N-acetylneuraminic acid (NANA) stimulates in situ cyclic AMP production in tentacles of sea anemone (Aiptasia pallida): possible role in chemosensitization of nematocyst discharge.

Cnidocytes, the stinging cells of cnidarians, optimally discharge nematocysts in response to combined physical contact and stimulation of specific chemoreceptors. In the tentacles of certain sea anemones, the primary chemoreceptors bind N-acetylated sugars, such as N-acetylneuraminic acid (NANA). Sensitization with NANA predisposes contact-sensitive mechanoreceptors (CSMs) to trigger discharge in response to physical contact. In the ectoderm of sea anemone tentacles, cnidocyte/supporting cell complexes (CSCCs) control and trigger nematocyst discharge. Previous findings have implicated cyclic AMP (cAMP) as a second messenger in NANA-sensitized nematocyst discharge. However, no reports have directly demonstrated that the cAMP content of tentacles changes in response to NANA stimulation. We now show that NANA elevates in situ cAMP levels in a dose-dependent manner in the ectoderm of tentacles from the sea anemone Aiptasia pallida. However, the endoderm of tentacles shows no detectable cAMP response to NANA. The effect of NANA on the cAMP content of the ectoderm is biphasic. Micromolar NANA increases the in situ cAMP level, with a maximal response occurring at 1.8x10(-5)mol x l(-1) NANA. At higher NANA concentrations, the cAMP content decreases to that of controls. Because the cAMP dose/response curve to NANA coincides precisely with the dose/response curves of NANA-sensitized nematocyst discharge and nematocyst-mediated adhesive force, a second-messenger role for cAMP in NANA-sensitized nematocyst discharge is strongly suggested. The addition of isobutyl-1-methylxanthine (IBMX) to the medium with sea anemones increases tissue cAMP levels both in the absence and in the presence of NANA. However, anesthetizing anemones in sea water containing high levels of Mg(2+) blocks the NANA-stimulated cAMP response of the ectoderm. In addition, our results suggest that NANA-stimulated cAMP may activate endogenous cAMP-dependent protein kinase (PKA) in broken cell preparations of tentacles. Thus, NANA-stimulated cAMP may function as a second messenger in the NANA chemosensory signaling pathway controlling nematocyst discharge.

Efferent Mechanisms of Discharging Cnidae: II. A Nematocyst Release Response in the Sea Anemone Tentacle.

Feeding behavior in cnidarians is a sequence of coordinated responses beginning with nematocyst discharge. The nematocyst response produces prey capture by envenomating prey and attaching prey to the tentacle. The strength of attachment of discharged nematocysts to the tentacle is termed intrinsic adherence and is calculated from measurements of adhesive force. Following prey capture, the feeding response involves movement of the tentacles toward the mouth and mouth opening. For ingestion to occur, nematocysts attaching the prey to the tentacles must be released from the tentacle. A nematocyst release response has been proposed, but never documented nor measured. Our criterion for a nematocyst release response is that the intrinsic adherence of discharged nematocysts must decrease to zero. The unit of nematocyst discharge in sea anemone tentacles is the cnidocyte/ supporting cell complex (CSCC). The nematocyst response includes nematocysts discharged from Type C CSCCs by physical contact alone and nematocysts discharged from the more numerous Type B CSCCs that require both chemosensitization and physical contact. We identify two prey-derived substances, N-acetylneuraminic acid (NANA) and glycine, both of which chemosensitize nematocyst discharge from Type B CSCCs at low concentrations. At higher concentrations NANA stimulates the release response of Type Cs, and glycine stimulates the release response of Type Bs.

Efferent Mechanisms of Discharging Cnidae: I. Measurements of Intrinsic Adherence of Cnidae Discharged From Tentacles of the Sea Anemone, Aiptasia pallida.

Two kinds of cnida predominate in the tentacles of the acontiate sea anemones: spirocysts and microbasic mastigophore nematocysts. These cnidae discharge in response to appropriate mechanical and chemical stimulation. In this paper, we calculate the strengths of attachment between the tentacle and the capsules (= "tentacle adherence") of discharged spirocysts and mastigophores by measuring adhesive force and by determining the numbers of spirocysts and mastigophores discharged onto targets under conditions where the attachment of everted cnida tubules to the target (= "cnida adhesion") exceeds tentacle adherence. Under these conditions, the average contribution of individual cnidae to adhesive force is called the intrinsic adherence. The intrinsic adherence is a measure of the average frictional force required to dislodge the capsule of individual discharged cnidae from the tentacle. The intrinsic adherence of discharged mastigophores varies inversely, from 0.45 to 0.15 mgf (4.41 to 1.47 µN), with the number of discharged mastigophores. The larger values characterize mastigophores discharged by mechanically triggering nonchemosensitized tentacles, whereas the lower values characterize the intrinsic adherence of mastigophores discharged from chemosensitized tentacles. In contrast, the intrinsic adherence of discharged spirocysts is very low to insignificant. Thus, by comparison to mastigophores, spirocysts contribute little, if any, to adhesive force, and, by inference, do not directly secure captured prey to the tentacle. Our measurements indicate that penetrable prey are primarily secured to the tentacle by discharged mastigophores and by the inherent stickiness of the tentacle surface.

Control of Cnida Discharge: III. Spirocysts are Regulated by Three Classes of Chemoreceptors.

Spirocysts are two to three times more abundant than nematocysts in the feeding tentacles of acontiate sea anemones. Despite their prevalence, little experimental work has been done on the discharge of spirocysts because of the difficulty in detecting and counting them after they have discharged. To circumvent this problem, we have developed a simple, reliable, enzyme-linked lectin sorbent assay (ELLSA) for quantifying discharged spirocysts. With this method, we have shown that the discharge of spirocysts, like that of mastigophore nematocysts, is chemosensitized in a dose-dependent manner by three classes of low molecular weight substances, typified by N-acetylneuraminic acid (NANA), glycine, and certain heterocyclic amino compounds, such as proline and histamine. We also show that spirocysts exhibit considerable agonist-specific variation in the dose-responses of discharge, suggesting the existence of multiple populations of spirocyst-bearing cnidocyte/supporting cell complexes (CSCCs). Our findings call into question commonly held views regarding the respective roles of spirocysts and mastigophore nematocysts in the retention of captured prey.