CLAVA MULTICORNIS (HYDROZOA): UN NUOVO MODELLO SPERIMENTALE DI CNIDARIO.

The organization of the cnidarian nervous system has been widely documented in polyps and medusae, but little is known about the nervous system of planula larvae, which give rise to adult forms after settling and metamorphosis. We described histological and cytological features of the nervous system in planulae of the hydrozoan Clava multicornis. These planulae crawl on the substrate by means of directional, coordinated ciliary movement coupled to lateral muscular bending movements associated with positive phototaxis. Histological analysis shows pronounced anteroposterior regionalization of the planula’s nervous system, with different neural cell types highly concentrated at the anterior pole. Levels of nervous system complexity are uncovered by neuropeptide-specific immunocytochemistry, which reveals distinct neural subsets having specific molecular phenotypes. During metamorphosis, the larva attaches to the substrate with the anterior end that develops into the basal foot region of the polyp, while the posterior end forms the oral region. To elucidate the fate of the neural cells in C. multicornis, we investigated the distribution pattern of GLWamide and RFamide positive sensory cells at different stages of metamorphosis. We observed that immunoreactivity was still present at the anterior end of the larva during early settlement, but gradually disappeared through the following stages of metamorphosis. Only at later stages, positive cells appeared around the mouth of the newly formed polyp. By TUNEL assay, apoptotic nuclei were identified in the anterior end of the settled larva, in the same region occupied by sensory cells. These results suggest that at least part of the neurons of the larva degenerates during metamorphosis by apoptosis and that at least part of the adult nervous system is formed by de novo differentiation. The WNT and FGF signaling pathways are involved in determining the body axis in several animal models. Moreover, the way of FGF is involved in cell differentiation. The lithium chloride acts by activating the path of canonical WNT signal. SU5402 is an antagonist of FGF receptors and inhibits the FGF signaling pathway. To obtain information regarding the presence of these pathways in cnidarians we analyzed the effects of treatment with lithium chloride and SU5402 on the embryonic development of Clava multicornis. The results obtained showed that the treated animals maintained a length comparable to the control specimens, while changing the shape of the two poles of the animals. In fact, the larvae treated with lithium chloride assume a rod-shaped instead of the typical club-shaped, while the larvae treated with SU5402 are wedge-shaped. Later in the treated animals have been seen cells containing GLWamide and RFamide peptides by immunolocalization. There are differences in the position of cells and their number in the larvae treated with lithium chloride than in those treated with SU5402. This shows that the two substances interfere with the proper development of the animal. The effects of these drugs have had on Clava multicornis suggest the likely presence of the signaling pathway of WNT and FGF in these animals. Retinoic acid (RA) is a well-known gradient morphogen governing the spatial patterning of anteroposterior elements in vertebrates and in other chordates. Components of RA machinery, such as synthetic and catalytic enzymes and receptors, have been isolated in a number of organisms, from nematods to echinoderms, suggesting that the morphogenetic role of this molecule is not exclusive to chordates.. We tested the effects of RA and of a RA antagonist on C. multicornis development by analyzing the nervous system organization and the photoresponsive behavior of larvae exposed to RA during their embryogenesis. After the exposure, the nervous system became completely disorganized, leading to the displacement of RFamide-IR cells. Moreover, RA treated larvae did not exhibit the typical phototropic behavior of control specimens. Our results suggest that RA can alter the normal development of nervous elements in this hydroid species, supporting the hypothesis that the morphogenetic activity of RA predates the origin of chordates. The regenerative capacity of cnidarians is remarkable. In this work we analyzed the regenerative processes of gastric columns of Clava multicornis, treating the latter with increasing concentrations of RA in order to evaluate their effects. Concentrations of RA 10-7 M did not have a material impact on the regeneration of amputated portions of the polyp, which regenerates similarly to control animals. The 10-6 M RA primarily involves the regeneration of stolons. Together these observations imply that the nervous system of the planula of Clava multicornis manifests a remarkable level of histological, cytological, and functional organization, the features of which may be reminiscent of those present in early bilaterian animals. In fact, these results open up important perspectives on the evolutionary significance of the antagonism between RA and FGF / WNT in basal metazoans.