Sexually dimorphic distribution of kiss1 and kiss2 in the brain of yellowtail clownfish, Amphiprion clarkii

Abstract

Kisspeptin system was shown to be a key factor in mediating social stress and reproduction. Yellowtail clownfish, Amphiprion clarkii, is a hermaphrodite fish, whose sex determination and gonadal development are affected by the social status of individuals. The yellowtail clownfish is a fantastic animal model to explore sex determination, but the social status and precise distribution of kiss mRNAs in the brain of this species are unknown. Hererin, a novel in situ hybridization technique, RNAscope, was used to investigate the distribution of kiss1 and kiss2 expressions in the brain of yellowtail clownfish. The coronal planes of brain showed that the kiss1 signal was mainly present in dorsal habenular nucleus (NHd) and kiss2 mRNA was widely expressed in telencephalon, midbrain, and hypothalamus, especially in dorsal part of the nucleus of the lateral recess (NRLd). Additionally, kiss1 and kiss2 signals have sexually dimorphic distribution. The kiss1 mRNA was distributed in NHd, the telencephalon, and lateral part of the diffuse nucleus of the inferior lobe (NDLIl) of females but in NHd and NDLIl of males. kiss2 signals were stronger in females than that in males. The distribution of kiss1 and kiss2 neurons in NHd of habenula and NRLd of hypothalamus may suggest that kiss genes associate environmental signaling and reproductive function in yellowtail clownfish.

Keywords: RNAscope; kisspeptin; sexually dimorphic distribution; social stress; yellowtail clownfish.

Figure 1

Histological slices were used to distinguish females and males according to the level of gonadal development in yellowtail clownfish. (A) The histological slices of the female gonad; (B) the histological slices of the male gonad. O1, oocytes in primary growth stage; O2, oocytes in cortical vesicle stage; O3, oocytes in vitellogenesis stage; SSC, secondary spermatocytes; S, spermatozoon. Scale = 200 μm.

Figure 2

The sagittal distribution of kiss1 and kiss2 mRNA in female yellowtail clownfish brain. (A, B, C and D) The kiss1-expressed brain regions in sagittal planes of the female brain; (E, F, G, H, I, J, K and L) the kiss2 expressed brain regions in sagittal planes of the female brain; (M) negative control; (N) positive control; (O) the level of the slices in the sagittal drawing view of female yellowtail clownfish brain; (P) the distribution of kiss1 and kiss2 in the female brain of yellowtail clownfish in sagittal planes. Tel, telencephalon; Vam, medial division of valvula cerebelli; CER, cerebellum; Hyp, hypothalamus; OT, optic tectum; DIE, diencephalon; MO, medulla oblongata; Hyp-NRLd, dorsal part of the nucleus of the lateral recess. Scale = 20 μm.

Figure 3

The distribution of kiss mRNA in the female and male brains of yellowtail clownfish in coronal planes. Black circles, the distribution of kiss mRNA in the female brain; open circles, the distribution of kiss mRNA in the male brain; black triangles, the distribution of kiss mRNA in the female brain; open triangles,the distribution of kiss mRNA in the male brain. CCe, corpus of the cerebellum; Cp, central posterior thalamic nucleus; DIp, lateral posterior part of the dorsal telencephalic area; Dm2, subdivision 2 of the medial dorsal telencephalic area; Dm3, subdivision 3 of the medial dorsal telencephalic area; Dp, posterior portion of the dorsal telencephalon; NAPv, anterior periventricular nucleus; NDLIl, lateral part of the diffuse nucleus of the inferior lobe; NGp, posterior part of glomerular nucleus; NHd, dorsal habenular nucleus; NHv, ventral habenular nucleus; NLTI, inferior part of the lateral tuberal nucleus; NLTv, ventral part of the lateral tuberal nucleus; nMLF, nucleus of the medial longitudinal fasciculus; NPC, central pretectal nucleus; NPGa, anterior preglomerular nucleus; NPGc, commissural preglomerural nucleus; NPOav, anteroventral part of the parvocellular preoptic nucleus; NPOPc, parvocellular part of the parvocellular preoptic nucleus; NPT, posterior tuberal nucleus; NRLd, dorsal part of the nucleus of the lateral recess; NRLv, ventral part of the nucleus of the lateral recess; NRP, nucleus of the posterior recces; LFB, lateral forebrain bundle; OC, optic chiasm; OT, optic tectum; PCo, posterior commissure; PGZ, periglomerular gray zone; PMgc, gigantocellular part of the magnocellular preoptic nucleus; PSp, parvocellular superficial pretectal nucleus; TEG, tegmentum; TLa, nucleus of the torus lateralis; TLo, torus longitudinalis; TPp, periventricular nucleus of the posterior tuberculum; VCe, valvula of the cerebellum; VL, ventrolateral thalamic nucleus; VM, ventromedial thalamic nucleus; VOT, ventral optic tract; VP, postcommissural part of the ventral telencephalon.

Figure 4

The coronal distribution of kiss1 mRNA in female and male yellowtail clownfish brain. (A, B, C and D) and (I, J and K) The kiss1-expressed brain regions in coronal planes of the female brain; (E, F, G and H) and (M, N and O) the kiss1-expressed brain regions in coronal planes of the male brain; (L) negative control; (P) positive control. Dm3, subdivision 3 of the medial dorsal telencephalic area; Dm2, subdivision 2 of the medial dorsal telencephalic area; DIp, lateral posterior part of the dorsal telencephalic area; Dp, posterior portion of the dorsal telencephalon; NHd, dorsal habenular nucleus; CCe, corpus of the cerebellum; NDLIl, lateral part of the diffuse nucleus of the inferior lobe. Scale = 20 μm.

Figure 5

The coronal distribution of kiss2 mRNA in female and male yellowtail clownfish brain. (A, B, C, D, E and F) and (M, N, O, P and Q) the kiss2expressed brain regions in coronal planes of the female brain; (G, H, I, J, K and L) and (S, T, U, V and W) the kiss2-expressed brain regions in coronal planes of the male brain; (R) negative control; (X) positive control. Dm3, subdivision 3 of the medial dorsal telencephalic area; Dm2, subdivision 2 of the medial dorsal telencephalic area; DIp, lateral posterior part of the dorsal telencephalic area; Dp, posterior portion of the dorsal telencephalon; OT, optic tectum; CCe, corpus of the cerebellum; TEG, tegmentum; TPp, periventricular nucleus of the posterior tuberculum; NGp, posterior part of glomerular nucleus; NRLd, dorsal part of the nucleus of the lateral recess; NDLIl, lateral part of the diffuse nucleus of the inferior lobe. Scale = 20 μm.