Colour and melanophore function in rainbow trout after long term exposure to the new antifoulant medetomidine Anna Lennquist a, * , Lena G.E. Mårtensson Lindblad a,b , Daniel Hedberg a , Erik Kristiansson a , Lars Förlin a a Department of Zoology/Zoophysiology, University of Gothenburg, Sweden b I-tech AB, Haraldsgatan 5, S-413 14 Gothenburg, Sweden article info Article history: Received 14 December 2009 Received in revised form 4 May 2010 Accepted 10 May 2010 Available online 9 June 2010 Keywords: Melanophore Fish Antifouling Pigment abstract Medetomidine is a new antifouling agent, and its effects in non-target aquatic organisms have been investigated. Earlier short-term studies in fish have shown a skin lightening response to medetomidine, but effects after chronic exposure have not been studied. In fish, the dark pigment melanin is contained within specialized cells, melanophores. Medetomidine binds to the melanophore a 2 -adrenoceptor, which stimulates pigment aggregation resulting in the light appearance. In the present study, rainbow trout (Oncorhynchus mykiss) was long-term exposed to 0.5 and 5.0 nM of medetomidine via water for 54 d. The fish were then photographed for paleness quantification and the images were analyzed using ImageJ analysis software. Additionally, scales were removed and used for in vitro function studies of the mela- nophores, monitoring the response to melanophore stimulating hormone (MSH) and subsequent mede- tomidine addition. The number of melanophores was also investigated. As a result of the medetomidine exposure, fish from the 5 nM treatment were significantly paler than control fish and the melanophores from these fishes were also more aggregated. Melanophores from all the treatments were functional, responding to MSH by dispersion and to subsequent medetomidine by aggregation. However, the results indicate a difference in sensitivity among treatments. The number of melanophores in the scales did not change significantly after long term exposure to medetomidine. These results suggest that the observed paleness may be reversible, even after chronic exposure. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction 1.1. A new antifouling agent Many commercially available antifouling paints contain biocides known to have adverse effects in the aquatic environment (Voulvou- lis et al., 2002; Konstantinou and Albanis, 2004; Thomas and Brooks, 2009). These findings have led to stricter legislation and intensive search for better alternatives. Medetomidine is a new antifouling agent, presently being evaluated according to the European Union Biocidal Products Directive. It is directed against barnacles and func- tions by altering the settling behavior. In Balanus improvisus settling was reversibly inhibited by medetomidine concentrations between 1 and 5 nM (0.2 and 1.0 lgL À1 ), while lethality occurred from 100 lM(Dahlström et al., 2000). This makes medetomidine promis- ing from an environmental perspective. Originally, medetomidine was introduced as an anesthetic and analgesic drug. Today it is widely used in veterinary medicine and wildlife management. It has been used in a variety of species, including amphibians (Brenner et al., 1994), reptiles (Bennett, 1998) and fish (Horsberg et al., 1999). In mammals, it is known to function as a potent and specific a 2 -adrenoceptor agonist, which makes it a useful tool in pharmacological studies (Savola et al., 1986; Virtanen et al., 1988). In fish, medetomidine has earlier been used in studies of pigment cells, since medetomidine induces paleness (Karlsson et al., 1989). The skin lightening response to medetomidine has also been used as a marker to demonstrate a 2 -adrenergic function in zebrafish (Ruuskanen et al., 2005). Medetomidine, or 4-[1-(2,3-dimethylphenyl)ethyl)-imidazole, has a molecular weight of 200.28 g mol À1 . Studies in marine Myti- lus edulis, Abra nitida, Crangon crangon and periphyton communi- ties have shown bioconcentration factors ranging from 2.8 to 1195 L kg À1 fresh weight (Hilvarsson et al., 2009). The half-life of mede- tomidine in rainbow trout at 10 °C is 5.5 h (Horsberg et al., 1999). 1.2. Colour and melanophore function in fish Fish colour has a number of different functions, where camou- flage and communication seem to be the most important (Bagnara 0045-6535/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2010.05.014 * Corresponding author. Address: Department of Zoology/Zoophysiology, University of Gothenburg, Box 463, SE 405 30 Göteborg, Sweden. Tel.: +46 31 786 36 83; fax: +46 31 786 38 07. E-mail addresses: anna.lennquist@zool.gu.se (A. Lennquist), lena.lindblad@zool. gu.se (L.G.E. Mårtensson Lindblad), daniel.hedberg@zool.gu.se (D. Hedberg), erik.kristiansson@zool.gu.se (E. Kristiansson), lars.forlin@zool.gu.se (L. Förlin). Chemosphere 80 (2010) 1050–1055 Contents lists available at ScienceDirect Chemosphere journal homepage: www.elsevier.com/locate/chemosphere