REGENERATION AND POST-METAMORPHIC DEVELOPMENT OF THE CENTRAL-NERVOUS-SYSTEM IN THE PROTOCHORDATE CIONA-INTESTINALIS - A STUDY WITH MONOCLONAL-ANTIBODIES

In this study, we use three monoclonal antibodies that recognise antig ens present in the central nervous system of the ascidian Ciona intest inalis to study regeneration and post-metamorphic development of the n eural ganglion. We have also used bromodeoxyuridine labelling to study generation of the neuronal precursor cells. The first antibody, CiN 1 , recognises all neurones in the ganglion, whereas the second, CiN 2, recognises only a subpopulation of the large cortical neurones. Wester n blotting studies show that CiN 2 recognises two membrane-bound glyco proteins of apparent Mr 129 and 100 kDa. CiN 1 is not reactive on West ern blots. Immunocytochemical studies with these antibodies show that CiN 1-immunoreactive neurone-like cells are present at the site of reg eneration as early as 5-7 days post-ablation, a sub-population of CiN 2-immunoreactive cells being detected by 9-12 days post-ablation. The third antibody, ECM 1, stains extracellular matrix components and reco gnises two diffuse bands on Western blots of whole-body and ganglion h omogenates. The temporal and spatial pattern of appearance of CiN 1 an d CiN 2 immunoreactivity both during post-metamorphic development and in regeneration occurs in the same sequence in both processes. Studies with bromodeoxyuridine show labelled nuclei in some neurones in the r egenerating ganglion. Plausibly these originate from the dorsal strand , an epithelial tube that reforms by cell proliferation during the ini tial phases of regeneration. A second population of cells, the large c ortical neurones, do not incorporate bromodeoxyuridine and thus must h ave been born prior to the onset of regeneration. This latter finding indicates a mechanism involving trans-differentiation of other cell ty pes or differentiation of long-lived totipotent stem cells.