Central distribution of kiss2 neurons and peri-pubertal changes in their expression in the brain of male and female red seabream Pagrus major Yuki Shimizu a,1 , Junko Tomikawa b,1 , Keisuke Hirano a , Yoko Nanikawa a , Yasuhisa Akazome c , Shinji Kanda c , Yukinori Kazeto d , Koichi Okuzawa e , Yoshihisa Uenoyama b , Satoshi Ohkura f , Hiroko Tsukamura b , Kei-ichiro Maeda g , Koichiro Gen d , Yoshitaka Oka c , Naoyuki Yamamoto a,⇑ a Laboratory of Fish Biology, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan b Laboratory of Reproductive Science, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan c Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan d Inland Station, National Research Institute of Aquaculture, Fisheries Research Agency, Tamaki, Mie 519-0423, Japan e National Research Institute of Aquaculture, Fisheries Research Agency, Minamiise, Mie 516-0193, Japan f Laboratory of Animal Production Science, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan g International Cooperation Center for Agricultural Education, Nagoya University, Nagoya 464-8601, Japan article info Article history: Received 19 July 2011 Revised 5 November 2011 Accepted 23 November 2011 Available online 7 December 2011 Keywords: Brain kiss2 Kisseptin Puberty Seabream Teleost abstract kisspeptins that are encoded by kiss1 gene are now considered the key regulator of reproduction from a number of studies in mammals. In most vertebrates, a paralogue of kiss1, called kiss2, is also present, and the functional significance of kisspeptins is not known precisely. In the present study, we have cloned kiss2 from a perciform teleost, the red seabream Pagrus major. The amino acid sequence deduced from the red seabream kiss2 contained a highly conserved 10-amino-acid residue, Kiss2(10) or kp-10. A kiss1-like transcript was also identified, but it appears to be non-functional due to the presence of a ‘‘premature’’ stop codon. Neurons that express kiss2 mRNA were distributed in the dorsal (NRLd) and ven- tral (NRLv) parts of nucleus recessi lateralis in the hypothalamus. In some fish a few kiss2-expressing neu- rons were detected in the preoptic area and nucleus ventralis tuberis. The number of kiss2-expressing neurons in the NRLd was larger during the first spawning season in both males and females compared with fish in the post-spawning periods. In males the number of kiss2 neurons in the NRLd of maturing fish was also larger than those in the post-spawning periods. In males the number of kiss2 neurons in the NRLv showed a similar pattern of changes to that of NRLd, while significant changes were not detected for females. The numbers of gonadotropin-releasing hormone 1 (GnRH1)-immunoreactive neu- rons in the preoptic area showed a similar pattern of change as those of kiss2 cells of the NRLd in both males and females (and also the NRLv in males). These results are in good agreement with the hypothesis that kiss2 neurons are involved in pubertal processes via regulatory influences on GnRH1 neurons in red seabream. Ó 2011 Elsevier Inc. All rights reserved. 1. Introduction In spite of the original identification of KISS1 as a metastasis suppressor gene in humans [19], the protein product of this gene or kisspeptin [16] that was first called metastin [29], is now recog- nized as the key molecule that regulates reproduction. In 2003, a mutation or knockout of the kisspeptin receptor gene (kissr1 or GPR54) was found to be associated with hypogonadotropic hypo- gonadism [5,35]. (We follow the terminology for genes and their product as proposed by a recent review [3].) Following these findings that imply critical roles of the kisspeptin-Kissr signaling pathway in the control of pubertal processes, a number of studies suggested that kisspeptin neurons mediate steroid feedback actions to gonadotropin-releasing hormone (GnRH) neurons, which results in an appropriate control on the reproductive func- tions through the hypothalamo–pituitary–gonadal axis in mam- mals [2,9,10,14,22,25,27,39]. In contrast to the situation in mammals where the physiology and functions of kisspeptin neurons are now fairly well understood, much remains to be studied regarding kisspeptins in non-mammalian vertebrates. In addition to kiss1 (medaka [12]; zebrafish and clawed toad [4]; zebrafish [41]; goldfish [42]), a paralogue of kiss1 called kiss2 has been found in a variety of non- mammalian (and one monotreme) species (lamprey, elephant shark, medaka, zebrafish, clawed toad, and platypus [20]; European seabass [6]; zebrafish and medaka [15]; goldfish [21]; chub 0016-6480/$ - see front matter Ó 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.ygcen.2011.11.038 ⇑ Corresponding author. Fax: +81 52 789 4083. E-mail address: nyama@agr.nagoya-u.ac.jp (N. Yamamoto). 1 These authors contributed equally to this work. General and Comparative Endocrinology 175 (2012) 432–442 Contents lists available at SciVerse ScienceDirect General and Comparative Endocrinology journal homepage: www.elsevier.com/locate/ygcen