Glen M. Watson

THE DEVELOPMENT OF A SEA ANEMONE TENTACLE SPECIALIZED FOR AGGRESSION: MORPHOGENESIS AND REGRESSION OF THE CATCH TENTACLE OF HALIPLANELLA LUCIAE (CNIDARIA, ANTHOZOA)

Three intermediate catch tentacle morphs were observed in the sea anemone Haliplanella luciae during catch tentacle development. Stage 1 catch tentacles, characterized by swollen bulb-like regions along their length, were histologically similar to feeding tentacles. Stage 2 catch tentacles, which tapered normally along most of their length and then constricted near the tip, were characterized by the presence of feeding tentacle cnidae in the tentacle coelenteron as they were removed from developing catch tentacles. Numerous cnidoblasts appeared in stage 2 tentacles and then synchro nously matured into small holotrich nematocysts, a cnida characteristic of mature catch tentacles. Stage 3 catch tentacles were characterized by the appearance of many large holotrich nematocysts. Such tentacles appeared similar to mature catch tentacles with wide, opaque, blunt tips. However, stage 3 catch tentacles had fewer large holotrichs per total tentacle cross section than mature catch tentacles. The numbers of large and small holotrich nematocysts decreased in regressing catch tentacles, which tapered to opaque, pointed tips. However, these cnidae did not move to the coelenteron as before but instead migrated to the epithelial surface. This migration suggested that they were externally expelled from the tentacles.

Comparative ultrastructure of catch tentacles and feeding tentacles in the sea anemone Haliplanella.

TEM observations of catch tentacles revealed that the tentacle tip epidermis is filled with two size classes of mature holotrich nematocysts and a gland cell filled with electron-dense vesicles. Vesicle production is restricted to upper-middle and tentacle tip regions, whereas holotrich development occurs in the lower-middle and tentacle base regions. Thus, catch tentacles have a maturity gradient along their length, with mature tissues concentrated at the tentacle tip. Occasional feeding tentacle cnidae (microbasic p-mastigophores and basitrichs) and mucus gland cells occur in proximal portions of catch tentacles, but are phagocytized by amoeboid granulocytes and transported to the gastrodermis for further degradation. No feeding tentacle cnidae or mucus cells occur distally in catch tentacles. Unlike catch tentacles, feeding tentacles are homogeneous in structure along their length with enidocytes containing mature spirocysts, microbasic p-mastigophore or basitrich nematocysts distributed along the epithelial surface. Cnidoblasts are recessed beneath cnidocytes, occurring along the nerve plexus. Mucus gland cells and gland cells filled with electron-dense vesicles are present in feeding tentacles, distributed at the epithelial surface. Granular phagocytes are rare in the feeding tentacle tip, but common in the tentacle base.

Ultrastructure of nematocyst discharge in catch tentacles of the sea anemone Haliplanella luciae (cnidaria: anthozoa)

The mature nematocyst lies just beneath the cnidodyte plasma membrane. A microtubule array surrounds the nematocyst capsule just beneath the capsule tip. We propose that the array helps to hold the capsule at the cnidocyte cell surface until discharge. The undischarged capsule tip is sealed by three apical flaps, joined together along complex radial seams. The seams are filled with subunits that appear to bind the flaps together. Upon discharge, the flaps separate along the radial seams to permit thread eversion. The everted thread is lined on both sides by subunits that are stained by antimonate, indicating that they bind calcium. We suggest that, together, the subunits hold the uneverted thread in its folded and coiled configuration. Thread eversion would follow subunit uncoupling. The capsule and thread interiors of partially discharged nematocysts are stained by antimonate. In contrast, the capsule and thread interiors of fully discharged nematocysts are not stained by antimonate. Thus, nematocyst calcium might be injected into the target tissue where it is presumed to act in conjunction with nematocyst venom to promote cell death.

The Cell Biology of Nematocysts

Publisher Summary This chapter focuses on the cell biology of nematocysts. Nematocysts, the most widely used term to describe the “stinging capsules” characteristic of the phylum Cnidaria, constitute the best-studied and most diverse group of cnidae, the secretory products of cnidocytes. All cnidae consist of a collagenous capsule containing an eversible tubule. The true diversity of cnida types may be higher due to the presence of distinct size classes of a single nematocyst type in different tissues of the animal. All cnidae discharge by everting the tubules to contact the target. Tubule eversion can be extremely rapid or fairly slow (seconds), depending on the cnida type. The tubules may be specialized for penetrating the target to permit injection of potent toxins into the target tissue, or for adhering to the surface of the target. Certain cells of the cnidarians exert a complex and sophisticated control over the utilization of nematocysts. In the end, the success of cnidarians as preditors can be attributed largely to the ingenuity of nematocysts and to the many forms of cellular control over their use.

Ultrastructure and sulfur cytochemistry of nematocyst development in catch tentacles of the sea anemone Haliplanella luciae (cnidaria: Anthozoa)

Nematocysts in catch tentacles of the sea anemone Haliplanella luciae are formed by a Golgi-microtubule complex in which the microtubules appear to support the developing nematocyst during its formation. The newly formed capsule becomes wrinkled with the discontinuation of microtubule support, but becomes straight again as the nematocyst wall thickens (possibly from the addition of new material onto the wall surface). Final wall maturation is characterized by a decrease in wall thickness by a factor of two. Wall thinning is apparently caused by disulfide bond formation among wall molecules, because it can be reversed in the mature nematocyst by performic acid oxidation. The mature thread wall stains with Alcian blue both prior to and after performic acid treatment, whereas the mature capsule wall is stained only after such treatment. It is proposed that the thread wall has “free” sulfur groups that are absent in the capsule wall.

Calcium cytochemistry of nematocyst development in catch tentacles of the sea anemone Haliplanella luciae (Cnidaria: Anthozoa) and the molecular basis for tube inversion into the capsule

A Golgi-deposition of calcium into developing holotrich nematocysts in catch tentacles of the sea anemone, Haliplanella luciae , was observed at all stages of nematocyst formation, as evidenced by dense antimonate staining in trans -Golgi cisternae and in the nematocyst interior. Such staining is removed by EGTA treatment. Electron-lucent, laminar granules are visible in the nematocyst capsule and external tube of antimonate-stained preparations (by negative staining), but are invisible in both EGTA-treated nematocysts and conventional TEM nematocyst preparations. These granules appear to solubilize prior to tube inversion into the capsule. Electron-lucent fibrils, 50 by 9 nm, along the outer wall of the inverting tube are in contact with similar fibrils along the inner wall of the external tube and might provide the motive force for tube inversion. Inversion is accompanied by an apparent increase in the nematocyst antimonate concentration, and by an increase in the fluid in the cell, suggesting that a dehydration of the nematocyst contents might occur at the time of tube inversion into the capsule.

Direct Monitoring of Intracellular Calcium Ions in Sea Anemone Tentacles Suggests Regulation of Nematocyst Discharge by Remote, Rare Epidermal Cells

In tentacles of sea anemones, cnidocytes and adjacent supporting cells are believed to be independent receptor-effector complexes that regulate nematocyst discharge in response to exogenous N-acetylated sugars. When sugar chemoreceptors on supporting cells are activated, nematocyst discharge is two- to threefold greater than discharge without chemosensitization. To examine the role of Ca2+ as a second messenger in chemodetection of sugars, we used fluo-3 to monitor Ca2+ levels in epidermal cells of intact anemone tentacles. Certain epidermal cells exhibit relatively high Ca2+ both with and without chemosensitization. With chemosensitization, a two-to threefold increase occurs in the abundance of relatively rare cells exhibiting the highest Ca2+ levels. Timecourses depicting abundances of these rare cells in chemosensitized specimens show positive correlations to timecourses for nematocyst discharge from chemosensitized specimens and for labeling of chemoreceptors. Cnidocyte/supporting-cell complexes discharging nematocysts are about three times more abundant than the rare cells exhibiting the highest intracellular Ca2+ levels. One interpretation of these data is that the Ca2+-dependent regulation of nematocyst discharge occurring with chemosensitization involves intense Ca2+ signaling by remote, rare cells. This interpretation is inconsistent with the current model that portrays cnidocyte/supporting-cell complexes as independent effectors of nematocyst discharge.

Self/Non-Self Recognition Affects Cnida Discharge and Tentacle Contraction in the Sea Anemone Haliplanella luciae

Certain species of sea anemone live in tightly packed communities, among clonemates and non-clonemates. Competition for space leads to intraspecific and interspecific aggressive interactions among anemones. The initial aggressive interactions appear to involve reciprocal discharge of cnidae triggered by contact with non-self feeding tentacles. We asked whether molecules contained in anemone-derived mucus constituted an important cue alone or in combination with cell surface molecules in stimulating aggressive or avoidance behaviors. In this study, we found that self and non-self stimuli differentially influenced two effector systems: cnida discharge and tentacle contraction. Interspecific mucus enhanced nematocyst discharge by 44% and spirocyst discharge by 90%, as compared to baseline discharge obtained in seawater alone. Conspecific stimuli accompanying touch inhibited specific tentacle contractions occurring on the far side of anemones relative to the site of contact. The greatest tentacle contractions occurred with exposure to interspecific mucus and tissue. Thus, several receptor systems are involved that integrate chemical and mechanical cues in order to initiate appropriate and graded effector responses during competition for space.