Morphological and functional studies on the epidermal cells of amphioxus (Branchiostoma belcheri tsingtauense) at different developmental stages

Epidermal cells of amphioxus at different developmental stages were investigated by electron microscopy and colloidal carbon tracing experiments. Amphioxus epidermal cells showed different ultrastructural characteristics at larval and adult stages. The epidermal cells at all larval stages studied (24–96 h) had numerous vesicles containing electron dense materials in their apical cytoplasm. In tracing experiments, carbon particles were found in apical vesicles and interoellular spaces. Under scanning electron microscope, many crater-like protrusions were observed on the surface of the cells. These results indicated that amphioxus larval epidermal cells may be capable of endocytosis. The epidermal cells of 3-month and adult amphioxus were obviously secretory ones characterized by well-developed peripheral filaments, a prominent Golgi apparatus and abundant apical secretory vesicles. This study also showed that adult amphioxus body surface mucus contained lectin that could agglutinate human red blood cells. The authors propose that the epidermal cells of amphioxus larva and adult may contribute to the immune defense of the amimal by different means. Project 3860811 supported by NSFC and study also supported by the Shandong Natural Science Foundation (Grant No. 92D1144).     

Laboratory observation on spawning,fecundity and larval development of amphioxus (Branchiostoma belcheri Tsingtaunese)

Although amphioxus is widespread in temperate and tropical seas, its population is diminishing because of environmental pollution. To keep the population of this evolutionarily important animal from diminishing, study on its reproduction and development is necessary. The main findings in this study on the spawning and fecundity of the amphioxus reared in laboratory and its larval development are as follows. 1. Water temperature markedly affected the spawning. It spawned only when water temperature reached 21°C. 2. Spawning of the amphioxus in laboratory was markedly extended. Initially, the amphioxus spawned at about 7:00 PM, but spawning time was postponed as spawning days went on. 3. The number of eggs produced by a female ranged from 1400 to 12800, average of 5800. This also represents the fecundity of the amphioxus because it shedded all eggs within the ovary at a time. 4. During the first few months of life of the amphioxus, its growth rate changed seasonally. The growth rate in summer and fall was greater than that in winter. 5. The pelagic larva became a benthic adult after 50 days. 6. The amphioxus reared in laboratory from fertilized eggs could produce fertile eggs and sperms. These findings can be a foundation for measures to address the problem of diminishing amphioxus population. Contribution No. 2274 from the Institute of Oceanology, Chinese Academy of Sciences This work was supported by CNNSF, and Chinese Academy of Sciences.     

花胚胎发育及仔鱼阶段同工酶的初步分析

<正>鱼类胚胎发育是一个复杂而有序的系统工程,是基因时空综合表达结果。已有的研究证明:机体在生长发育过程中会发生一系列复杂的同工酶时空表达变化,它是基因特异性表达的一种主要作用形式。同工酶研究是研究鱼类个体发育的有力手段之一,     

Developmental expression pattern of calmodulin gene in amphioxus Branchiostoma belcheri tsingtauense

We investigated the developmental expression pattern of AmphiCaM in cephalochordate amphioxus (Branchiostoma belcheri tsingtauense). We cultured and sampled the animals at different developmental stages (eggs and larvae), and used in-situ hybridization and northern blotting to document the spatial and temporal changes in AmphiCaM expression. The alimentary tract dominates the development from the late neurula stage to the adult stage. AmphiCaM expression increased significantly in the alimentary tract during the late neurula stage and remained elevated in the adults. Our results indicate that AmphiCaM is involved in the differentiation of the alimentary tract in amphioxus; and furthermore, provide an insight into the change in function of CaM genes during evolution.     

黑脊倒刺鲃仔稚鱼摄食的习性与生长

研究了养殖条件下黑脊倒刺仔稚鱼摄食习性与生长,结果表明:仔鱼4日龄开口摄食,全长9.0~9.99mm;仔稚鱼的摄食具有明显的昼夜节律性;仔稚鱼个体间生长速度具有一定的差异性;其体重与摄食量的关系为y=0.064 4X+1.433 1,全长与日龄的关系为L=8.861 4×exp(0.042 9T),体重与日龄的关系为W=3.736×exp(0.151 3T),全长与体重的关系为W=1.7×10-3×L3.531 4。     

Toxicity of organophosphorus pesticide sumithion on larval stages of stinging catfish <Emphasis Type="Italic">Heteropneustes fossilis</Emphasis>

Sumithion is widely used to control brittle in paddy fields and tiger bug in fish larval rearing ponds. The objective of this study was to elucidate the toxic effects of sumithion on larval stages of stinging catfish Heteropneustes fossilis. Larvae were exposed to two concentrations (150 and 250 μg/L) of sumithion with one control in three replicates of each. Larvae samples were collected at 20- and 24-h intervals followed by observation under a digital microscope. Exposures of stinging catfish larvae to sumithion produced deformities including irregular head shape, lordosis, yolk sac edema, body arcuation, tissue ulceration, etc. The mortality rates of larvae were significantly increased in response to increase in sumithion concentrations. Furthermore, around 30% of the total adult stinging catfish reared in sumithiontreated aquaculture ponds were found to be deformed permanently. These findings highlight that exposure of stinging catfish to sumithion at the critical and sensitive stages in their life cycle may significantly reduce the number of returning adults. Therefore, the use of sumithion for crop protection needs to be considered carefully and alternatives to sumithion should to be developed for controlling aquatic insects in aqua-ponds during larval rearing.     

The Gastric Shield and the Undrelying Epithelium of Chlamys Farreri:Morphological,Histological and Histochemical Characterizations

The gastric shield and underlying stomach epithelium of Chlymas ffarreri were examined at the light and ultrastructural levels.The results showed that the gastric shield consisted of two different size lobes joined together by a narrow middle piece,the thicker lobe was shaped like a funnel,but unclosed at the lateral side;the other lobe was irregularly triangular-shaped.The transverse section of the thicker lobe was obviously laminated and gradually decreased in thickness from the peak to the margins of the shield.The underlying epithelium bore numerous about 3μm diameter spherical processes formed by the apical plasmalemma of the epithelial cells becoming blunt pseudopodia.Microvilli and some interspersed cilia were present in the areas among the spherical processes regions where only microvilli existed.Rough endoplasmic reticulum,Golgi apparatus,different-sized electron-dense secretory granules and electron-lucent vacuoles as well las abundant mitochondria were present in the underlying epithelial cells.Fused droplets of the secretion from the underlying epithelial cells formed the gastric shield.     

Antioxidant enzyme activities in different genders and tissues of amphioxus Branchiostoma belcheri tsingtauense

Information regarding antioxidant enzymes in amphioxus remains lacking, and this study was carried out to examine the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) in different genders and tissues of amphioxus Branchiostoma belcheri tsingtauense. Results show that (1) CuZn-SOD, CAT and GPX activities in the whole amphioxus B. belcheri tsing- tauense were basically at the same levels in male and female amphioxus, whereas both T-SOD and Mn-SOD activities in male amphioxus were significantly higher than that in the female (P<0.05); (2) The testis had significantly higher T-SOD and CuZn-SOD activities than the ovary (P<0.05); (3) CuZn-SOD activity was undetectable in the guts of male and female amphioxus; (4) For both male and female am- phioxus, the activities of CAT and GPX in the gonads including testis and ovary were the lowest (P<0.05) among the tissues examined; (5) The gut and gill had the same level GPX activities while the gut had a higher CAT activity; (6) There was no clear difference in CAT and GPX activities in the corresponding tissues between male and female amphioxus. The study on SOD, CAT and GPX activities in different genders and tissues of the protochordate provides data for future comparison of amphioxus antioxidant enzymes with those of invertebrates and vertebrates.     

The Gastric Shield and the Underlying Epithelium of Chlamys Farreri:Morphological, Histological and Histochemical Characterizations

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INDUCTION OF ECTODERMAL CELLS FROM VEGETAL- ENDODERMAL BLASTOMERES OF AMPHIOXUS(BRANCHIOSTOMA BELCHERI TSINGTAUNESE) EMBRYOS BY THE CALCIUM IONOPHORE A23187

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Induction of ectodermal cells from vegetal-endodermal blastomeres of amphioxus (Branchiostoma belcheri tsingtaunese) embryos by the calcium ionophore A23187

Timing of vegetal-endodermal cell determination in amphioxus embryos remains uncertain. We tentatively tested effects of A23187, the calcium ionophore, on the development of vegetal blastomeres isolated at the 16-cell stage. It was found that when vegetal blastomeres committed to endoderm were treated with A23187 prior to gastrulation, they were transformed into ectodermal cells as evidenced by the cell morphology and function characteristic of epidermis. However, the developmental fate of the same blastomeres untreated or treated with DMSO at the same stage or of those treated with A23187 after gastrulation remained unchanged. Thus, vegetal-endodermal cells in amphioxus embryos are not irreversibly determined before the gastrula stage, and artificial increase in intracelluar Ca²⁺ concentration can induce transdetermination of the predetermined endodermal cells into ectodermal cells. Project 39470091 supported by NSFC and partly supported by Shandong Natural Science Foundation, Grant No.93D0140.     

施氏獭蛤幼虫和稚贝发育及行为的研究

进行了施氏獭蛤(Lutraria sieboldii Reeve)的人工育苗、幼虫发育及行为观察实验。结果显示:在施氏獭蛤幼虫发育过程中,壳顶后期幼虫只有鳃丝及足出现,而没有眼点出现,发达的足和鳃丝是幼虫变态的标志;幼虫变态期明显的分为3～5d的爬行期和3～5d的附着期;爬行期幼虫有单条水管形成,附着期幼虫只有一条分叉的足丝,水管为单管型;底栖稚贝期开始形成两条水管,底栖稚贝早期的幼虫不具备潜沙能力,只进行爬行活动,随着水管的快速发育,幼虫逐渐具备潜沙能力,幼虫壳长达2500μm、水管长达6000μm以上时,最终具备和成体一样的完全潜沙能力。     

Initiation of primary cell culture from amphioxus Branchiostoma belcheri tsingtauense

Amphioxus, a cephalochordate, is an important model fish for studies in evolution and comparative biology. A successful cell culture from amphioxus tissues in vitro would help understanding some basic issues. To determine the optimal culture conditions for proliferation of amphioxus cells, primary cultures were initiated from buccal cirri, tail, gill, gut and metapleural fold of amphioxus Branchiostoma belcheri tsingtauense. The media tested were L-15, F-12, M 199, MEM, DMEM, PRMI 1640 and LDF, each was supplemented with 20% fetal bovine serum. The optimal conditions include tail tissue cultured in L-15 or F-12 with supplement of 20% FBS and 1.5% NaCl at about 25°C.     

凿贝才女虫发育阶段的形态观察

对寄生在九孔鲍 (HaliotisdiversicoloraquatilisReeve)贝壳上的凿贝才女虫 (Polydoraciliata)进行了观察 ,将其发育过程分为卵、胚胎、幼虫和成虫 4个阶段 ,并对这 4个阶段的形态结构进行了描述和讨论。由于凿贝才女虫寄生 ,九孔鲍贝壳内侧形成弯曲的管道和隆起 ,其外套膜被破坏 ,严重影响了九孔鲍生长 ,甚至导致其死亡。     

Initiation of primary cell culture from amphioxus <Emphasis Type="Italic">Branchiostoma belcheri tsingtauense</Emphasis>

Amphioxus, a cephalochordate, is an important model fish for studies in evolution and comparative biology. A successful cell culture from amphioxus tissues in vitro would help understanding some basic issues. To determine the optimal culture conditions for proliferation of amphioxus cells, primary cultures were initiated from buccal cirri, tail, gill, gut and metapleural fold of amphioxus Branchiostoma belcheri tsingtauense. The media tested were L-15, F-12, M 199, MEM, DMEM, PRMI 1640 and LDF, each was supplemented with 20% fetal bovine serum. The optimal conditions include tail tissue cultured in L-15 or F-12 with supplement of 20% FBS and 1.5% NaCl at about 25°C. Supported by Doctoral Initial Fund of Ludong University (No.43304)     

Response of regeneration diversity of <Emphasis Type="Italic">Carex Lasiocarpa</Emphasis> community to different water levels in Sanjiang Plain,China

The species diversity at the regeneration stage, inflenced by different water levels, is important for community composition in the later growing season. Regeneration diversity of Carex lasiocarpa community under different water levels was studied at two stages, recruitment and adult, in the Sanjiang Plain, Heilongjiang Province, China. The results showed that, at the two growing stages, important value of C. lasiocarpa population and species richness of the community decreased with the increasing water level, while the Simpson and Shannon-Wiener diversity indexes and Pielou evenness index increased. Under different water levels, community diversities were higher at the recruitment stage, while population important values of C. lasiocarpa were higher at the adult stage. Indexes in vegetation evaluation must be chosen prudentially for successful restoration and effective management of wetlands, and especially for wetland restoration, the optimal time should be selected according to the restoration objectives and costs.     

Response of Regeneration Diversity of Carex Lasiocarpa Community to Different Water Levels in Sanjiang Plain,China

The species diversity at the regeneration stage, inflenced by different water levels, is important for community composition in the later growing season.Regeneration diversity of Carex lasiocarpa community under different water levels was studied at two stages, recruitment and adult, in the Sanjiang Plain, Heilongjiang Province, China.The results showed that, at the two growing stages, important value of C.lasiocarpa population and species richness of the community decreased with the increasing water level, while the Simpson and Shannon-Wiener diversity indexes and Pielou evenness index increased.Under different water levels, community diversities were higher at the recruitment stage, while population important values of C.lasiocarpa were higher at the adult stage.Indexes in vegetation evaluation must be chosen prudentially for successful restoration and effective management of wetlands, and especially for wetland restoration, the optimal time should be selected according to the restoration objectives and costs.     

Interplay between amphioxus complement with sea bass macrophages: Opsonic activity of amphioxus humoral fluids

Previous studies have shown the existence of a complement system in the amphioxus Branchiostoma japonicum. However, whether it has an opsonic activity similar to that of vertebrates remains unknown. We demonstrated that the humoral fluid (HF) of amphioxus promoted the phagocytosis of yeast cells with sea bass (Lateolabrax japonicus) macrophages, whereas the C3-depleted and heated HF significantly lost the phagocytosis-promoting capacity. In addition, the precipitation of factor B (Bf) led to a marked loss of opsonic activity. Moreover, C3 fragments in the HF were found to bind to yeast cell surfaces. The results indicate that the amphioxus complement system is an important element involved in the opsonic activity, which promotes the sea bass macrophage phagocytosis by tagging yeast cells with C3 fragments via the activation of alternative complement pathway.     

MORPHOLOGICAL CHANGES IN DEVELOPMENT OF PHARYNGEAL TEETH IN MYLOPHARYNGODON PICEUS

Adult pharyngeal teeth in Mylopharyngodon piceus are molariform. Based on SEM observations ofthe developing teeth, this paper describes the morphological diversification of pharyngeal teeth in M.piceus. The larval and juvenile teeth are changed from conical to adult molariform teeth through sevenstages. Comparisons are made between each stage and corresponding types in some species of differentsubfamilies in Cyprinidae. It is considered that the ontogenetic resemblances of the pharyngeal teehbear relationship to the phylogeny of cyprinids.     

Developmental Dynamics of the Larval Muscle System of Bay Scallop (Argopecten irradians)

Though the larval development of bivalves has been extensively studied for commercial purposes,the dynamic development of larval muscle system remains largely unknown.In this study,we characterized the larval muscle system at different developmental stages(D-shaped veligers,umbo veligers and spats)in the bay scallop(Argopecten irradians)by phalloidin staining and under a confocal microscopy.The functional muscles are initially established at the early stage of veligers,which have four pairs of velar retractors and one anterior adductor.At the veliger stage,the velum and posterior retractor muscles are functionally important for velar contractility but undergo an irreversible shrink until they disappear at the end of the larval stage.During metamorphosis,three crucial modifications take place in the larval muscle system.The metamorphosis process involves the gradual degeneration of velum retractors,mantle margin development from an unfolded to a three-fold state,and remodeling of the adductor muscle system from dimyarian(two adductors)to monomyarian condition(one adductor)as in juveniles/adults.All retractor muscles are composed of striated muscle,but both anterior and posterior adductors have smooth and striated components.These findings highlight that the morphological changes at different stages are typical features of myogenesis in scallops.The present knowledge on the developmental dynamics of myogenesis in the bay scallop will not only improve our understanding of phenotypic diversity of larval myoanatomy in bivalves,but also provide useful information on the larval culture in hatcheries.