carrión et al., 2005, Sangiao-Alvarellos et al., 2005a). In these studies, we have also analysed the osmoregulatory and metabolic action of estrogens like E2 (Guzman et al., 2004; Sangiao-Alvarellos et al., 2005b), but to date there are no information about the possible osmoregulatory and metabolic role of T in such processes in gilthead sea bream. This species can develop gonads in estuarine areas, characterised by the presence of brackishwater but spawning always happen in seawater (Arias, 1976). It is known that in gilthead sea beam there is a “U-shaped” relationship between environmental salinity and gill Na+,K+-ATPase activity, with lower activities at isosmotic environments (estuarine areas) and higher at hyperosmotic environments (sea water of open sea) (Laiz-Carrión et al., 2005). Therefore, high levels of T necessary to stimulate gonadal development are present once gilthead sea bream enters seawater where osmoregulatory (increased gill Na+,K+-ATPase activity and hypoosmoregulatory capacity) and metabolic changes occur. Therefore, increasing levels of T observed during last phases of gonadal development take place when gilthead seabream is in seawater where osmoregulatory (increased gill Na+-K+-ATPase activity and hypoosmoregulatory capacity) and metabolic changes occur. Therefore, we aimed to assess a possible relationship between increased T levels and osmoregulatory and metabolic changes. This situation simulating the increased levels of this hormone occurring in male gilthead seabream during spawning in seawater.

Section snippets

Fish

Sexually immature male gilthead sea bream (Sparus auratus L.; 81 ± 2 g body weight) were provided by Planta de Cultivos Marinos (CASEM, Universidad de Cádiz, Puerto Real, Cádiz, Spain) and transferred to the laboratories of the Faculty of Marine Sciences (Puerto Real, Cádiz). Inmature specimens of approximately 1-year-old were used in order to eliminate the possible interference of endogenous androgens (the first sexual maturation in male gilthead sea bream occurs when fish reach approximately 400 

Results

The parameters assessed did not show any differences between untreated fish (day 0) and fish implanted with coconut oil alone (control) (data not shown). P-values resulting from the two-way ANOVA of all parameters analysed are displayed on Table 1.

Plasma T levels increased in treated fish at day 1 after implant and decreased throughout the experiment. However, plasma T levels always showed higher values in treated fish compared with controls and presented a lineal relationship with doses of T

Discussion

Implants of coconut oil plus T elevated chronically plasma levels of this hormone, which decreased slightly with time but being always significantly higher than controls. The levels observed in treated fish after 1–3 days (10–30 ng ml−1) were higher than those observed in gilthead sea bream during its sexual cycle, ca. 0.4–0.5 ng ml−1 during spermatogenesis and spawning and, around 1 ng ml−1 at post-spawning (Gothilf et al., 1997, Meiri et al., 2002). However, levels observed in treated fish after 7

Acknowledgments

This study was partly supported by Grants VEM2003-20062 (Ministerio de Ciencia y Tecnologı´a and FEDER, Spain), and PGIDT04PXIC31208PN (Xunta de Galicia, Spain) to J.L.S., and Grant BFU2004-04439-C02-01B (Ministerio de Educación y Ciencia and FEDER, Spain) to J.M.M. The authors wish to thank Planta de Cultivos Marinos (CASEM, Universidad de Cádiz, Puerto Real, Cádiz, Spain) for providing the experimental fish.

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