Rapid modulation of gene expression profiles in the telencephalon of male goldfish following exposure to waterborne sex pheromones Wudu E. Lado a,1 , Dapeng Zhang b , Jan A. Mennigen c , Jacob M. Zamora a , Jason T. Popesku d,2 , Vance L. Trudeau a,⇑ a Department of Biology, University of Ottawa, Ottawa, Canada b National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USA c Institut National de la Recherche Agronomique, UR Nutrition Aquaculture et Génomique, Nutrition Métabolisme et Aquaculture, Quartier Ibarron, 64310 St Pée-sur-Nivelle, France d Life Sciences Institute, Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada article info Article history: Available online 22 June 2013 Keywords: Microarray Ependymin Calmodulin Aldolase C GnRH Pheromones Sperm release abstract Sex pheromones rapidly affect endocrine physiology and behaviour, but little is known about their effects on gene expression in the neural tissues that mediate olfactory processing. In this study, we exposed male goldfish for 6 h to waterborne 17,20bP (4.3 nM) and PGF 2a (3 nM), the main pre-ovulatory and post-ovu- latory pheromones, respectively. Both treatments elevated milt volume (P = 0.001). Microarray analysis of male telencephalon following PGF 2a treatment identified 71 unique transcripts that were differentially expressed (q < 5%; 67 up, 4 down). Functional annotation of these regulated genes indicates that PGF 2a pheromone exposure affects diverse biological processes including nervous system functions, energy metabolism, cholesterol/lipoprotein transport, translational regulation, transcription and chromatin remodelling, protein processing, cytoskeletal organization, and signalling. By using real-time RT-PCR, we further validated three candidate genes, ependymin-II, calmodulin-A and aldolase C, which exhibited 3–5-fold increase in expression following PGF 2a exposure. Expression levels of some other genes that are thought to be important for reproduction were also determined using real-time RT-PCR. Expression of sGnRH was increased by PGF 2a, but not 17,20bP, whereas cGnRH expression was increased by 17,20bP but not PGF 2a . In contrast, both pheromones increase the expression of glutamate (GluR2a, NR2A) and c-aminobutyric acid (GABA A c2) receptor subunit mRNAs. Milt release and rapid modulation of neuronal transcription are part of the response of males to female sex pheromones. Ó 2013 Elsevier Inc. All rights reserved. 1. Introduction Chemical communication is important for intra-species com- munication. Vertebrates have evolved cryptic pheromonal signal- ling to help navigate the complex environment of cues while avoiding eavesdropping. One such example of signalling is sex pheromones, which vertebrates have used to signal reproductive status and readiness. This strategy obviates the need for species to spend time and energy inspecting one another for reproductive cues, thereby allowing them to expand their range for feeding while expending their energy on growth, development and preda- tor avoidance. Species-specificity of pheromone signalling is achieved through chemical form, size, polarity and blends depend- ing on biosynthetic and environmental constraints (Brennan and Zufall, 2006; Symonds and Elgar, 2008). Sex pheromones are important for goldfish reproduction be- cause they are oviparous and live in murky waters where visibility is greatly reduced. Therefore, in order for male and female goldfish to coordinate their reproduction, they have established an elabo- rate and sophisticated chemical communication system to aid in advertising their reproductive status to conspecifics (Stacey, 2003). The sex pheromones in goldfish are sex hormones that have been co-opted through evolution and given new functionality. For example, 17a,20b dihydroxy-4-pregnene-3-one (17,20bP) – the prevovulatory priming pheromone – is an oocyte maturation- inducing steroid hormone in females, and when it is released in the water acts on males to induce milt production and courtship (Sorensen et al., 1998; Stacey, 2003). In addition, prostaglandin- F 2a (PGF 2a ) is a female hormone that stimulates female sexual behaviour but when released to the water by the female it becomes a postovulatory releaser pheromone that triggers milt production and male sexual behaviour (Sorensen et al., 1988; Stacey et al., 2003). It is now well established that 17,20bP and PGF 2a are the primary sex pheromones controlling spawning in goldfish. Female goldfish, responding to environmental cues, release a priming pheromone (17,20bP and its metabolites) triggering an increase 0016-6480/$ - see front matter Ó 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.ygcen.2013.06.015 ⇑ Corresponding author. E-mail address: trudeauv@uottawa.ca (V.L. Trudeau). 1 Current address: Cell and Systems Biology, University of Toronto, 25 Harbord Street, Toronto, Ontario, Canada M5S 3G5. 2 Current address. General and Comparative Endocrinology 192 (2013) 204–213 Contents lists available at SciVerse ScienceDirect General and Comparative Endocrinology journal homepage: www.elsevier.com/locate/ygcen