New observations on green hydra symbiosis

New observations on green hydra symbiosis are described. Herbicide norflurazon was chosen as a "trigger" for analysis of these observations. Green hydra (Hydra viridissima Pallas, 1766) is a typical example of endosymbiosis. In its gastrodermal myoeptihelial cells it contains individuals of Chlorella vulgaris Beij. (KESSLER & HUSS 1992). Ultrastructural changes were observed by means of TEM. The newly described morphological features of green hydra symbiosis included a widening of the perialgal space, missing symbiosomes and joining of the existing perialgal spaces. Also, on the basis of the newly described mechanisms, the recovery of green hydra after a period of intoxication was explained. The final result of the disturbed symbiosis between hydra and algae was the reassembly of the endosymbiosis in surviving individuals.     

Modulatory action of 12-O-tetradecanoylphorbol-13-acetate on bud production inHydra

Summary A low concentration of 12-O-tetradecanoylphorbol-13-acetate (TPA, 1.0 ng/ml) induced a transient inhibition of bud production in hydra which were fed daily. However, when hydra were starved following TPA-treatment, they produced further buds. Phorbol (1.0 ng/ml) and dimethyl sulfoxide (0.001%) did not influence bud production under either feeding or starvation conditions. These results indicate that TPA modulates asexual reproduction in hydra.     

The aberrant,a morphological mutant of Hydra attenuata,has altered inhibition properties

Transplantation experiments have led to the idea that developmental gradients are involved in the patterning processes of hydra. Head activation and head inhibition gradients appear to be involved in head formation. More could be learned about pattern formation if these gradients could be altered in the animal. One approach to this end is to study mutants suspected of having altered gradients. Such mutants may be identified by their altered morphology. In this investigation the aberrant, a morphological mutant of Hydra attenuata, was examined for alterations in its gradients. The aberrant was found to have changes in its inhibition and polarity properties. The inhibition gradient was steeper than normal and the level of inhibition was higher than normal near the head. In addition, the amount of inhibition transmitted from the head was greater than normal, while the amount of inhibition transmitted through the gastric region was less than normal. The polarity of the aberrant gastric region was more difficult than normal to reverse, and the aberrant extremities caused faster polarity reversal than normal extremities. The significance of these changes is discussed in relation to pattern formation.     

Infection of Hydra by Microsporidia

A microsporidian infection in a laboratory clone of Hydra littoralis has been observed, and the parasite has been tentatively identified as a species of Plistophora . Infected hydra continue to bud and regenerate normally and show no significant physiological or morphological changes. Sexual crossing of infected and non-infected animals shows that the infection is transmitted by the ovum but not by the sperm. Continuous exposure of infected hydra to Fumidil B in solution resulted in the disappearance of all Plistophora spores after a five week period of treatment, and the clones of the treated animals have remained parasite-free for more than a year.     

Molecular, biochemical and functional analysis of a novel and developmentally important fibrillar collagen (Hcol-I) in hydra

The body wall of hydra (a member of the phylum Cnidaria) is structurally reduced to an epithelial bilayer with an intervening extracellular matrix (ECM). Previous studies have established that cell-ECM interactions are important for morphogenesis and cell differentiation in this simple metazoan. The ECM of hydra is particularly interesting because it represents a primordial form of matrix. Despite progress in our understanding of hydra ECM, we still know little about the nature of hydra collagens. In the current study we provide a molecular, biochemical and functional analysis of a hydra fibrillar collagen that has similarity to vertebrate type I and type II collagens. This fibrillar collagen has been named hydra collagen-I (Hcol-I) because of its structure and because it is the first ECM collagen to be identified in hydra. It represents a novel member of the collagen family. Similar to vertebrate type I and II collagens, Hcol-I contains an N-terminal propeptide-like domain, a triple helical domain containing typical Gly-X-Y repeats and a C-terminal propeptide domain. The overall identity to vertebrate fibrillar collagens is about 30%, while the identity of the C-terminal propeptide domain is 50%. Because the N-terminal propeptide domain is retained after post-translational processing, Hcol-I does not form thick fibers as seen in vertebrates. This was confirmed using transmission electron microscopy to study rotary shadow images of purified Hcol-I. In addition, absence of crucial lysine residues and an overall reduction in proline content, results in reduced crosslinking of fibrils and increased flexibility of the molecule, respectively. These structural changes in Hcol-I help to explain the flexible properties of hydra ECM. Immunocytochemical studies indicate that Hcol-I forms the 10 nm fibrils that comprise the majority of molecules in the central fibrous zone of hydra ECM. The central fibrous zone resides between the two subepithelial zones where hydra laminin is localized. While previous studies have shown that basal lamina components like laminin are expressed by the endoderm, in situ hybridisation studies show that Hcol-I mRNA expression is restricted to the ectoderm. Hcol-I expression is upregulated during head regeneration, and antisense studies using thio-oligonucleotides demonstrated that blocking the translation of Hcol-I leads to a reversible inhibition of head morphogenesis during this regenerative process. Taken in total, the data presented in this study indicate that Hcol-I is required for morphogensis in hydra and represents a novel fibrillar collagen whose structural characteristics help to explain the unique biophysical properties of hydra ECM. Interestingly, the structure of Hcol-I mimics what is seen in Ehlers-Danlos syndrome type VII in humans; an inherited pathological condition that leads to joint and skin abnormalities. Hcol-I therefore illustrates an adaptive trait in which the normal physiological situation in hydra translates into a pathological condition in humans.     

Both the epithelial cells and the nerve cells are involved in the head inhibition properties in Hydra attenuata

The aberrant, a morphological mutant of Hydra attenuata, has been shown to have altered inhibition properties(D. I. Rubin and H. R. Bode, 1982, Develop. Biol. 89, 316–331). Based on transplantation experiments, the inhibition gradient in the aberrant is steeper and higher near the head than that in the normal. Also, the amount of inhibition transmitted by the aberrant head is higher. Both epithelial cells and nerve cells have been implicated in the patterning process of hydra. The cell types involved in the altered inhibition properties of the aberrant were determined by making chimeras consisting of epithelial cells from one strain and nerve cells from the other. Experiments with these chimeras demonstrated that the epithelial cells were responsible for the altered inhibition gradient of the aberrant. In contrast, both the epithelial cells and the nerve cells were involved in the higher amount of inhibition transmitted by the aberrant head. Thus, both cell types are involved in this aspect of patterning.     

Epithelial morphogenesis in hydra requires de novo expression of extracellular matrix components and matrix metalloproteinases

As a member of the phylum Cnidaria, the body wall of hydra is organized as an epithelium bilayer (ectoderm and endoderm) with an intervening extracellular matrix (ECM). Previous studies have established the general molecular structure of hydra ECM and indicate that it is organized as two subepithelial zones that contain basement membrane components such as laminin and a central fibrous zone that contains interstitial matrix components such as a unique type I fibrillar collagen. Because of its simple structure and high regenerative capacity, hydra has been used as a developmental model to study cell-ECM interaction during epithelial morphogenesis. The current study extends previous studies by focusing on the relationship of ECM biogenesis to epithelial morphogenesis in hydra, as monitored during head regeneration or after simple incision of the epithelium. Histological studies indicated that decapitation or incision of the body column resulted in an immediate retraction of the ECM at the wound site followed by a re-fusion of the bilayer within 1 hour. After changes in the morphology of epithelial cells at the regenerating pole, initiation of de novo biogenesis of an ECM began within hours while full reformation of the mature matrix required approximately 2 days. These processes were monitored using probes to three matrix or matrix-associated components: basement membrane-associated hydra laminin beta1 chain (HLM-beta1), interstitial matrix-associated hydra fibrillar collagen (Hcol-I) and hydra matrix metalloproteinase (HMMP). While upregulation of mRNA for both HLM-beta1 and Hcol-I occurred by 3 hours, expression of the former was restricted to the endoderm and expression of the latter was restricted to the ectoderm. Upregulation of HMMP mRNA was also associated with the endoderm and its expression paralleled that for HLM-beta1. As monitored by immunofluorescence, HLM-beta1 protein first appeared in each of the two subepithelial zones (basal lamina) at about 7 hours, while Hcol-I protein was first observed in the central fibrous zone (interstitial matrix) between 15 and 24 hours. The same temporal and spatial expression pattern for these matrix and matrix-associated components was observed during incision of the body column, thus indicating that these processes are a common feature of the epithelium in hydra. The correlation of loss of the ECM, cell shape changes and subsequent de novo biogenesis of matrix and matrix-associated components were all functionally coupled by antisense experiments in which translation of HLM-beta1 and HMMP was blocked and head regeneration was reversibly inhibited. In addition, inhibition of translation of HLM-beta1 caused an inhibition in the appearance of Hcol-I into the ECM, thus suggesting that binding of HLM-beta1 to the basal plasma membrane of ectodermal cells signaled the subsequent discharge of Hcol-I from this cell layer into the newly forming matrix. Given the early divergence of hydra, these studies point to the fundamental importance of cell-ECM interactions during epithelial morphogenesis.     

A novel hydra matrix metalloproteinase (HMMP) functions in extracellular matrix degradation, morphogenesis and the maintenance of differentiated cells in the foot process

As a member of Cnidaria, the body wall of hydra is structurally reduced to an epithelial bilayer with an intervening extracellular matrix (ECM). Biochemical and cloning studies have shown that the molecular composition of hydra ECM is similar to that seen in vertebrates and functional studies have demonstrated that cell-ECM interactions are important to developmental processes in hydra. Because vertebrate matrix metalloproteinases (MMPs) have been shown to have an important role in cell-ECM interactions, the current study was designed to determine whether hydra has homologues of these proteinases and, if so, what function these enzymes have in morphogenesis and cell differentiation in this simple metazoan. Utilizing a PCR approach, a single hydra matrix metalloproteinase, named HMMP was identified and cloned. The structure of HMMP was similar to that of vertebrate MMPs with an overall identity of about 35%. Detailed structural analysis indicated some unique features in (1) the cysteine-switch region of the prodomain, (2) the hinge region preceding the hemopexin domain, and (3) the hemopexin domain. Using a bacterial system, HMMP protein was expressed and folded to obtain an active enzyme. Substrate analysis studies indicated that recombinant HMMP could digest a number of hydra ECM components such as hydra laminin. Using a fluorogenic MMP substrate assay, it was determined that HMMP was inhibited by peptidyl hydroxamate MMP inhibitors, GM6001 and matlistatin, and by human recombinant TIMP-1. Whole-mount in situ studies indicated that HMMP mRNA was expressed in the endoderm along the entire longitudinal axis of hydra, but at relatively high levels at regions where cell-transdifferentiation occurred (apical and basal poles). Functional studies using GM6001 and TIMP-1 indicated that these MMP inhibitors could reversibly block foot regeneration. Blockage of foot regeneration was also observed using antisense thio-oligo nucleotides to HMMP introduced into the endoderm of the basal pole using a localized electroporation technique. Studies with adult intact hydra found that GM6001 could also cause the reversible de-differentiation or inhibition of transdifferentiation of basal disk cells of the foot process. Basal disk cells are adjacent to those endoderm cells of the foot process that express high levels of HMMP mRNA. In summary, these studies indicate that hydra has at least one MMP that is functionally tied to morphogenesis and cell transdifferentiation in this simple metazoan.     

The Basal Inhibition in Hydra may be Mediated by a Diffusing Substance

The decay, following basal disk removal, of the disk-mediatedinhibition of disk formation in hydra is as slow as would beexpected, were the inhibition mediated by a diffusing substance. When a hydra's size is changed, either by adding or by removingmass, the inhibition intensity changes in the manner expectedif the vehicle of the inhibition is a substance diffusing fromthe disk and destroyed by all the cells of the body.     

Cloning of noggin gene from hydra and analysis of its functional conservation using Xenopus laevis embryos

SUMMARY Hydra, a member of phylum Cnidaria that arose early in evolution, is endowed with a defined axis, organized nervous system, and active behavior. It is a powerful model system for the elucidation of evolution of developmental mechanisms in animals. Here, we describe the identification and cloning of noggin‐like gene from hydra. Noggin is a secreted protein involved at multiple stages of vertebrate embryonic development including neural induction and is known to exert its effects by inhibiting the bone morphogenetic protein (BMP)‐signaling pathway. Sequence analysis revealed that hydra Noggin shows considerable similarity with its orthologs at the amino acid level. When microinjected in the early Xenopus embryos, hydra noggin mRNA induced a secondary axis in 100% of the injected embryos, demonstrating functional conservation of hydra noggin in vertebrates. This was further confirmed by the partial rescue of Xenopus embryos by hydra noggin mRNA from UV‐induced ventralization. By using animal cap assay in Xenopus embryos, we demonstrate that these effects of hydra noggin in Xenopus embryos are because of inhibition of BMP signaling by Noggin. Our data indicate that BMP/Noggin antagonism predates the bilaterian divergence and is conserved during the evolution.     

Models for the generation of the embryonic body axes: ontogenetic and evolutionary aspects

Coelenterates including hydra are assumed to be close to the last common ancestor before bilaterality evolved. Models based on local self-enhancement and long-range inhibition account for pattern formation and regeneration along this ancestral axis. The body of a hydra-like ancestor evolved into the brain and heart of higher organisms, accounting for the close relationship of both patterning processes. Bilateria require a long-extended organizing region to pattern their dorsoventral axis. Models reveal the difficulties in the generation of such a stripe-like organizer and account for different mechanisms realized in vertebrates and insects. Common pathways involved in hydra budding and in the formation of appendages in higher organisms suggest a possible link.     

Evidence that polycystins are involved in <Emphasis Type="Italic">Hydra</Emphasis> cnidocyte discharge

Like other cnidarians, the freshwater organism Hydra is characterized by the possession of cnidocytes (stinging cells). Most cnidocytes are located on hydra tentacles, where they are organized along with sensory cells and ganglion cells into battery complexes. The function of the battery complexes is to integrate multiple types of stimuli for the regulation of cnidocyte discharge. The molecular mechanisms controlling the discharge of cnidocytes are not yet fully understood, but it is known that discharge depends on extracellular Ca²⁺ and that mechanically induced cnidocyte discharge can be enhanced by the presence of prey extracts and other chemicals. Experiments in this paper show that a PKD2 (polycystin 2) transient receptor potential (TRP) channel is expressed in hydra tentacles and bases. PKD2 (TRPP) channels belong to the TRP channel superfamily and are non-selective Ca²⁺ channels involved in the transduction of both mechanical and chemical stimuli in other organisms. Non-specific PKD2 channel inhibitors Neo (neomycin) and Gd³⁺ (gadolinium) inhibit both prey capture and cnidocyte discharge in hydra. The PKD2 activator Trip (triptolide) enhances cnidocyte discharge in both starved and satiated hydra and reduces the inhibition of cnidocyte discharge caused by Neo. PKD1 and 2 proteins are known to act together to transduce mechanical and chemical stimuli; in situ hybridization experiments show that a PKD1 gene is expressed in hydra tentacles and bases, suggesting that polycystins play a direct or indirect role in cnidocyte discharge.     

Biosynthetic events of hydrid regeneration

Summary The results of a combined morphological and biochemical study of the role of DNA synthesis during distal regeneration inHydra oligactis revealed that a burst of³H-thymidine incorporation into DNA preceded the elaboration of each of the initial three tentacles. In addition, the relative level of each burst of precursor incorporation relfected the number of tentacles formed at that time. Cytological localization of concentrated amounts of labeled material in nuclei of the hypostome and tentacle regions provided corroborative evidence for the biochemical findings.Evidence that the increased DNA specific activity levels described above are associated with tentacle initiation derived from studies in which regenerating hydra were cultured in hydroxyurea and studies in which hydra regenerated proximally rather than distally. Hydra regenerating in 8 mg/ml (0.105 M) hydroxyurea developed morphologically recognizable hypostomes but no tentacles, and incorporated³H-thymidine into DNA at a level distinctly below that exhibited by uncut, untreated animals. Similarly, hydra regenerated a normal, functional basal disc in the absence of any increased DNA specific activity. Therefore, it is suggested that tentacle initiation inH. oligactis requires concomitant DNA synthesis and, as such, represents an epimorphic phenomenon.     

Notch-signalling is required for head regeneration and tentacle patterning in Hydra

Local self-activation and long ranging inhibition provide a mechanism for setting up organising regions as signalling centres for the development of structures in the surrounding tissue. The adult hydra hypostome functions as head organiser. After hydra head removal it is newly formed and complete heads can be regenerated. The molecular components of this organising region involve Wnt-signalling and β-catenin. However, it is not known how correct patterning of hypostome and tentacles are achieved in the hydra head and whether other signals in addition to HyWnt3 are needed for re-establishing the new organiser after head removal. Here we show that Notch-signalling is required for re-establishing the organiser during regeneration and that this is due to its role in restricting tentacle activation. Blocking Notch-signalling leads to the formation of irregular head structures characterised by excess tentacle tissue and aberrant expression of genes that mark the tentacle boundaries. This indicates a role for Notch-signalling in defining the tentacle pattern in the hydra head. Moreover, lateral inhibition by HvNotch and its target HyHes are required for head regeneration and without this the formation of the β-catenin/Wnt dependent head organiser is impaired. Work on prebilaterian model organisms has shown that the Wnt-pathway is important for setting up signalling centres for axial patterning in early multicellular animals. Our data suggest that the integration of Wnt-signalling with Notch-Delta activity was also involved in the evolution of defined body plans in animals.     

水螅的替代食物筛选

在水螅的培养中,常出现食物缺乏。为了获得替代食物,比较水螅对一些常见肉类的捕食行为。结果表明,水螅对蚯蚓肉、虾肉、鱼肉、鸡肉、牛肉和猪肉均能捕捉,对各食物的捕捉比例无显著差异;但水螅对食物的摄食比例在食物类型间存在显著差异,对虾肉较为偏好。可选择虾肉作为培养时的替代食物。     

Chemical anatomy of hydra nervous system using antibodies against hydra neuropeptides: a review

Polyclonal antibodies against hydra neuropeptides allow us to visualize the hydra nerve net. Chemical anatomy of the hydra nerve net was archived by means of immunocytochemistry with various antibodies to hydra neuropeptides. Our goal is to describe the hydra nerve net both qualitatively and quantitatively. The present chemical anatomy results indicate (1) differences in peptide expression between ganglion cells and sensory cells, (2) large differences in neuropeptide expression between ectodermal nerve cells and endodermal nerve cells, and (3) the usefulness of quantitative chemical anatomy to analyze the entire nervous system of hydra.     

Hydra of Hawaii: phylogenetic relationships with continental species

The presence of freshwater hydra in the Hawaiian Islands, over 3,700 km from the nearest continental margin, provokes the question of how these animals could reach the islands. We examined three islands for hydra and found two species (the brown hydra, Hydra vulgaris Pallas 1766, and green hydra, H. viridissima Pallas 1766) present at multiple locations on Oahu and Kauai, and at a single site on Hawaii. Phylogenetic analysis based on the internal transcribed spacer (ITS1 and ITS2) regions of ribosomal DNA indicates that all collected strains of brown hydra were closely related to each other—consistent with a single introduced origin. The analysis also shows that all sampled Hawaiian brown hydra fall within a specific clade of H. vulgaris. This clade is sister to a North American clade and nested within a deeper North/Central America clade. The clade with all the Hawaiian brown hydra includes also individuals from Southern California, suggesting a Californian origin for the brown Hawaiian hydra. Hawaiian hydra were probably transported to the islands by man.     

光周期对中国水螅种群增长、无性出芽生殖及抗氧化酶活力的影响

单细胞真核绿藻在中国水螅(Hydra sinensis)内胚层皮肌细胞中共生是有较高科研价值的特殊生物学现象。水螅宿主细胞为共生藻提供CO2、氮源及矿物质,而共生藻通过光合作用可能为宿主提供碳水化合物等有机物营养,因此水螅与共生藻间代谢流是以共生藻光合作用为中心,但基于代谢流二者间的互作机制目前尚未阐明。水螅通过营养积累进行出芽生殖,从母体脱落的芽体数量间接反映水螅营养积累的相对量。而光暴露时长能影响共生绿藻光合作用,如果共生藻的确能向水螅细胞转移光合作用产物,那光暴露时长应该能间接影响水螅的营养积累、从而进一步影响水螅无性出芽生殖。为证实该假说,本研究应用种群累积培养法,观察了光周期对中国水螅种群增长、无性出芽生殖及抗氧化酶(SOD和CAT)活力的影响。结果显示,光周期对中国水螅种群增长具有明显的影响。培养15 d后,所有实验组水螅的种群密度均为正增长,其中8L∶16D(在一个24h周期内光暴露8 h、黑暗16 h)实验组种群密度最大、而0L∶24D(持续黑暗)实验组种群密度最小。另外,随着光暴露时长的增加,中国水螅SOD及CAT活力整体均呈下降趋势。结果表明,从光周期对中国水螅无性出芽生殖及两种抗氧化酶活力的影响来看,中国水螅对光周期的生理学响应较为敏感,这个现象可能源于共生绿藻能通过向宿主细胞转移光合作用产物的方式为水螅提供营养补充。