Effects of early embryonic exposure to genistein on male copulatory behavior and vasotocin system of Japanese quail Carla Viglietti-Panzica, Elena Mura, GianCarlo Panzica ⁎ Laboratory of Neuroendocrinology, Neuroscience Institute of Torino (NIT), Department of Anatomy, Pharmacology and Forensic Medicine, University of Torino, Corso M. D’Azeglio 52, 10126, Torino, Italy Received 13 September 2006; revised 14 November 2006; accepted 4 December 2006 Available online 22 December 2006 Abstract Genistein is a phytoestrogen, particularly abundant in soybeans that can bind estrogen receptors and sex hormone binding proteins, exerting both estrogenic and antiestrogenic activity. In this study we used the Japanese quail embryo as a test end-point to investigate the effects of early embryonic exposure to genistein on male copulatory behavior and on vasotocin parvocellular system. Both differentiate by the organizational effects of estradiol during development and may therefore represent an optimal model to study the effects of xenoestrogens. We injected two doses of genistein (100 and 1000 μg) into the yolk of 3-day-old Japanese quail eggs. Other eggs were treated with either 25 μg of estradiol benzoate or sesame oil as positive and negative controls. At the age of 6 weeks, behavioral tests revealed a significant decrease of all aspects of copulatory behavior (in comparison to the control group) in estradiol-treated birds. In contrast, genistein-treated animals demonstrated various degrees of decrease in the mean frequencies of some aspects of the sexual behavior. The computerized analysis of vasotocin innervation in medial preoptic, stria terminalis and lateral septum nuclei revealed a statistically significant decreased immunoreactivity in treated animals compared to control ones. These results demonstrate that genistein, similarly to estradiol, has an organizational effect on quail parvocellular vasotocin system and on copulatory behavior. In conclusion, present results confirm, in this avian model, that embryonic exposure to phytoestrogens may have life-long effects on sexual differentiation of brain structures and behaviors. © 2006 Elsevier Inc. All rights reserved. Keywords: Phytoestrogens; Development; Avian brain; Bed nucleus of the stria terminalis; Organizational effects Genistein is the simplest isoflavonoid compound produced by Leguminosae, particularly abundant in soybeans. In plants, together with the other isoflavonoids collectively called phytoestrogens, it has antimicrobial activity (Dixon and Ferreira, 2002), as well as a specific activity to protect plants from several parasites (insects: Wang et al., 1999; Boué and Raina, 2003), this action is mediated by its activity as a ligand of ecdysone (the molt steroid hormone of invertebrates) receptor (Obertdorster et al., 2001). Genistein shares structural features with the 17β-estradiol (the phenolic ring and the distance between some hydroxyl groups); therefore, it can bind to vertebrate estrogen receptors (ERs) and sex hormone binding proteins (Kuiper et al., 1997; Morito et al., 2001). Thus, genistein and other similar phytoestrogens can exert both estrogenic and antiestrogenic activity, the latter by competing with estradiol for receptor binding (Bramlett et al., 2001). In addition, genistein is a tyrosine kinase inhibitor (Constantinou and Huberman, 1995) that can regulate the calcium-dependent phosphorylation processes controlling brain aromatase activity (Balthazart et al., 2003a). Due to their widespread occurrence, phytoestrogens are largely present in human and laboratory animal diets, where they range from 200 to 800 μg per gram, becoming a possible variable cause when examining reproductive and endocrine parameters in research studies (Lephart et al., 2002, 2005). Genistein, as well as other xenoestrogens, may directly influence the expression of estrogen-inducible genes (Leffers Hormones and Behavior 51 (2007) 355 – 363 www.elsevier.com/locate/yhbeh Abbreviations: AVPV, anteroventral periventricular nucleus; BST, bed nucleus of stria terminalis; CCM, cloacal contact movements; CNS, central nervous system; DES, diethylstilbestrol; EB, estradiol benzoate; FA, fractional area; GEN100, 100 μg genistein; GEN1000, 1000 μg genistein; M, mount; MA, mount attempt; NG, neck grab; OIL, sesame oil; PBS, phosphate-buffered saline; POM, medial preoptic nucleus; SL, lateral septum; SON, supraoptic nucleus; TH, tyrosine hydroxylase; VT, vasotocin; VT-ir, VT immunoreactivity. ⁎ Corresponding author. Fax: +39 011 2367970. E-mail address: giancarlo.panzica@unito.it (G. Panzica). 0018-506X/$ - see front matter © 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.yhbeh.2006.12.003