Induction of c-fos transcription in the medaka brain (Oryzias latipes) in response to mating stimuli

https://doi.org/10.1016/j.bbrc.2010.11.143Get rights and content

Abstract

Immediate-early genes (IEGs) are useful for mapping active brain regions in various vertebrates. Here we identified a c-fos homologue gene in medaka and demonstrated that the amounts of c-fos transcripts and proteins in the medaka brain increased in relation to an artificially evoked seizure, suggesting that the homologue gene has the characteristics of IEGs, which are used as markers of neural activity. Next, quantitative reverse-transcription-polymerase chain reaction revealed that female mating behaviors upregulated c-fos transcription in some brain regions including the telencephalon, optic tectum, and cerebellum. In addition, we performed in situ hybridization with a c-fos intron probe to detect the de novo synthesis of c-fos transcripts and confirmed induction of c-fos transcription in these brain regions after mating. This is the first report of IEG induction in response to mating stimuli in teleost fish. Our results indicated that c-fos expression was induced in response to behavioral stimuli in the medaka brain and that medaka c-fos could be a useful marker of neural activity.

Research highlights

c-fos expression in the medaka brain was induced by convulsant agent treatment within 30-min. ► c-fos expression was induced in response to mating stimuli in the medaka brain. ► Medaka c-fos is a useful marker of neural activity.

Introduction

The medaka (Oryzias latipes) is a freshwater teleost fish native to East Asia that has long been an ornamental fish in Japan. Medaka exhibit various social interactions, such as schooling behavior [1], [2], aggressive behavior [3], and a female mating preference for large males [4]. The medaka mating behaviors, for example, comprise a series of behavioral steps. First, the male medaka approaches the female and swims underneath her [5], [6]. Then the male swims rapidly in a small circle, which is called a “quick circle”. If the female is receptive to the male courtship display, the fish copulate by crossing their cloaca. As the medaka is a model organism for molecular genetics [7], functional analysis of the neural circuits involved in social interactions using advanced genetic methods will contribute to a better understanding of the neural/molecular basis underlying vertebrate social interactions.

Immediate-early gene (IEG) expression is induced in neurons by stimuli naturally associated with behaviors and the localization of IEG expression is a useful marker of neural activity. Based on IEG expression, brain regions that are active in response to (social) behavioral stimuli have been identified in vertebrate brains, such as rodents [8], songbirds, [9], and anurans [10]. Brain regions associated with mating behavior have been successfully mapped. For example, mating in rodent significantly increases the number of Fos-immunoreactive neurons in several brain regions, including the medial preoptic area, bed nucleus of the stria terminalis, medial amygdala, hypothalamic ventromedial nucleus, subparafascicular thalamic nucleus, and midbrain central tegmental field [8], [11]. Although IEG mapping studies have been extensively performed in mammalian and avian model systems, similar analyses in fish have been limited to a few studies in teleost fish [12], [13], [14]. Here we identified a medaka c-fos homologue gene and demonstrated that IEG expression was induced in response to mating stimuli.

Section snippets

Fish

Medaka fish (O. latipes, dr-R strain) were maintained in like groups in plastic aquariums (12 cm × 13 cm × 19 cm). Sexually mature male and female adult medaka fish (more than 3 months after hatching) were used for the cDNA cloning, real-time PCR, Western blotting, and in situ hybridization studies.

Mating condition

The adult female and male medaka fish were separated by two tanks overnight, prior to mating (Fig. 4A). The next morning, the male and female were placed together in a single tank and then the pair began to

Medaka c-fos homologue has characteristics of an immediate-early gene

To search for active regions in the medaka brain, we focused on the c-fos gene as an IEG that is transiently expressed in active neuronal cells. We found an exon encoding a putative medaka c-fos homologue in the medaka genome database and isolated a full-length cDNA using the 5′-and 3′-rapid amplification of the cDNA ends (RACE) method (Genbank No. AB572350). The open reading frame encoded 364 amino acids, which had the highest identities (57% and 60%) with fos homologues in mouse (Genbank No. NM_010234

Discussion

Here we identified a c-fos homologue gene in medaka and characterized the time course of its gene expression after pharmacologic stimulation. The c-fos expression peaked at 30-min post-induction and was markedly reduced at 90 min (Fig. 1A). The time course of c-fos mRNA induction was similar to that in mammals [24]. A small increase in c-fos expression was also detected in control fish (Fig. 1A), which might be due to handling during transfer. Furthermore, qRT-PCR and in situ hybridization

Acknowledgments

We thank the Medaka National BioResource Project for providing the medaka strains. This work was supported by the Ministry of Education, Culture, Sports, Science, and Technology, Scientific Research on the Grant-in-Aid (Wakate B). The first author was supported by Grant-in-Aid for JSPS Fellows. This study was carried out under the NIBB Cooperative Research Program (10-104).

References (28)

  • H. Imada et al.

    Coordinated and cohesive movement of two small conspecific fish induced by eliciting a simultaneous optomotor response

    PLoS ONE

    (2010)
  • K. Nakayama et al.

    Effects of polychlorinated biphenyls on the schooling behavior of Japanese medaka (Oryzias latipes)

    Environ. Toxicol. Chem.

    (2005)
  • J.J. Magnuson

    An analysis of aggressive behaviour, growth, and competition for food and space in medaka (Oryzias latipes)

    Can. J. Zool.

    (1962)
  • T. Ono et al.

    Mating ethogram in Oryzias latipes

    J. Fac. Sci. Hokkaido. Univ.

    (1957)
  • Cited by (0)

    View full text