Identification of nerve plexi in connective tissues of the sea cucumber Holothuria glaberrima by using a novel nerve-specific antibody.

The echinoderm nervous system is one of the least studied among invertebrates, partly because the tools available to study the neurobiology of this phylum are limited. We have now produced a monoclonal antibody (RN1) that labels a nervous system component of the sea cucumber Holothuria glaberrima. Western blots show that our antibody recognizes a major band of 66 kDa and a minor band of 53 kDa. Immunohistological experiments show that, in H. glaberrima, the antibody distinctly labels most of the known nervous system structures and some components that were previously unknown or little studied. A surprising finding was the labeling of nervous plexi within the connective tissue compartments of all organs studied. Double labeling with holothurian neuropeptides and an echinoderm synaptotagmin showed that RN1 labeled most, if not all, of the fibers labeled by these neuronal markers, but also a larger component of cells and fibers. The presence of a distinct connective tissue plexus in holothurians is highly significant since these organisms possess mutable connective tissues that change viscosity under the control of the nervous system. Therefore, the cells and fibers recognized by our monoclonal antibodies may be involved in controlling tensility changes in echinoderm connective tissue.

Spherulocytes in the echinoderm Holothuria glaberrima and their involvement in intestinal regeneration.

The holothuroid echinoderm Holothuria glaberrima can regenerate its intestine after a process of evisceration. Spherule-containing cells, the spherulocytes, appear to be associated with intestinal regeneration. We have used histochemistry and immunocytochemistry to characterize these cells and their role in the regeneration process. Spherulocytes are 10-20 microm in diameter with an acrocentric nucleus and spherule-like structures within their cytoplasm. They are found in the connective tissue of the intestine and mesentery of noneviscerated and regenerating animals. During the second week of regeneration, the number of spherulocytes in the regenerating intestine increases and a dramatic change in their morphology occurs. Together with the morphological change, the immunohistochemical labeling of the cells also changes; the antibodies not only recognize the spherule structures but also label the cellular cytoplasm in a more homogeneous pattern. Moreover, immunohistochemical labeling also appears to be dispersed within the extracellular matrix, suggesting that the cells are liberating their vesicular contents. Spherulocytes are found in other tissues of H. glaberrima, always associated with the connective tissue component. Our data strongly suggest that spherulocytes are involved in intestinal regeneration but their specific role remains undetermined. In summary, our data expand our knowledge of the cellular events associated with regeneration processes in echinoderms and provide for comparisons with similar processes in vertebrates.