Changes to chloride and rodlet cells in gills, kidney and intestine of Dicentrarchus labrax (L.) exposed to reduced salinities

The effect of osmotic shock was investigated mainly in the chloride cells (CCs) and rodlet cells (RCs) of gills, and RCs of intestine and kidney of the European sea bass Dicentrarchus labrax obtained from a farm in the northern Adriatic Sea. During the experiment, fish were abruptly transferred from sea water to a salinity of 15 (15 SW) or to fresh water (FW). Numeric variation and ultrastructural changes of both cell types were evaluated at 24, 48 and 96 h after the transfer to lower salinity levels, using light and transmission electron microscopy (TEM). Exposure to FW produced a significant increase ( P < 0·05) in the number of branchial CCs and RCs within 96 and 24 h, respectively. Following osmotic challenge (either transfer to 15 SW or FW), kidney and intestine showed an evident increase in RC numbers. The cellular damage detected by TEM was the same for each sampling time (24, 48 and 96 h), but appeared more severe in fish exposed to FW (higher osmotic shock) than in those exposed to 15 SW. In RCs cytoplasmic vacuolizations, autophagosomes and autophagolysosomes with myelinoid bodies, dissolution and shrinkage of the typical inclusions were documented. Nevertheless, CCs showed vacuolization of endoplasmic reticulum and cytoplasmic dissolution and maintained the apical crypt typical of seawater acclimated fish. Renal tubular cells and intestinal epithelial cells showed similar changes to those reported for CCs and RCs.     

Rodlet cells in teleosts: a new insight into their nature and functions

The nature of rodlet cells (RCs) and their functions is subject to a number of different interpretations. This review provides a detailed analysis of the parasitic and endogenous origin of these cells. Two new functional aspects of RCs are considered in detail. The possible function of RCs as immune cells was derived from studies that reported an increase in the number of RCs in fish infected with protozoan and metazoan parasites, particularly at the site of the pathogen infection and/or attachment. Accordingly, RCs represent inflammatory cells, with a similar role to eosinophile granule cells, epithelioid cells and mesothelial cells. Rodlet cells may potentially act as biomarkers. Experimental studies that examined the response of RCs in fish exposed to chemical substances such as metals and herbicides reported an increase in the number of RCs in the tissues of the fish. Fish exposed to these substances expressed myelinic figures in the cytoplasm of the RCs and various degrees of rodlet degeneration and high vacuolization of RC cytoplasm were often noticed. Further lines of research are suggested that might elucidate the true function of these enigmatic cells.     

Seasonal changes in intra- and interorgan occurrence of rodlet cells in freshwater bream

Significant seasonal differences in intra- and interorgan prevalence, abundance and distribution of rodlet cells (RC) were observed in freshwater bream caught in May and August 1997 from a PCB-polluted lake, Lake Kernaala, Finland. In May, the majority of RCs in liver were seen in blood vessel endothelium, while in August most cells were located in the epithelium of the intrahepatic biliary ducts. In kidney, a significant increase in RCs in the columnar epithelium of the collecting ducts was observed from May to August. RCs in spleen were extremely rare in May, but were seen frequently in the endothelium of the splenic blood vessels in August. In gill and intestine, RCs were found in every tissue sample studied, with no statistical differences in their abundance or distribution between seasons. No RCs were seen in gonads. Total numbers of RCs increased dramatically from May to August in kidney and in spleen. The frequency of RCs corresponded strongly with the current phase of parasitic infections in liver, kidney, and spleen. It is concluded that RCs play a role in host defence mechanisms in fish, displaying induced expression at the times of peak parasitic prevalences.     

Distribution and characteristics of D-amino acid and D-aspartate oxidases in fish tissues

The distributions of D-amino acid oxidase (D-AAO, EC 1.4.3.3) and D-aspartate oxidase (D-AspO, EC 1.4.3.1) activities were examined on several tissues of various fish species. Both enzyme activities were commonly high in kidney and liver and low in intestine with some exceptions. After oral administration of D-alanine at 5 micromol /g body weight(-1)day(-1) to carp for 30 days, D-AAO activity increased by about 8-, 3-, and 1.5-fold in intestine, hepatopancreas, and kidney, respectively, whereas no increase was found in brain. In contrast, oral administration of D-glutamate or D-aspartate did not show any increase of D-AspO activity in any tissues. D-AAO and D-AspO of common carp kidney and hepatopancreas were subcellularly localized in peroxisomes, as clarified in mammals. D-proline was the best substrate for D-AAO in rainbow trout kidney, common carp kidney, and hepatopancreas, followed by D-alanine and D-phenylalanine. N-methyl-D-aspartate was the best substrate for D-AspO in rainbow trout kidney and common carp hepatopancreas. The optimal pH for D-AAO in rainbow trout kidney was broad, from 7.4 to 8.2, and that for D-AspO was around 10. D-AAO was inhibited by benzoate known as D-AAO inhibitor and D-AspO was strongly inhibited by meso-tartarate as D-AspO inhibitor. From these results, at least D-AAO in fish is considered to work as a metabolizing agent of exogenous and endogenous free D-alanine that is abundant in aquatic invertebrates such as crustaceans and bivalve mollusks, which are potential food sources of these fishes.     

二龄草鱼脾脏、肝脏组织高表达甘露糖结合凝集素mRNA

Innate immunity is expected to be very important in fish. Mannose-bingding lectin (MBL) participates in the innate immune system as an activator of the complement system and as an opsonin after binding to certain carbohydrate structures on microorganisms. In this experiment, total mRNA was isolated from spleen, liver, gills, thymus, head kidney and kidney of adult and immature grass carp Ctenopharygodon idllus. The cDNA of MBL was obtained by RT-PCR using total mRNA from the spleen of carp as template. Such cDNA was labled with ^32p and used as probe for Northern analysis, and autoradiographic signals were quantified by densitometry analysis. The results showed that MBL was high expressed in the spleen and liver and low in gills, thymus, head kidney and kidney of adult grass carp, and MBL was much lower expressed in spleen and liver of immature grass carp than those of adult grass carp. The results might partially explain why immature grass carp are vulnerable to grass carp hemorrhage virus (GCHV) whereas adult grass carp are not.This suggested that MBL mav be an imoortant anti-GCHV factor [Acta Zoologica Sinica 50 (1): 137 - 140. 2004].     

草鱼免疫相关基因Prkrip1的克隆和表达分析

随着草鱼养殖规模的扩大, 草鱼的病毒性疾病极大地影响着草鱼的产量。开展鱼类病毒免疫反应相关功能基因的研究意义重大。研究首先通过同源克隆的方法从草鱼中克隆到了一段Prkrip1基因的EST序列, 进一步通过RACE、长片段PCR和Genome walking的方法获得了该基因的全长cDNA序列、基因组DNA序列和启动子区序列。氨基酸序列分析显示, Prkrip1含有3个核定位信号和一个双链RNA结合区, 并具有与PKR结合的保守N端区; 荧光报告基因的表达证实我们所克隆到的启动子区是有活性的, 可用于后续该基因的转录调控分析; Real-time PCR分析发现, Prkrip1 基因在草鱼的肝和血中表达量最高, GCRV感染后在大部分免疫组织中均上调表达, 说明该基因确实与病毒感染相关。研究结果为Prkrip1基因在硬骨鱼类的功能研究提供了线索, 也为鱼类天然免疫反应中调控PKR信号通路的系统研究提供了理论依据。       

The effect of large doses of vitamin C and magnesium on stress responses in common carp, Cyprinus carpio.

1. Plasma magnesium, cortisol, lactate and ascorbic acid were examined in common carp subjected to various dietary treatments and following handling stress. 2. Under conditions of satisfied dietary magnesium and ascorbate requirements, plasma cortisol concentration after stress increased less pronouncedly than in fish fed large doses of ascorbate and/or magnesium. 3. Plasma lactate increased significantly in all groups after stress, although the increase seemed to be more severe (detrimental) in fish on large doses of ascorbate, either as ascorbic acid (AA) or ascorbic monophosphate Mg salt (AP). 4. Large doses of dietary ascorbate, both AA and AP, resulted in a significant increase of total ascorbate concentration in kidney and hepatopancreas of carp in comparison to pre-experimental level. 5. Kidney total ascorbate concentration decreased by 10-23% in all groups but one in which fish fed diet supplemented with AA displayed a significant increase (30%) of tissue ascorbate. The opposite trend was found in hepatopancreas of AA group with 21.5% ascorbate depletion. 6. The present results suggest that plasma cortisol and kidney (steroidogenesis site) and hepatopancreas ascorbate concentration responses to stress may not be related. Our results also do not support the hypothesis of the primary role of the high concentration of ascorbate in the kidney inhibiting steroidogenesis.     

A study of sequential histopathology of Trypanoplasma borreli (Protozoa: Kinetoplastida) in susceptible common carp Cyprinus carpio

The tissue response of common carp Cyprinus carpio to the kinetoplastid blood parasite Trypanoplasma borreli Laveran & Mesnil, 1901 was investigated during a laboratory infection of a highly susceptible carp line. With the development of the parasitaemia an increased proliferation of the lymphoid renal interstitial tissue was induced, which resulted in a progressive depression and deterioration of renal tubules. In heavily infected carp at Days 20 to 28 post inoculation (PI), a tubulonephrosis, a glomerulitis caused by a massive accumulation of leukocytes in glomerular capillaries, and large numbers of trypanoplasms in blood vessels and renal interstitium were observed. Corresponding with rising T. borreli numbers in the peripheral blood, splenic lymphocytes showed increasing proliferation rates, and the capillaries of the liver, gills, heart and intestine were infiltrated with lymphocytes and trypanoplasms. In heavily infected carp, congestion of liver sinusoids, focal necroses of hepatic tissue, extensive accumulations of erythrocytes in the spleen and in the blood marked anaemia were observed. These carp often showed abdominal distension, exophthalmus and swimming disorders described as 'sleeping sickness of carp'. Proliferation of cells from the interstitial lymphoid tissue of the kidney, which bears a close resemblance to the bone marrow of higher vertebrates, is considered a normal immune response of fish to antigen challenge. We here describe the unique case of a severe but ineffective immune reaction which results in the destruction of excretory renal structures. This has to be considered a severe disturbance of osmoregulation in affected carp, which, together with a decrease in oxygen uptake due to anaemia, is likely a major cause of death in these carp.     

Cloning, characterization, and expression analysis of Toll-like receptor-7 cDNA from common carp, Cyprinus carpio L.

The Toll-like receptor 7 (TLR7) is activated by single strand RNA and RNA-like compounds (imidazoquinoline), and it induces interferon production. We identified and described carp TLR7 cDNA and its mRNA expression. The full-length cDNA of carp TLR7 gene is 3427 bp, encoding 1049 amino acids (AB553573). The similarities of carp TLR7 with zebrafish, rainbow trout, fugu, and human TLR7 were 89.6, 83.4, 80.6 and 74.6%, respectively, at the amino acid sequence level. Furthermore, the expression of TLR7 mRNA was investigated in normal tissues of carp by semi-quantitative RT-PCR analysis. Carp TLR7 expression was exhibited in healthy tissues (kidney, brain, spleen, skin, intestine, muscle, liver, gills and heart) and though the expression level in each tissue varied among healthy fish. Carp TLR7 expression was significantly increased in head kidney stimulated with TLR7 agonist, imiquimod, at 8, 24 and 48 h in vitro when compared to expression in the control group. Moreover, carp head kidney leukocytes produced elevated levels of pro-inflammatory and type 1 interferon cytokine mRNA in response to imiquimod stimulation.     

Phylogeny and ontogeny of fish leucocytes

In contrast to higher vertebrates, most fish species hatch at the embryonic stage of life. Consequently, they have to defend against a variety of micro-organisms living in their aquatic environment. This paper is focussed on the development of leucocytes functioning within this early innate system and later on in the acquired immune system (B and T cells). Most of the data are derived from cyprinid fish (zebrafish, carp), which are excellent models to study early ontogeny. Attention is also paid to the phylogeny of leucocytes, with special attention to early chordates. It is clear that young fish use innate mechanisms during the first weeks/months of their development. In zebrafish, a variety of hematopoietic genes have been sequenced which allow a detailed picture of the development of the distinct leucocytes and their precursors. In cyprinids and sea bass, the thymus is the first lymphoid organ and T cells appear to be selected there much earlier than the first detection of T cell-dependent antibody responses. The first B cells are most probably generated in head kidney. Although T cells are selected earlier than B cells, T cell independent responses occur earlier than the T cell-dependent responses. The very early (pre-thymic) appearance of T-like cells in gut of sea bass and carp suggests an extra-thymic origin of these cells. However, B cells populate the GALT much later than spleen or kidney, indicating a rather late appearance of mucosal immunity. The first plasma cells are found long after the intake of food in cyprinids, but in many marine fish they appear around the first food intake. In general, acquired immunity is not correlated to food intake.     

Ichthyophthirius multifiliis (Fouquet) in the mirror carp, Cyprinus carpio L.

Mirror carp were infected with Ichthyophthirius multifiliis (Fouquet) under standardized conditions. The size and number of parasites at selected sites on the body were recorded during the course of the infection. Initial exposure to 40 mature parasites resulted in a mild infection with 100% recovery after 18 days. Recovered fish did not appear to be carriers of the parasite. Exposure to 400 parasites resulted in 100% mortality between 22–25 days. The growth rate of the parasite was linear. Parasites were more numerous in the dorsal surface of the fish than in the lateral or ventral surface. The increase in parasite numbers during the disease was greater in the gills than in the skin.     

Ontogeny of sexual development in the roach (Rutilus rutilus) and its interrelationships with growth and age

The roach (Rutilus rutilus) has become a sentinel species for the study of sexual disruption in wild fish populations as a consequence of exposure to endocrine disrupting chemicals (EDCs). Little is known, however, about the normal ontogeny of sexual development in this species. Here, we analyzed the ontogeny of sexual development in captive‐bred roach and assessed how growth rate and fish size affected the timing of both sexual differentiation and sexual development over a 2‐year period. Ovarian differentiation was first recorded at 68 days post‐fertilization (dpf) and this preceded testicular differentiation (first recorded at 98 dpf). In contrast, sexual maturation occurred at an earlier age in males (300 dpf) compared with females (728 dpf). No differences in body size (length or weight) were recorded between male and female roach until the fish were 415 dpf. Studies on three populations of roach which grew at different rates showed that the timing of sexual differentiation was highly variable and more related to fish size than to fish age. Time to sexual maturation was also variable among populations but, subsequent to their first year of life, gonadal status was less well associated with fish size. Interestingly, the sex ratio of the population was biased towards females in populations that grew more rapidly during early life. The findings presented here provide a valuable foundation of work to support both field‐ and laboratory‐based assessments on the effects of EDCs, and other stressors, on sexual differentiation and development in the roach. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.     

The effect of digested sludge on the common carp, Cyprinus carpio L.

The effects of sewage sludge on the survival of the common carp, Cyprinus carpio, were investigated. It was noted that, when the concentrations of the sludge used for rearing carp exceeded 0.6%, the fish died within 42 days, whereas those reared in 0.2% sludge remained alive and healthy during the trial period. The body and the gills of the dead fish were covered by a veil-like film which looked like coagulated mucus and was formed by the heavy metal ions from the sludge reacting with some constituents of the mucus secreted by the gill. The histopathological symptoms of the various organs further demonstrated that internal injury in liver and kidney was also an important feature of the intoxicants as well as external damage, especially the particles observed around the gills. It was concluded that the optimum concentration of sludge to any species of fish should be determined by testing a series of concentrations before using it as a supplementary fish feed or fish pond fertilizer.     

饲料丙二醛对草鱼生长、肝胰脏及肠道结构和功能的影响

为了探讨丙二醛(MDA)对草鱼Ctenopharyngodon idellus(74.821.49) g生长、肝胰脏及肠道结构和功能的影响并初步对比MDA与其他油脂氧化产物的毒副作用, 本试验以新鲜豆油、低氧化程度的鱼油为饲料脂肪源, 制成豆油组(S组)、鱼油组(F组), 并在豆油组中喷涂不同浓度的MDA, 制成MDA水平为61.59 (M1组)、123.92 (M2组)、185.04 (M3组) mg/kg的5种等氮等能的试验饲料。经72d池塘网箱养殖后, 试验结果显示:(1)饲料中MDA及油脂其他氧化产物均可显著增加草鱼饲料系数(FCR) (P0.05), 显著降低特定生长率(SGR)、蛋白质沉积率(PRR)(P0.05), MDA还可显著降低草鱼脂肪沉积率(LRR) (P0.05); (2)饲料中MDA及油脂其他氧化产物均可显著降低血清总胆汁酸(TBA)含量(P0.05), 并使血清总胆固醇(TC)、甘油三酯(TG)、MDA含量及超氧化物歧化酶(SOD)酶活性显著上升(P0.05), 饲料中MDA还可显著增加血清丙氨酸氨基转移酶(ALT)含量(P0.05), 显著降低血清高密度脂蛋白与低密度脂蛋白之比(HDL/LDL)、白蛋白与球蛋白之比(A/G)比值(P0.05); (3)饲料中MDA及油脂及其他氧化产物会显著增加肝胰脏脂肪(P0.05), 且MDA还会显著增加肝胰脏SOD含量(P0.05), 导致草鱼肝胰脏氧化应激; (4)饲料中MDA会损伤肝胰脏细胞线粒体, 降低细胞核数量, 使肝胰脏细胞有明显纤维化趋势; (5)饲料MDA及鱼油其他氧化产物均会引起草鱼肠道黏膜杯状细胞数量增加, 损伤肠道微绒毛, 并会损伤肠道紧密连接结构, 增加肠道通透性, 导致血清内毒素及D-乳酸含量显著增加(P0.05)。上述结果表明: (1)饲料MDA会引起草鱼鱼体应激, 并通过干扰正常胆汁酸循环来干扰草鱼对脂肪的消化吸收, 最终导致草鱼生长性能下降; (2)MDA会引起肝胰脏氧化应激, 并可通过损伤肝胰脏细胞线粒体内部结构来损伤草鱼肝胰脏, 增加其发生脂肪性肝炎机率, 而鱼油其他氧化产物则是通过影响线粒体膜结构改变线粒体形态来损伤肝胰脏; (3)饲料MDA及鱼油其他氧化产物均会损伤草鱼肠道绒毛和微绒毛来降低其消化吸收能力, 还可损伤肠道紧密连接结构, 增加肠道通透性。     

饲料氧化鱼油引起草鱼肠道结构损伤、通透性增加

为了探讨饲料氧化鱼油对草鱼(Ctenopharyngodon idellus)肠道组织结构及其通透性的影响, 本实验以豆油、鱼油及氧化鱼油作为饲料脂肪源, 分别设计鱼油组(6F)、豆油组(6S)、2%氧化鱼油(4S2OF)、4%氧化鱼油(2S4OF)及6%氧化鱼油(6OF) 5组等氮、等能的半纯化饲料。经72d池塘网箱养殖后, 实验结果显示: (1)氧化鱼油显著增加(P0.05)草鱼血清和肠道MDA含量、增加肠道GSH含量(P0.05), 但随氧化产物含量上升GSH含量出现下降。(2)氧化鱼油会显著降低肠道内胆汁酸的含量(P0.05)。(3)氧化鱼油会显著增加肠道绒毛中杯状细胞的数量(P0.05), 且随着氧化产物的增加, 肠道微绒毛高度呈现先上升后下降趋势。(4)氧化鱼油会导致肠道紧密连接间隙增大, 增加肠道通透性, 使血清中D-乳酸及内毒素含量显著增加(P0.05)。结果表明, 饲料中鱼油氧化产物损伤了草鱼肠道组织结构, 尤其是肠道上皮细胞紧密连接结构损伤严重, 从而破坏了肠道黏膜的机械屏障功能, 使肠道通透性显著增加, 肠道细菌内毒素等发生转移。鱼油氧化产物会引起草鱼肠道氧化与抗氧化应激反应, 干扰草鱼肝-肠正常胆汁酸循环, 致使草鱼肠道胆汁酸不足。     

Effects of Cortisol on the Intestinal Mucosal Immune Response during Cohabitant Challenge with IPNV in Atlantic Salmon (Salmo salar)

Infectious pancreatic necrosis virus (IPNV) causes high incidence of disease in salmonids during the first period after SW transfer. During this period as well as during periods of stress, cortisol levels increase and indications of a relationship between IPNV susceptibility and cortisol have been suggested. The intestine is an entry route and a target tissue for IPNV displaying severe enteritis and sloughing of the mucosa in infected fish. The mechanisms behind effects of the virus on the intestinal tissue and the impact of cortisol on the effect remain unclear. In the present study, Atlantic salmon post smolts treated with or without slow release cortisol implants were subjected to a cohabitant IPNV challenge. Analysis of genes and proteins related to the innate and acquired immune responses against virus was performed 6 days post-challenge using qPCR and immunohistochemistry. An increased mRNA expression of anti-viral cytokine interferon type I was observed in the proximal intestine and head kidney as a response to the viral challenge and this effect was suppressed by cortisol. No effect was seen in the distal intestine. T-cell marker CD3 as well as MHC-I in both intestinal regions and in the head kidney was down regulated at the mRNA level. Number of CD8α lymphocytes decreased in the proximal intestine in response to cortisol. On the other hand, mRNA expression of Mx and IL-1β increased in the proximal intestine and head kidney in IPNV challenged fish in the presence of cortisol suggesting that the immune activation shifts in timing and response pathway during simulated stress. The present study clearly demonstrates that IPNV infection results in a differentiated epithelial immune response in the different intestinal regions of the Atlantic salmon. It also reveals that the epithelial immune response differs from the systemic, but that both are modulated by the stress hormone cortisol.     

Egg Cortisol Exposure Enhances Fearfulness in Larvae and Juvenile Rainbow Trout

We investigated the effects of an early boost of cortisol exposure in rainbow trout (Oncorhynchus mykiss) eggs during fertilisation on subsequent behavioural responses when exposed to a sudden stimulus in larvae and juveniles. At 55 d post‐fertilisation (dpf), treatment had no effect on high accelerations occurring after a sudden event. At 146 dpf, these high accelerations were more frequent in cortisol‐treated fish than in controls. At 146 dpf also, swimming activity was increased in cortisol‐treated fish both before and after the sudden stimulus. This study underlines the important behavioural modifications in both larvae and juveniles, linked to a change in the surrounding environment of the embryo. Indeed, fish exposed to cortisol as eggs showed a higher level of fearfulness later in life. Our findings are of major interest for stress management in an aquaculture context and also allow for a better understanding of the long‐lasting effects of a permanent and/or acute stress – mediated by cortisol – that could be encountered by females, affecting population's life history trajectory.     

Ontogenic expression profiles of gonadotropins (fshb and lhb) and growth hormone (gh) during sexual differentiation and puberty onset in female zebrafish

In the zebrafish model, the ontogenic expression profiles of all pituitary hormones have been reported except gonadotropins, partly because they are not supposed to be expressed in the embryonic stage. The spatiotemporal expression patterns of gonadotropins, namely follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh), in this species therefore remain largely unknown. As the master hormones controlling reproduction, the information on this issue would be valuable for understanding the roles of gonadotropins in early sexual development. Using double-colored fluorescent in situ hybridization (FISH) and real-time quantitative PCR (qPCR), this study was undertaken to analyze the ontogenic expression patterns of FSHbeta (fshb) and LHbeta (lhb) subunits in the zebrafish pituitary, with particular emphasis on the stage of sexual differentiation (～25-30 dpf [days postfertilization]) and puberty onset (～45 dpf). As a control, growth hormone (gh) was also examined throughout the study. The zebrafish were collected at different time points of early development, including 4, 5, 6, 8, 10, 13, 16, 19, 22, 25, 28, 38, 48, and 53 dpf. The head of each fish, including the brain and pituitary, was sampled for double-colored FISH analysis, whereas the body was fixed for histological examination of sex and gonadal developmental stage. Our results showed that the expression of fshb started much earlier than that of lhb, with its mRNA signal detectable (～2-3 cells per pituitary) shortly after hatching (4 dpf). In contrast, lhb expression became detectable much later, at the time of sex differentiation (～25 dpf). In female zebrafish, the first morphological sign for puberty is the first wave of follicle transition from the primary growth to previtellogenic stage, which occurs around 45 dpf and is marked by the appearance of cortical alveoli in the oocytes. Interestingly, the number of lhb-expressing cells was very low (～5-6 cells per pituitary) before this transition but increased dramatically during and after the transition. In contrast, the expression of fshb was abundant before puberty, with only a slight increase in cell number during puberty onset. The increased expression of fshb and lhb at puberty was also supported by real-time qPCR analysis at the single pituitary level. Interestingly, the fshb-expressing cells changed their spatial distribution significantly during puberty, from a predominantly peripheral to a central location. As the control, the expression of gh was abundant throughout prepubertal and pubertal periods. Our results strongly suggest an important role for Lh at the puberty onset of female zebrafish, similar to the situation in mammals, and its expression could be a sign for puberty at the pituitary level. However, the significance of the location change of Fsh cells during this period will be interesting to investigate.