水螅畸形触手转分化过程的观察

水螅胃区诱导出的触手芽，１４天后在茎区诱导体柱组织分化头结构。胃区诱导出的畸形触手芽，８天后在出芽区脱落，在母体上留下一个基盘，脱落的畸形触手呈２触手水螅个体，其直径相当于头部触手直径的２／３。在人工帮助下能吞食１个未分化的蚤状卵。第１２天能自行捕食幼。最后对２种额外触手芽的不同演化结果做了简单比较。     

水螅营养积累对基盘组织更新进程的影响

目的观察大乳头水螅（Hydra magnipapillata ）基盘组织更新进程,探讨水螅营养积累对基盘组织更新进程的影响。方法设定水螅喂食频率梯度（代表不同的营养积累水平）,记录和观察喂食频率对水螅更新基盘组织进程的影响。通过ABTS细胞化学染色法检测水螅基盘分子标志物过氧化物酶的表达,观察水螅老基盘组织脱落后水螅体主体新生基盘组织的再生过程。结果喂食频率对水螅更新基盘组织进程有明显的影响。水螅基盘组织更新的标准过程如下;在一定的喂食频率下培养水螅,水螅体出芽区逐渐有芽体产生,随后在出芽区和基盘之间靠近芽体的位置出现缢痕,最后水螅体在缢痕处断裂为水螅体主体和老基盘组织两部分。缢痕断裂后对水螅体主体保持既定的喂食频率,其伤口能愈合但不能再生出新的基盘组织;对其降低喂食频率直至其伤口上方的芽体全部脱落后伤口处重新启动新生基盘组织的再生进程。另外,脱落的老基盘组织有两种不同的命运,即大部分老基盘组织不能发育成正常水螅体、最终解体;而小部分的老基盘组织能发育成正常的水螅体。结论水螅营养积累可能促进基盘组织更新进程,靠近断裂伤口处的芽体能抑制水螅体主体新生基盘的再生进程。     

水螅芽体发育与头部再生进程间的相互作用

目的观察具有单个芽体的大乳头水螅(Hydra magnipapillata)头部再生进程,探讨水螅头部结构的再生进程与芽体发育过程之间可能存在的相互作用。方法选取具有单个年幼芽体的水螅体,在水螅母体上紧贴芽体着生部位的上方进行切除手术,观察母体头部再生进程。通过ABTS细胞化学染色法检测水螅基盘分子标志物过氧化物酶的表达,观察水螅芽体基盘与母体间的结构联系。结果水螅母体伤口在手术后2h内愈合。随着再生时间的延长,出现两种不同命运的芽体发育方式。一种情况是水螅芽体基盘紧贴母体手术切口,芽体发育成熟后可正常脱离母体;在芽体脱离母体前母体头部再生进程被抑制,在芽体脱落后母体头部再生进程重新启动、且在其后48h内母体头部再生完成。另一种情况是水螅芽体基盘组织与母体手术切口不产生结构联系而向外突出生长,母体头部再生进程完全停止,芽体胃区与母体相连且芽体发育成熟后不脱离母体。结论靠近水螅母体手术伤口的年幼芽体能延迟或阻断母体头部的再生进程,而手术切口也可能干扰了发育成熟的芽体与母体脱离的正常机制。     

水螅触手对垂唇的形态发生的影响

本文采用移植触手于水螅胃区,同时切除水螅头的方法,仔细胞了４２个水螅的切面和胃哎的垂唇的形发生过程。其中发现了３例有触手无口的特殊畸形水螅。     

水螅触手的诱导作用

从3个不同的面切割水螅,观察愈合时及愈合后各触手的变化情况。第1组中有少数原位于触手环上的触手外移出触手环,移至茎区诱导成头,但如果切去已移出触手环外的触手,则无头形成。第2组中多数基部带有组织块的触手环外的触手移至茎区诱导成头,但如果切去触手环外的触手,则只有还保留有由组织块形成的圆锥状基部的可形成头;而切去圆锥状基部的则无头形成。第3组中再生性的位于触手环外的触手可在原水螅头的对应末端诱导成头。以上3个方面的观察结果都说明了水螅触手在一定条件下具有诱导成头的能力。     

水螅的无性生殖

淡水水螅的无性繁殖方式是出芽生殖。水螅胃区的皮肌细胞摄取的营养物质,经细胞间传递方式,转移到芽体的内、外胚层,为芽体的发育提供了能量。胃区的干细胞与增殖的细胞不断迁移到芽体,保障芽体发生时的细胞数。水螅的头部与基盘对芽体的发生,同时存在着激活与抑制2对位置信息素,共同控制并决定了芽体在体柱上的发生位置。芽体先发生垂唇与触手芽,其发生位置必需远离母体头部抑制素的作用。芽体发育后期,发生基盘时,必须远离母体基盘抑制素的作用。     

骨骼肌细胞的凋亡

细胞凋亡是受基因调控的细胞死亡方式,与机体组织的萎缩和退化关系密切,近年来有人将骨骼肌的萎缩和退化与细胞凋亡联系起来,认为细胞凋亡可能在骨骼肌萎缩和退化中发挥重要作用,本文综述了近年来对骨骼肌肌肉细胞和未分化肌细胞的凋亡及其基因调控的研究,以期进一步阐明骨骼肌萎缩和退化的发生机制,为临床上寻找延缓骨骼肌萎缩的治疗方法提供一些思路。     

水螅在异物穿插状态下的组织动态

本文于１９９３年用３号昆虫针纵穿水螅肠腔、切除水螅头足纵穿腔肠及横穿水螅体柱等方法，实验了２４０个水螅、观察表明，所有被针穿的个体都能在１—１５天内脱逃。纵穿腔肠的个体以纵裂体壁的方式脱逃，横穿身体的个体是分别把插针向口端或体侧或基部推移出体外。切除头、足后纵穿腔肠的个体，体柱一侧组织收缩，收缩部按原极性分化再生新的头和足，相对一侧体壁则发生纵裂并脱逃。最后作了简单讨论。     

双头无基盘水螅的初步观察

自然形成的双头无基盘畸形体水螅极为罕见,为探索其形成原因,人工嫁接水螅14例,获得5例相同的双头水螅。对其分别进行连续观察发现:嫁接的双头水螅的双头可同时摄食,食物集聚于1个胃区。随个体生长其体柱延长,逐渐发育为2个胃区,在2个胃区间发生2~6个芽体,所有子代发育正常。出芽位置至触手环的距离同正常水螅。之后分化为2个出芽区,出芽区之间体柱较透明,同茎区组织。第21天开始发生基盘,发生过程异常缓慢。     

大乳头水螅基盘再生进程中过氧化物酶的表达

目的观察大乳头水螅(Hydra magnipapillata)基盘再生进程中基盘过氧化物酶的表达情况,探讨水螅基盘过氧化物酶的生理作用。方法通过ABTS细胞化学染色法显示水螅基盘过氧化物酶的表达。结果水螅基盘再生20h后其基盘过氧化物酶开始出现少量表达,其后过氧化物酶表达量逐渐增加;基盘再生52h后该酶表达量趋于稳定。过氧化物酶仅在基盘周边区域外胚层中表达,而在基盘中央区域(反口孔)外胚层中无表达。结论水螅基盘再生进程中过氧化物酶的表达量逐渐增加直接反映了基盘再生时细胞分化过程,基盘表达的过氧化物酶可能在维持基盘结构的稳定上起一定的作用。     

Formation of pattern in regenerating tissue pieces of Hydra attenuata: II. Degree of proportion regulation is less in the hypostome and tentacle zone than in the tentacles and basal disc

The relative sizes of the various structures in Hydra attenuata were compared over a broad range of animal sizes to determine in detail the ability to regulate proportions during regeneration. The three components of the head, namely hypostome, tentacles, and tentacle zone from which the tentacles emerge, the body column, and the basal disc were all measured separately. Ectodermal cell number was used as the measure of size. The results showed that the basal disc proportioned exactly over a 40-fold size range, and the tentacle tissue proportioned exactly over a 20-fold size range. In contrast, the hypostome and tentacle zone proportioned allometrically. With decreasing size, the hypostome and tentacle zone became an increasing fraction of the animal at the expense of body tissue, and in the very smallest regenerates at the expense of tentacle tissue. In their current form, the reaction-diffusion models proposed for pattern regulation in hydra are not consistent with the data.     

Formation of pattern in regenerating tissue pieces of Hydra attenuata: I. Head-body proportion regulation

The precision with which an almost uniform sheet of hydra cells develops into a complete animal was measured quantitatively. Pieces of tissue of varying dimensions were cut from the body column of an adult hydra and allowed to regenerate. The regenerated animals were assayed for number of heads (hypostomes plus tentacle rings), head attempts (body tentacles), and basal discs. To ascertain whether the head and body were reformed in normal proportions, the average number of epithelial cells in the heads and bodies was measured. Pieces of tissue, from $\text{1}\text{2}$ to $\text{1}\text{20}$ an adult in size, formed heads that were a constant fraction of the regenerate. Thus, over a 10-fold size range, a proportioning mechanism was operating to divide the tissue into head area and body area quite precisely, but appeared to reach limits at the extremes of the range. However, the regenerates were not all normal miniatures with one hypostome and one basal disc. As the width-length ratio of the cut piece was increased beyond the circumference-length ratio of the intact body column, the incidence of extra hypostomes in the “head” and body tentacles and extra basal discs in the “body” rose dramatically. A proportioning mechanism based on the Gierer-Meinhardt model for pattern formation is presented to explain the results.     

Expression of a novel receptor tyrosine kinase gene and a paired-like homeobox gene provides evidence of differences in patterning at the oral and aboral ends of hydra

Axial patterning of the aboral end of the hydra body column was examined using expression data from two genes. One, shin guard, is a novel receptor protein-tyrosine kinase gene expressed in the ectoderm of the peduncle, the end of the body column adjacent to the basal disk. The other gene, manacle, is a paired-like homeobox gene expressed in differentiating basal disk ectoderm. During regeneration of the aboral end, expression of manacle precedes that of shin guard. This result is consistent with a requirement for induction of peduncle tissue by basal disk tissue. Our data contrast with data on regeneration of the oral end. During oral end regeneration, markers for tissue of the tentacles, which lie below the extreme oral end (the hypostome), are detected first. Later, markers for the hypostome itself appear at the regenerating tip, with tentacle markers displaced to the region below. Additional evidence that tissue can form basal disk without passing through a stage as peduncle tissue comes from LiCl-induced formation of patches of ectopic basal disk tissue. While manacle is ectopically expressed during formation of basal disk patches, shin guard is not. The genes examined also provide new information on development of the aboral end in buds. Although adult hydra are radially symmetrical, expression of both genes in the bud's aboral end is initially asymmetrical, appearing first on the side of the bud closest to the parent's basal disk. The asymmetry can be explained by differences in positional information in the body column tissue that evaginates to form a bud. As predicted by this hypothesis, grafts reversing the orientation of evaginating body column tissue also reverse the orientation of asymmetrical gene expression.     

Proportioning a Hydra

SYNOPSIS. Pieces of hydra tissue of various sizes and shapeswere cut from the body columns of adult hydra and allowed toregenerate. The proportions of the resulting animals were determinedfirst by counting cells in the head and body, and secondly bymeasuring the structures directly using an ocular micrometer. Head-body proportions were found to be constant over a tenfoldsize range. Very small regenerates had a larger head fractionand large budding regenerates had a smaller head fraction. Extrastructures developed in certain shape pieces, but total head-bodytissue remained proportional. More detailed measurement of thehead showed that the hypostome regulated only slightly withtotal size change, while the tentacle tissue varied considerablyto maintain the head-body ratio. This suggested that the patterningof the hypostome and the tentacles might involve separate processes,with the latter being responsible for proportion regulation.While the body mass appeared to be determined by the proportioningmechanism, its circumference was related to the circumferenceof the hypostome, suggesting a causal relationship between thetwo organizers and the column shaping. The basal disc remainedproportional to the body except in the smallest pieces. A Gierer-Meinhardtpattern formation scheme could account for the results found.     

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.     

Hym-301, a novel peptide, regulates the number of tentacles formed in hydra

Hym-301 is a peptide that was discovered as part of a project aimed at isolating novel peptides from hydra. We have isolated and characterized the gene Hym-301, which encodes this peptide. In an adult, the gene is expressed in the ectoderm of the tentacle zone and hypostome, but not in the tentacles. It is also expressed in the developing head during bud formation and head regeneration. Treatment of regenerating heads with the peptide resulted in an increase in the number of tentacles formed, while treatment with Hym-301 dsRNA resulted in a reduction of tentacles formed as the head developed during bud formation or head regeneration. The expression patterns plus these manipulations indicate the gene has a role in tentacle formation. Furthermore, treatment of epithelial animals indicates the gene directly affects the epithelial cells that form the tentacles. Raising the head activation gradient, a morphogenetic gradient that controls axial patterning in hydra, throughout the body column results in extending the range of Hym-301 expression down the body column. This indicates the range of expression of the gene appears to be controlled by this gradient. Thus, Hym-301 is involved in axial patterning in hydra, and specifically in the regulation of the number of tentacles formed.     

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.     

Perturbations in morphogen gradients induce budding in hydra.

Lateral grafting of small pieces of midregion tissue into different levels of the hydra body column was done to assess the influence of the host hypostome and basal disc (or, of the underlying morphogenetic gradients) in inducing secondary structures in the transplanted tissue; and also to identify the role, if any, of the induced secondary structures (or, perturbed morphogen gradients) on the pattern of the host. The same midpiece tissue differentiated to a basal disc when grafted near the host hypostome, and to a small hypostome with tentacles when grafted near the host basal disc. Chimeras with induced secondary basal discs showed a phenomenal increase in budding compared to the controls and to the chimeras having induced hypostomes. These results indicate a positive cross-reaction between both organizing regions during patterning in hydra.     

Feeding and wounding responses in Hydra suggest functional and structural polarization of the tentacle nervous system

The nervous system of Hydra, a freshwater cnidaria, occurs as dispersed, or diffuse, nerve net throughout the animal. It is widely accepted that in a diffuse nervous system an external stimulus is conducted in all directions over the net. Here I report observations that hydra tentacles respond to feeding and wounding stimuli in a unidirectional manner. Upon contact of a tentacle with a brine shrimp larva during feeding, tissue on the proximal side of the point of contact contracted strongly, whereas tissue on the distal side contracted only very weakly. Feeding a tentacle to which a second tentacle was grafted to the proximal end in the reversed orientation showed that unidirectional conduction, once initiated, was blocked by the reversal of polarity, demonstrating that the distal to proximal polarity of tissue is crucial for unidirectional conduction. Unidirectional conduction was obtained also by mechanically pinching the tissue. The response of tentacles devoid of neurons examined was bidirectional, demonstrating that the nervous system is responsible for the unidirectional responses. These observations suggest that polarized property of the nerve net in hydra tentacles is responsible for the unidirectional tentacle contraction.     

Cngsc, a homologue of goosecoid, participates in the patterning of the head, and is expressed in the organizer region of Hydra.

We have isolated Cngsc, a hydra homologue of goosecoid gene. The homeodomain of Cngsc is identical to the vertebrate (65-72%) and Drosophila (70%) orthologues. When injected into the ventral side of an early Xenopus embryo, Cngsc induces a partial secondary axis. During head formation, Cngsc expression appears prior to, and directly above, the zone where the tentacles will emerge, but is not observed nearby when the single apical tentacle is formed. This observation indicates that the expression of the gene is not necessary for the formation of a tentacle per se. Rather, it may be involved in defining the border between the hypostome and the tentacle zone. When Cngsc(+) tip of an early bud is grafted into the body column, it induces a secondary axis, while the adjacent Cngsc(-) region has much weaker inductive capacities. Thus, Cngsc is expressed in a tissue that acts as an organizer. Cngsc is also expressed in the sensory neurons of the tip of the hypostome and in the epithelial endodermal cells of the upper part of the body column. The plausible roles of Cngsc in organizer function, head formation and anterior neuron differentiation are similar to roles goosecoid plays in vertebrates and Drosophila. It suggests widespread evolutionary conservation of the function of the gene.