An expression and secretion plasmid for brain-derived neurotrophic factor (BDNF) was constructed with a constitutive promoter and the Bacillus subtilis subtilisin signal sequence. For the first time, BDNF expression and secretion by recombinant B. subtilis were showed by measuring the biological activity of the medium with dorsal root ganglion neurons of 8-d chick embryos. The presence of BDNF in the medium was revalidated by the observation of lower biological activity with the BDNF antibody, but the estimated BDNF level was very low. In addition, 1.5 times higher biological activity was achieved in the glucose medium than in the glucose-free medium because of catabolite repression of protease expression resulting in lower proteolytic degradation of the product.