Immunoreactive insulin from mouse brain cells in culture and whole rat brain

Abstract

Fetal mouse brain cells were cultured as described previously without added insulin and without fetal calf serum after 12 days in culture. Examination by phase-contrast microscopy showed that these modifications did not appear to affect growth and development of the cells adversely. Ag impregnation of the cultures and indirect immunofluorescence following reaction with tetanus toxin showed that a high proportion of the cells resembled neurons. Analysis of concentrated culture medium by radioimmunoassay and high-pressure liquid chromatography (HPLC) revealed that the cells produced 2 main forms of immunoreactive insulin which differed from authentic pancreatic insulin in retention time. Immunoreactive somatostatin was also produced in culture and this was resolved into at least 3 forms by HPLC. Immunoreactive insulin was also extracted from whole rat brain by using 2 published procedures. The method of Havrankova, Schmechel, Roth and Brownstein consistently gave greater yields of insulin than did that of Eng and Yalow and the concentration was about three times that of plasma. The extracted insulin was further characterized by HPLC in each case and behaved like authentic pancreatic insulin. The production of insulin and somatostatin by fetal mouse brain cells in culture suggests that they may be a useful model system for studies of neuropeptide biosynthesis.