This proposal will test the hypothesis that trophic factors can influence whether precursor cells differentiate into neurons or into glia. These factors will be tested in an explant culture system using regenerating spinal cord of the teleost, Apteronotus albifrons. This newly-developed culture system is one of the very few instances in which neurogenesis and initial neuronal differentiation have been demonstrated in vitro. Cultures derived from both normal and regenerating Apteronotus spinal cord show good neurite out-growth and incorporation of 3H-thymidine into cells that differentiate into neurons. This study proposes to test the following factors for their effects on precursor cell mitosis and subsequent differentiation in vitro: calcium levels; cyclic AMP; nerve growth factor; fibroblast growth factor; brain homogenates; the corticosteroids: cortisone; thyroxine; ACTH; conditioned media; and the substrate materials: polylysine; collagen; laminin. The differentiation of neuronal and glial cell types will be assessed by immunoreactivity with neuronal- and glial-specific monoclonal antibodies, and by measurements on the morphology and growth parameters of the cells in vitro. The criterion for precursor cell mitosis will be nuclear incorporation of tritiated thymidine. An initial determination of the membrane properties of the cultured precursor cells and differentiating neurons will be made by patch clamp recording. Studies examining 3H-thymidine incorporation in rostral Apteronotus cord in vivo will determine whether any low level of neurogenesis normally occurs in this morphologically mature cord which is the source of neurogenesis during regeneration in vivo and in vitro. Since regeneration is extremely limited and neurogenesis non- existent in mammalian spinal cord, the Apteronotus explant culture system provides an important new model in which the control of these processes can be examined. The ultimate goal is that a thorough definition of the processes of neurogenesis and neuronal differentiation in tissue cultures of Apteronotus spinal cord may lead to potential mechanisms for reactivating mitosis or creating a permissive environment for neuronal repair in the mammalian spinal cord.
| Anderson, M J; Rossetto, D L; Lorenz, L A (1994) Neuronal differentiation in vitro from precursor cells of regenerating spinal cord of the adult teleost Apteronotus albifrons. Cell Tissue Res 278:243-8 |
| Anderson, M J (1993) Differences in growth of neurons from normal and regenerated teleost spinal cord in vitro. In Vitro Cell Dev Biol 29A:145-52 |
