The existence of β-nerve growth factor (βNGF) in mouse cerebral cortical synaptosomal extracts was assessed using a newly developed radioimmunoasay and by bioassay. Displacement curves of labeled βNGF in the radioimmunoassay by synaptosomal extracts were parallel to the βNGF standard curves. In contrast, both mitochondrial and nonsynaptosomal extracts produced displacement curves in the βNGF radioimmunoassay which were nonparallel to standard βNGF. Intraventricular injections of purified βNGF in amounts 5 and 100 ng/mouse (six to 120 times that of whole brain βNGF concentrations) did not alter the endogenous synaptosomal βNGF concentrations. Synaptosomal extracts contained βNGF in relatively higher concentrations than the extracts of whole brain or cerebral cortex. Synaptosomal βNGF levels undergo a developmental change. The βNGF concentrations were highest in synaptosomal extracts of 12-day-old mouse cerebral cortex (the youngest age studied), and levels decreased with increasing age. Synaptosomal extracts of 12-day-old mouse cerebral cortex produced neurite outgrowth in the pheochromocytoma cell line 12 bioassay system and this response was completely inhibited by βNGF antibodies. In the pheochromocytoma cell line 12 cell bioassay, synaptosomal extracts also caused cell-cell aggregation which was unaffected by βNGF antibodies. These results provide evidence for the existence of βNGF in relatively high concentration in nerve terminals of mouse cerebral cortex.