During the development of a primary culture of chromaffin cells from adult rat adrenals, neurite outgrowth from chromaffin cells was sometimes observed. Subsequently we collected the adrenal medullary culture media in which chromaffin cell neurite outgrowth was observed and found that it stimulated neurite out-growth from cultured adrenal chromaffin cells and also from PC-12 cells. The experimental results to date indicate that this neurite out-growth activity is a heat labile protein but is distinct from NGF, aFGF, bFGF, EGF and IGF-I. Recently, it has been reported that various protease inhibitors are capable of initiating neurite-outgrowth from various cells. Because of this, various protease inhibitors, alpha2-macroglobulin, soybean trypsin inhibitor, alpha1-antitrypsin, aprotinin, leupeptin and antipain were examined and found to exert no neurite-outgrowth activity on PC- 12 cells. Our previous experiments also indicate that adrenal medullary non-chromaffin cells may release this neurite-outgrowth activity into the culture media during culture. In this study, large quantities of adrenal medullary cells were prepared for extractions of the neurite outgrowth promoting factor(s) and some extracts were found to be active in initiating neurite-outgrowth from PC-12 cells. We plan to establish an optimal extraction condition and then to isolate this neurite-outgrowth factor(s) for further characterization.
