Insulin, insulin growth factor-I (IGF-I) and insulin growth factor-II (IGF-11) are known to support neuronal survival as well as stimulate cell proliferation and vigorous neuritic outgrowth. While the concept has emerged that insulin and the insulin-like growth factors are neuronotrophic factors, where in the nervous system the factors are produced and what transmembrane signal they employ remain unknown. We plan to use immunohistochemistry to determine the distribution of IGF-I and II in normal and regenerating rat sciatic nerve. Later, IGF-I and II production sites will be localized with in situ hybridization techniques. In vitro specific IGF-I and Il receptor binding sites will be determined in a model system, the SH-SY5Y neuroblastoma clone, using iodinated ligands, competitive binding curves and affinity crosslinking. IGF-I and Il mRNA will be measured in the neuroblastomas as well as a defined Schwann cell line. We believe the transmembrane signal of the insulin-like growth factors may be mediated through the activity of phosphoinositide (PI) turnover. We have preliminary results which show an IGF-II stimulation of PI turnover in the SH-SY5Y neuroblastoma cells. The kinetics of IGF-I and II stimulated formation of 3H-inositol phosphates will be measured along with a comparison between receptor occupancy and PI turnover. Intracellular calcium and protein kinase C activity will be assayed. Our long term goal is to understand how the insulin-like growth factors regulate growth in the nervous system and to bridge the gap between receptor activation, signal induction and neurite outgrowth.
